ORCID Profile
0000-0003-0532-8331
Current Organisation
Monash University
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Research Topics
In Research Link Australia (RLA), "Research Topics" refer to ANZSRC FOR and SEO codes. These topics are either sourced from ANZSRC FOR and SEO codes listed in researchers' related grants or generated by a large language model (LLM) based on their publications.
Top 5 Research Topics
ANZSRC Field of Research (FoR)
Biochemistry and Cell Biology | Proteomics and Intermolecular Interactions (excl. Medical Proteomics) | Structural Biology (incl. Macromolecular Modelling) | Medical Biochemistry: Proteins And Peptides | Immunology | Biological And Medical Chemistry | Tumor Immunology | Cellular Immunology | Cellular Immunology | Biochemistry And Cell Biology Not Elsewhere Classified | Medicinal and Biomolecular Chemistry | Biotechnology Not Elsewhere Classified | Proteins and Peptides | Receptors and Membrane Biology | Enzymes | Information Storage, Retrieval And Management | Autoimmunity | Medical Infection Agents (Incl. Prions) | Invertebrate Biology | Analytical Biochemistry | Medical Biochemistry: Proteins and Peptides (incl. Medical Proteomics) | Analytical Biochemistry | Bioinformatics | Animal Systematics and Taxonomy | Cell Neurochemistry
ANZSRC Socio-Economic Objective (SEO)
Expanding Knowledge in the Biological Sciences | Immune system and allergy | Cancer and related disorders | Prevention—biologicals (e.g. vaccines) | Infectious diseases | Preventive medicine | Nervous System and Disorders | Nervous system and disorders | Biological sciences | Expanding Knowledge in the Medical and Health Sciences | Expanding Knowledge in the Chemical Sciences | Immune System and Allergy | Infectious Diseases |
Related Links
Publications
The central role played by peptides in the immune response and the design of peptide-based vaccines against infectious diseases and cancer.
Publisher: Bentham Science Publishers Ltd.
Date: 04-2002
Abstract: Vaccines are one of the most cost effective methods of improving public health thereby increasing the quality of life. Prophylactic and therapeutic treatment by vaccines can prevent infectious diseases and some cancers and could also be used in the treatment of autoimmune disorders. An appreciation of this potential has resulted in a burgeoning literature which not only describes the scientific efforts being made into designing new and improved vaccines but also drives the efforts being made by public health organizations world-wide in delivering vaccines to the community. At the forefront of technologies being applied to the design of vaccines is the use of synthetic peptides the chemical technologies used to assemble peptides have made great strides over the last decade and assembly of hi-fidelity peptides which can be of high molecular weight, multimeric or even branched is now almost routine. Together with the advances in peptide technology our understanding of the molecular events that are necessary to induce immune responses has also made great strides. The central role that peptides play in immune recognition is now recognised and rules are emerging that are being applied to the construction of peptide-based vaccines that, in the right context, can induce humoral (antibody) and cellular (cytotoxic and helper T cell) immune responses. Synthetic peptides are exquisitely placed to answer questions about immune recognition and along the way to provide us with new and improved vaccines.
The Structure of HLA-B8 Complexed to an Immunodominant Viral Determinant: Peptide-Induced Conformational Changes and a Mode of MHC Class I Dimerization
Publisher: The American Association of Immunologists
Date: 11-2002
DOI: 10.4049/JIMMUNOL.169.9.5153
Abstract: EBV is a ubiquitous human pathogen that chronically infects up to 90% of the population. Persistent viral infection is characterized by latency and periods of viral replication that are kept in check by a strong antiviral CTL response. Despite the size of the EBV genome, CTL immunity focuses on only a few viral determinants but expands a large primary and memory response toward these epitopes. In unrelated HLA-B8+ in iduals, the response to the immunodominant latent Ag FLRGRAYGL from Epstein Barr nuclear Ag 3A is largely comprised of CTL clones with identical conserved αβ TCR structures. To better understand the structural correlates of Ag immunodominance and TCR selection bias, we have solved the crystal structure of the HLA-B8-FLRGRAYGL peptide complex to a resolution of 1.9 Å. The structure confirms the importance of P3-Arg, P5-Arg, and P9-Leu as dominant anchor residues involved in peptide binding to HLA-B8. A bulged conformation of the bound peptide provides a structural basis for the critical role of the P7-Tyr residue in T cell recognition. The peptide also induces backbone and side-chain conformational changes in HLA-B8 that are transmitted along the peptide-binding groove in a domino effect. The HLA-B8-FLRGRAYGL complex crystallizes as a dimer in the asymmetric unit and is oriented such that both peptide ligands are projected in the same plane suggesting a higher order arrangement of MHC-peptide complexes that could be involved in formation of the class I Ag-loading complex or in T cell activation.
Differential Roles of Plasma Protein Corona on Immune Cell Association and Cytokine Secretion of Oligomeric and Fibrillar Beta-Amyloid.
Publisher: American Chemical Society (ACS)
Date: 10-10-2019
Supplementary Figure and Table Legends from Spliced Peptides and Cytokine-Driven Changes in the Immunopeptidome of Melanoma
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/2326-6066.22542576.V1
Abstract: Supplementary figure legends and tables descriptors
The interplay between citrullination and HLA-DRB1 polymorphism in shaping peptide binding hierarchies in rheumatoid arthritis
Publisher: Elsevier BV
Date: 03-2018
Rapid microscale in-gel processing and digestion of proteins using surface acoustic waves
Publisher: Royal Society of Chemistry (RSC)
Date: 2010
DOI: 10.1039/C001501F
Abstract: A new method for in-gel s le processing and tryptic digestion of proteins is described. S le preparation, rehydration, in situ digestion and peptide extraction from gel slices are dramatically accelerated by treating the gel slice with surface acoustic waves (SAWs). Only 30 minutes total workflow time is required for this new method to produce base peak chromatograms (BPCs) of similar coverage and intensity to those observed for traditional processing and overnight digestion. Simple set up, good reproducibility, excellent peptide recoveries, rapid turnover of s les and high confidence protein identifications put this technology at the fore-front of the next generation of proteomics s le processing tools.
Human CD8
Publisher: Springer Science and Business Media LLC
Date: 18-02-2019
DOI: 10.1038/S41590-019-0320-6
Abstract: Influenza A, B and C viruses (IAV, IBV and ICV, respectively) circulate globally and infect humans, with IAV and IBV causing the most severe disease. CD8
Can α- and β-Alanine Containing Peptides Be Distinguished Based on the CID Spectra of Their Protonated Ions?
Publisher: American Chemical Society (ACS)
Date: 12-2008
DOI: 10.1016/J.JASMS.2008.09.007
Abstract: The fragmentation reactions of isomeric dipeptides containing alpha- and beta-alanine residues (alphaAla-alphaAla, alphaAla-betaAla, betaAla-alphaAla, and betaAla-betaAla) were studied using a combination of low-energy and energy resolved collision induced dissociation (CID). Each dipeptide gave a series of different fragment ions, allowing for differentiation. For ex le, peptides containing an N-terminal beta-Ala residue yield a diagnostic imine loss, while lactam ions at m/z 72 are unique to peptides containing beta-Ala residues. In addition, MS(3) experiments were performed. Structure-specific fragmentation reactions were observed for y(1) ions, which help identify the C-terminal residue. The MS(3) spectra of the b(2) ions are different suggesting they are unique for each peptide. Density functional theory (DFT) calculations predict that b(2) ions formed via a neighboring group attack by the amide are thermodynamically favored over those formed via neighboring group attack by the N-terminal amine. Finally, to gain further insight into the unique fragmentation chemistry of the peptides containing an N-terminal beta-alanine residue, the fragmentation reactions of protonated beta-Ala-NHMe were examined using a combination of experiment and DFT calculations. The relative transition-state energies involved in the four competing losses (NH(3), H(2)O, CH(3)NH(2), and CH(2)=NH) closely follow the relative abundances of these as determined via CID experiments.
Molecular chaperones are targets of autoimmunity in Ro(SS-A) immune mice
Publisher: Oxford University Press (OUP)
Date: 02-1999
DOI: 10.1046/J.1365-2249.1999.00794.X
Abstract: We have used a murine model of experimental anti-Ro(SS-A) autoimmunity to dissect additional intermolecular interactions between the 52-kD Ro (Ro52) and 60-kD Ro (Ro60) autoantigens and molecular chaperones. Immune responses to members of the heat shock protein hsp70 and hsp90 families were measured by immunoblotting and ELISA in sera from mice immunized and boosted with purified recombinant Ro52, Ro60 and La (SS-B). All Ro52 and Ro60 immune sera immunoblotted the inducible glucose-regulated protein grp78 and hsp70 species but not constitutive hsc70 or hsp90. The kinetics of antibody production and reciprocal affinity purification experiments indicated that the grp78 and hsp70 responses were cross-reactive but distinct from immune responses to the primary Ro52 and Ro60 immunogens and the endoplasmic reticulum (ER)-resident chaperone calreticulin. No responses to molecular chaperones were detected in the La-immunized mice. Control immunizations indicated that the recruited grp78 and hsp70 responses were specific for the Ro proteins and not due to immunization with denatured protein. The rapid spreading of immunity to the inducible grp78 and hsp70 in Ro52- and Ro60-immunized mice suggests that these components may co-localize and physically associate under certain physiological conditions which may promote autoimmunization. The potential importance of the ER-resident chaperones grp78 and calreticulin is further supported by their co-localization with Ro in small apoptotic membrane blebs and the finding of a novel putative grp78 binding motif in the carboxyl-terminal region of Ro52.
Repertoire of Nonclassical MHC I (HLA-E, HLA-F, HLA-G, and Orthologues)
Publisher: Elsevier
Date: 2016
Highly Divergent T-cell Receptor Binding Modes Underlie Specific Recognition of a Bulged Viral Peptide bound to a Human Leukocyte Antigen Class I Molecule
Publisher: Elsevier BV
Date: 05-2013
T-cell autoreactivity to citrullinated autoantigenic peptides in rheumatoid arthritis patients carrying HLA-DRB1 shared epitope alleles
Publisher: Springer Science and Business Media LLC
Date: 2012
DOI: 10.1186/AR3848
Understanding the Role of HLA Class I Molecules in the Immune Response to Influenza Infection and Rational Design of a Peptide-Based Vaccine
Publisher: MDPI AG
Date: 21-11-2022
DOI: 10.3390/V14112578
Abstract: Influenza A virus is a respiratory pathogen that is responsible for regular epidemics and occasional pandemics that result in substantial damage to life and the economy. The yearly reformulation of trivalent or quadrivalent flu vaccines encompassing surface glycoproteins derived from the current circulating strains of the virus does not provide sufficient cross-protection against mismatched strains. Unlike the current vaccines that elicit a predominant humoral response, vaccines that induce CD8+ T cells have demonstrated a capacity to provide cross-protection against different influenza strains, including novel influenza viruses. Immunopeptidomics, the mass spectrometric identification of human-leukocyte-antigen (HLA)-bound peptides isolated from infected cells, has recently provided key insights into viral peptides that can serve as potential T cell epitopes. The critical elements required for a strong and long-living CD8+ T cell response are related to both HLA restriction and the immunogenicity of the viral peptide. This review examines the importance of HLA and the viral immunopeptidome for the design of a universal influenza T-cell-based vaccine.
An Extensive Antigenic Footprint Underpins Immunodominant TCR Adaptability against a Hypervariable Viral Determinant
Publisher: The American Association of Immunologists
Date: 12-2014
Abstract: Mutations in T cell epitopes are implicated in hepatitis C virus (HCV) persistence and can impinge on vaccine development. We recently demonstrated a narrow bias in the human TCR repertoire targeted at an immunodominant, but highly mutable, HLA-B*0801–restricted epitope (1395HSKKKCDEL1403 [HSK]). To investigate if the narrow TCR repertoire facilitates CTL escape, structural and biophysical studies were undertaken, alongside comprehensive functional analysis of T cells targeted at the natural variants of HLA-B*0801–HSK in different HCV genotypes and quasispecies. Interestingly, within the TCR–HLA-B*0801–HSK complex, the TCR contacts all available surface-exposed residues of the HSK determinant. This broad epitope coverage facilitates cross-genotypic reactivity and recognition of common mutations reported in HCV quasispecies, albeit to a varying degree. Certain mutations did abrogate T cell reactivity however, natural variants comprising these mutations are reportedly rare and transient in nature, presumably due to fitness costs. Overall, despite a narrow bias, the TCR accommodated frequent mutations by acting like a blanket over the hypervariable epitope, thereby providing effective viral immunity. Our findings simultaneously advance the understanding of anti-HCV immunity and indicate the potential for cross-genotype HCV vaccines.
High-performance liquid chromatography of amino acids, peptides and proteins CXV. Thermodynamic behaviour of peptides in reversed-phase chromatography
Publisher: Elsevier BV
Date: 02-1992
DOI: 10.1016/0021-9673(92)80274-X
Abstract: The thermodynamic behaviour of three peptides, bombesin, beta-endorphin and glucagon, was studied under reversed-phase high-performance liquid chromatographic conditions. Experimental data related to the interactive surface contact area (S values) and solute affinity (log k0) were derived over a range of temperatures between 5 and 85 degrees C. These experimental conditions allowed changes in the secondary structure of the solute to be monitored. The influence of the nature of the stationary phase ligand on the relative conformational stability of the three peptides was analysed by acquiring data with n-octadecyl silica (C18) and n-butyl silica (C4) sorbents. Values for the relative changes in entropy and enthalpy associated with the interactive process were also determined. The results provide further insight into the factors involved with the stabilization of secondary structure and the mechanism of the interaction of peptides with hydrophobic surfaces.
A class of γδ T cell receptors recognize the underside of the antigen-presenting molecule MR1
Publisher: American Association for the Advancement of Science (AAAS)
Date: 20-12-2019
Abstract: The ligands bound by γδ T cell receptors (TCRs) are less well characterized than those of their αβ TCR cousins, which are antigens presented by major histocompatibility complex (MHC) and related proteins. Le Nours et al. identified a phenotypically erse γδ T cell subset in human tissues that reacts to MHC-related protein 1 (MR1), which presents vitamin B derivatives. A crystal structure of a γδ TCR–MR1–antigen complex revealed that some of these TCRs can bind underneath the MR1 antigen-binding cleft instead of recognizing the presented antigen. This work thus uncovers an additional ligand for γδ T cells and reconceptualizes the nature of T cell antigen recognition. Science , this issue p. 1522
Antigen Ligation Triggers a Conformational Change within the Constant Domain of the αβ T Cell Receptor
Publisher: Elsevier BV
Date: 06-2009
DOI: 10.1016/J.IMMUNI.2009.03.018
Abstract: Ligation of the alphabeta T cell receptor (TCR) by a specific peptide-loaded major histocompatibility complex (pMHC) molecule initiates T cell signaling via the CD3 complex. However, the initial events that link antigen recognition to T cell signal transduction remain unclear. Here we show, via fluorescence-based experiments and structural analyses, that MHC-restricted antigen recognition by the alphabeta TCR results in a specific conformational change confined to the A-B loop within the alpha chain of the constant domain (Calpha). The apparent affinity constant of this A-B loop movement mirrored that of alphabeta TCR-pMHC ligation and was observed in two alphabeta TCRs with distinct pMHC specificities. The Ag-induced A-B loop conformational change could be inhibited by fixing the juxtapositioning of the constant domains and was shown to be reversible upon pMHC disassociation. Notably, the loop movement within the Calpha domain, although specific for an agonist pMHC ligand, was not observed with a pMHC antagonist. Moreover, mutagenesis of residues within the A-B loop impaired T cell signaling in an in vitro system of antigen-specific TCR stimulation. Collectively, our findings provide a basis for the earliest molecular events that underlie Ag-induced T cell triggering.
Novel tumor derived peptides in vaccines
Publisher: Elsevier
Date: 2013
Human leukocyte antigen-associated drug hypersensitivity
Publisher: Elsevier BV
Date: 02-2013
DOI: 10.1016/J.COI.2012.10.002
Abstract: A growing number of associations between adverse drug reactions and alleles of the human leukocyte antigen (HLA) genes are now known. Although several models have been proposed to explain these associations, an underlying molecular basis has only recently been described. The associations between HLA-B*57:01 and abacavir hypersensitivity syndrome, and HLA-B*15:02 and carbamazepine-induced bullous skin disease have provided new insights into the mechanism associated with hypersensitivity reactions to these drugs. Here we discuss recent evidence that small molecules can interact with specific HLA to distort self-peptide presentation leading to autoimmune-like drug hypersensitivities that potentially provide clues to the mechanisms underlying other immunopathologies.
Immunodominance hierarchies and gender bias in direct TCD8 -cell alloreactivity
Publisher: Elsevier BV
Date: 2008
DOI: 10.1111/J.1600-6143.2007.02044.X
Abstract: Allogeneic solid organ transplantation often occurs across multiple donor-recipient HLA mismatches with consequent risk of allograft rejection. However, there is growing evidence that not all HLA mismatches are equivalent in their stimulation of allogeneic T cells making it important to determine which of these might be more significant as predictors of allograft rejection. To this end, we used defined antigen-presenting cell (APC) transfectants expressing single MHC-I allotypes as target cells that could discriminate the relative contribution of in idual mismatched MHC-I allotypes to direct T-cell alloreactivity. We demonstrate remarkably reproducible patterns of immunodominance in reactivity across mismatched MHC-I allotypes. These patterns are HLA context-dependent, partly reflecting alloantigenic competition in responder cell responses. In strong alloresponses, we also observed an increased percentage of alloreactive T(CD8) cells in female responders, regardless of the stimulator gender, highlighting HLA-independent factors in the potency of the alloresponse. This approach provides a potential measure of specific alloreactive T cells that could be used in clinical practice for selection of donors, assessment of posttransplant outcomes, modulation of immunosuppression and detection of rejection episodes.
Specialisation of the venom gland proteome in predatory cone snails reveals functional diversification of the conotoxin biosynthetic pathway.
Publisher: American Chemical Society (ACS)
Date: 11-08-2011
DOI: 10.1021/PR1012976
Abstract: Conotoxins, venom peptides from marine cone snails, ersify rapidly as speciation occurs. It has been suggested that each species can synthesize between 1000 and 1900 different toxins with little to no interspecies overlap. Conotoxins exhibit an unprecedented degree of post-translational modifications, the most common one being the formation of disulfide bonds. Despite the great ersity of structurally complex peptides, little is known about the glandular proteins responsible for their biosynthesis and maturation. Here, proteomic interrogations on the Conus venom gland led to the identification of novel glandular proteins of potential importance for toxin synthesis and secretion. A total of 161 and 157 proteins and protein isoforms were identified in the venom glands of Conus novaehollandiae and Conus victoriae, respectively. Interspecies differences in the venom gland proteomes were apparent. A large proportion of the proteins identified function in protein eptide translation, folding, and protection events. Most intriguingly, however, we demonstrate the presence of a multitude of isoforms of protein disulfide isomerase (PDI), the enzyme catalyzing the formation and isomerization of the native disulfide bond. Investigating whether different PDI isoforms interact with distinct toxin families will greatly advance our knowledge on the generation of cone snail toxins and disulfide-rich peptides in general.
Transcriptional signature in microglia associated with Aβ plaque phagocytosis
Publisher: Springer Science and Business Media LLC
Date: 21-05-2021
DOI: 10.1038/S41467-021-23111-1
Abstract: The role of microglia cells in Alzheimer’s disease (AD) is well recognized, however their molecular and functional ersity remain unclear. Here, we isolated amyloid plaque-containing (using labelling with methoxy-XO4, XO4 + ) and non-containing (XO4 − ) microglia from an AD mouse model. Transcriptomics analysis identified different transcriptional trajectories in ageing and AD mice. XO4 + microglial transcriptomes demonstrated dysregulated expression of genes associated with late onset AD. We further showed that the transcriptional program associated with XO4 + microglia from mice is present in a subset of human microglia isolated from brains of in iduals with AD. XO4 − microglia displayed transcriptional signatures associated with accelerated ageing and contained more intracellular post-synaptic material than XO4 + microglia, despite reduced active synaptosome phagocytosis. We identified HIF1α as potentially regulating synaptosome phagocytosis in vitro using primary human microglia, and BV2 mouse microglial cells. Together, these findings provide insight into molecular mechanisms underpinning the functional ersity of microglia in AD.
The molecular basis of cross-reactivity in the Australian Snake Venom Detection Kit (SVDK)
Publisher: Elsevier BV
Date: 12-2007
DOI: 10.1016/J.TOXICON.2007.07.023
Abstract: The Snake Venom Detection Kit (SVDK) is of major medical importance in Australia, yet it has never been rigorously characterised in terms of its sensitivity and specificity, especially when it comes to reports of false-negative and false-positive results. This study investigates reactions and cross-reactions of five venoms the SVDK is directed against and a number of purified toxins. Snakes showing the closest evolutionary relationships demonstrated the lowest level of cross-reactivity between groups. This was, instead, far more evident between snakes that are extraordinarily evolutionary separated. These snakes: Pseudechis australis, Acanthophis antarcticus and Notechis scutatus, in fact displayed more false-positive results. Examination of in idual toxin groups showed that phospholipase A(2)s (PLA(2)s) tends to react strongly and display considerable cross-reactivity across groups while the three-finger toxins (3FTx) reacted poorly in all but the Acanthophis well. The hook effect was evident for all venoms, particularly Oxyuranus scutellatus. The results of this study show considerable variation in toxin detection, with implications in further development of venom detection, both in Australia and other countries.
Supplementary Table S3 from Spliced Peptides and Cytokine-Driven Changes in the Immunopeptidome of Melanoma
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/2326-6066.22542558.V1
Abstract: Table S3
The structural basis for autonomous dimerization of the pre-T-cell antigen receptor.
Publisher: Springer Science and Business Media LLC
Date: 10-2010
DOI: 10.1038/NATURE09448
Abstract: The pre-T-cell antigen receptor (pre-TCR), expressed by immature thymocytes, has a pivotal role in early T-cell development, including TCR β-selection, survival and proliferation of CD4(-)CD8(-) double-negative thymocytes, and subsequent αβ T-cell lineage differentiation. Whereas αβTCR ligation by the peptide-loaded major histocompatibility complex initiates T-cell signalling, pre-TCR-induced signalling occurs by means of a ligand-independent dimerization event. The pre-TCR comprises an invariant α-chain (pre-Tα) that pairs with any TCR β-chain (TCRβ) following successful TCR β-gene rearrangement. Here we provide the basis of pre-Tα-TCRβ assembly and pre-TCR dimerization. The pre-Tα chain comprised a single immunoglobulin-like domain that is structurally distinct from the constant (C) domain of the TCR α-chain nevertheless, the mode of association between pre-Tα and TCRβ mirrored that mediated by the Cα-Cβ domains of the αβTCR. The pre-TCR had a propensity to dimerize in solution, and the molecular envelope of the pre-TCR dimer correlated well with the observed head-to-tail pre-TCR dimer. This mode of pre-TCR dimerization enabled the pre-Tα domain to interact with the variable (V) β domain through residues that are highly conserved across the Vβ and joining (J) β gene families, thus mimicking the interactions at the core of the αβTCR's Vα-Vβ interface. Disruption of this pre-Tα-Vβ dimer interface abrogated pre-TCR dimerization in solution and impaired pre-TCR expression on the cell surface. Accordingly, we provide a mechanism of pre-TCR self-association that allows the pre-Tα chain to simultaneously 's le' the correct folding of both the V and C domains of any TCR β-chain, regardless of its ultimate specificity, which represents a critical checkpoint in T-cell development. This unusual dual-chaperone-like sensing function of pre-Tα represents a unique mechanism in nature whereby developmental quality control regulates the expression and signalling of an integral membrane receptor complex.
Identification of native and posttranslationally modified HLA-B*57:01- restricted HIV envelope derived epitopes using immunoproteomics
Publisher: Wiley
Date: 10-04-2018
Abstract: The recognition of pathogen-derived peptides by T lymphocytes is the cornerstone of adaptive immunity, whereby intracellular antigens are degraded in the cytosol and short peptides assemble with class I human leukocyte antigen (HLA) molecules in the ER. These peptide-HLA complexes egress to the cell surface and are scrutinized by cytotoxic CD8+ T-cells leading to the eradication of the infected cell. Here, naturally presented HLA-B*57:01 bound peptides derived from the envelope protein of the human immunodeficiency virus (HIVenv) are identified. HIVenv peptides are present at a very small percentage of the overall HLA-B*57:01 peptidome (<0.1%) and both native and posttranslationally modified forms of two distinct HIV peptides are identified. Notably, a peptide bearing a natively encoded C-terminal tryptophan residue is also present in a modified form containing a kynurenine residue. Kynurenine is a major product of tryptophan catabolism and is abundant during inflammation and infection. Binding of these peptides at a molecular level and their immunogenicity in preliminary functional studies are examined. Modest immune responses are observed to the modified HIVenv peptide, highlighting a potential role for kynurenine-modified peptides in the immune response to HIV and other viral infections.
Characterization of the Antigen Processing Machinery and Endogenous Peptide Presentation of a Bat MHC Class I Molecule
Publisher: The American Association of Immunologists
Date: 06-2016
Abstract: Bats are a major reservoir of emerging and re-emerging infectious diseases, including severe acute respiratory syndrome–like coronaviruses, henipaviruses, and Ebola virus. Although highly pathogenic to their spillover hosts, bats harbor these viruses, and a large number of other viruses, with little or no clinical signs of disease. How bats asymptomatically coexist with these viruses is unknown. In particular, little is known about bat adaptive immunity, and the presence of functional MHC molecules is mostly inferred from recently described genomes. In this study, we used an affinity purification/mass spectrometry approach to demonstrate that a bat MHC class I molecule, Ptal-N*01:01, binds antigenic peptides and associates with peptide-loading complex components. We identified several bat MHC class I–binding partners, including calnexin, calreticulin, protein disulfide isomerase A3, tapasin, TAP1, and TAP2. Additionally, endogenous peptide ligands isolated from Ptal-N*01:01 displayed a relatively broad length distribution and an unusual preference for a C-terminal proline residue. Finally, we demonstrate that this preference for C-terminal proline residues was observed in Hendra virus–derived peptides presented by Ptal-N*01:01 on the surface of infected cells. To our knowledge, this is the first study to identify endogenous and viral MHC class I ligands for any bat species and, as such, provides an important avenue for monitoring and development of vaccines against major bat-borne viruses both in the reservoir and spillover hosts. Additionally, it will provide a foundation to understand the role of adaptive immunity in bat antiviral responses.
Family presence during resuscitation (FPDR): A qualitative descriptive study exploring the experiences of emergency personnel post resuscitation
Publisher: Elsevier BV
Date: 07-2019
DOI: 10.1016/J.HRTLNG.2018.09.016
Abstract: Family presence during resuscitation (FPDR), remains inconsistently implemented by emergency personnel. The benefits for family members is well documented, providing opportunities for family to say goodbye, facilitates closure and enables family to provide emotional support to the patient. The aim of this study was to explore the experiences and attitudes of emergency personnel towards FPDR immediately post resuscitation events. A descriptive qualitative design was used to explore the experiences of emergency personnel with FPDR. Data was collected from single rural and metropolitan emergency departments in the state of Victoria, Australia. The participants consisted of nurses and doctors who took active roles during resuscitation events. Following transcription of the audiotaped interviews Creswell's (2003) six step analysis process was employed. A total of 29 interviews of key personnel, following 6 paediatric and 18 adult resuscitation events. Interviews were conducted over a period of two weeks in each venue. The data was organised into six themes following analysis including: care coordinators inconsistently called, gate keepers to implementation, effective communication strategies helping to deliver bad news, life experience generates confidence, allocation of family support person, and family members roles dependent on age of patient. FPDR is common practice in paediatric events however remains inconsistently implemented during adult resuscitations. A designated family support person is essential to successful implementation of FPDR and should be incorporated in to the allocation of the resuscitation team roles during both adult and paediatric resuscitation events. Education and training is important for clinicians to learn essential communication skills, building practice confidence, which is required to successfully implement FPDR.
Tapasin-Mediated Retention and Optimization of Peptide Ligands During the Assembly of Class I Molecules
Publisher: The American Association of Immunologists
Date: 07-2000
DOI: 10.4049/JIMMUNOL.165.1.322
Abstract: The murine class I H-2Kb molecule achieves high level surface expression in tapasin-deficient 721.220 human cells. Compared with their behavior in wild-type cells, Kb molecules expressed on 721.220 cells are more receptive to exogenous peptide, undergo more rapid surface decay, and fail to form macromolecular peptide loading complexes. As a result, they are rapidly transported to the cell surface, reflecting a failure of endoplasmic reticulum retention mechanisms in the absence of loading complex formation. Despite the failure of Kb molecules to colocalize to the TAP and their rapid egress to the cell surface, Kb is still capable of presenting TAP-dependent peptides in the absence of tapasin. Furthermore, pool sequencing of peptides eluted from these molecules revealed strict conservation of their canonical H-2Kb-binding motif. There was a reduction in the total recovery of peptides associated with Kb molecules purified from the surface of tapasin-deficient cells. Comparison of the peptides bound to Kb in the presence and absence of tapasin revealed considerable overlap in peptide repertoire. These results indicate that in the absence of an interaction with tapasin, Kb molecules fail to assemble with calreticulin and TAP, yet they are still capable of acquiring a erse array of peptides. However, a significant proportion of these peptides appear to be suboptimal, resulting in reduced cell surface stability of Kb complexes. Taken together, the findings indicate that tapasin plays an essential role in the formation of the class I loading complex, which retains class I heterodimers in the endoplasmic reticulum until optimal ligand selection is completed.
Synergistic Combination of Polymyxin B and Enrofloxacin Induced Metabolic Perturbations in Extensive Drug-Resistant Pseudomonas aeruginosa
Publisher: Frontiers Media SA
Date: 03-10-2019
Tentacle Transcriptomes of the Speckled Anemone (Actiniaria: Actiniidae: Oulactis sp.): Venom-Related Components and Their Domain Structure
Publisher: Springer Science and Business Media LLC
Date: 24-01-2020
DOI: 10.1007/S10126-020-09945-8
Abstract: Cnidarians are one of the oldest known animal lineages (ca. 700 million years), with a unique envenomation apparatus to deliver a potent mixture of peptides and proteins. Some peptide toxins from cnidarian venom have proven therapeutic potential. Here, we use a transcriptomic roteomic strategy to identify sequences with similarity to known venom protein families in the tentacles of the endemic Australian 'speckled anemone' (Oulactis sp.). Illumina RNASeq data were assembled de novo. Annotated sequences in the library were verified by cross-referencing in iduals' transcriptomes or protein expression evidence from LC-MS/MS data. Sequences include pore-forming toxins, phospholipases, peptidases, neurotoxins (sodium and potassium channel modulators), cysteine-rich secretory proteins and defensins (antimicrobial peptides). Fewer than 4% of the sequences in the library occurred across the three in iduals examined, demonstrating high sequence variability of an in idual's arsenal. We searched for actinoporins in Oulactis sp. to assess sequence similarity to the only described toxins (OR-A and -G) for this genus and examined the domain architecture of venom-related peptides and proteins. The novel putative actinoporin of Oulactis sp. has a greater similarity to other species in the Actiniidae family than to O. orientalis. Venom-related sequences have an architecture that occurs in single, repeat or multi-domain combinations of venom-related (e.g. ShK-like) and non-venom (e.g. whey acid protein) domains. This study has produced the first transcriptomes for an endemic Australian sea anemone species and the genus Oulactis, while identifying nearly 400 novel venom-related peptides and proteins for future structural and functional analyses and venom evolution studies.
The Cellular Redox Environment Alters Antigen Presentation
Publisher: Elsevier BV
Date: 10-2014
Modulation of conotoxin structure and function is achieved through a multienzyme complex in the venom glands of cone snails
Publisher: Elsevier BV
Date: 10-2012
Spliced HLA-bound peptides: a Black Swan event in immunology
Publisher: Oxford University Press (OUP)
Date: 28-03-2021
DOI: 10.1111/CEI.13589
Abstract: Peptides that bind to and are presented on the cell surface by human leucocyte antigen (HLA) molecules play a critical role in adaptive immunity. For a long time it was believed that all the HLA-bound peptides were generated through simple proteolysis of linear sequences of cellular proteins, and therefore are templated in the genome and proteome. However, evidence for untemplated peptide ligands of HLA molecules has accumulated during the last two decades, with a recent global analysis of HLA-bound peptides suggesting that a considerable proportion of HLA-bound peptides are potentially generated through splicing/fusion of discontinuous peptide segments from one or two distinct proteins. In this review, we will evaluate recent discoveries and debates on the contribution of spliced peptides to the HLA class I immunopeptidome, consider biochemical rules for splicing and the potential role of these spliced peptides in immune recognition.
Modification of the cyclopropyl moiety of abacavir provides insight into the structure activity relationship between HLA‐B*57:01 binding and T‐cell activation
Publisher: Wiley
Date: 09-10-2019
DOI: 10.1111/ALL.14057
Abstract: Abacavir is associated with hypersensitivity reactions in in iduals positive for the HLA‐B*57:01 allele. The drug binds within the peptide binding groove of HLA‐B*57:01 altering peptides displayed on the cell surface. Presentation of these HLA‐abacavir‐peptide complexes to T‐cells is hypothesized to trigger a CD8 + T‐cell response underpinning the hypersensitivity. Thus, the aim of this study was to explore the relationship between the structure of abacavir with HLA‐B*57:01 binding and the CD8 + T‐cell activation. Seventeen abacavir analogues were synthesized and cytokine secretion from abacavir/abacavir analogue‐responsive CD8 + T‐cell clones was measured using IFN‐γ ELIspot. In silico docking studies were undertaken to assess the predicted binding poses of the abacavir analogues within the HLA‐B*57:01 peptide binding groove. In parallel, the effect of selected abacavir analogues on the repertoire of self‐peptides presented by cellular HLA‐B*57:01 was characterized using mass spectrometry. Abacavir and ten analogues stimulated CD8 + T‐cell IFN‐γ release. Molecular docking of analogues that retained antiviral activity demonstrated a relationship between predicted HLA‐B*57:01 binding orientations and the ability to induce a T‐cell response. Analogues that stimulated T‐cells displayed a perturbation of the natural peptides displayed by HLA‐B*57:01. The antigen‐specific CD8 + T‐cell response was dependent on the enantiomeric form of abacavir at both cyclopropyl and cyclopentyl regions. Alteration of the chemical constitution of abacavir generates analogues that retain a degree of pharmacological activity, but have variable ability to activate T‐cells. Modelling and immunopeptidome analysis delineate how drug HLA‐B*57:01 binding and peptide display by antigen presenting cells relate to the activation of CD8 + T‐cells.
MhcVizPipe: A Quality Control Software for Rapid Assessment of Small- to Large-Scale Immunopeptidome Datasets
Publisher: Elsevier BV
Date: 2022
Short Duration Alagebrium Chloride Therapy Prediabetes Does Not Inhibit Progression to Autoimmune Diabetes in an Experimental Model
Publisher: MDPI AG
Date: 28-06-2021
Abstract: Mechanisms by which advanced glycation end products (AGEs) contribute to type 1 diabetes (T1D) pathogenesis are poorly understood. Since life-long pharmacotherapy with alagebrium chloride (ALT) slows progression to experimental T1D, we hypothesized that acute ALT therapy delivered prediabetes, may be effective. However, in female, non-obese diabetic (NODShiLt) mice, ALT administered prediabetes (day 50–100) did not protect against experimental T1D. ALT did not decrease circulating AGEs or their precursors. Despite this, pancreatic β-cell function was improved, and insulitis and pancreatic CD45.1+ cell infiltration was reduced. Lymphoid tissues were unaffected. ALT pre-treatment, prior to transfer of primed GC98 CD8+ T cell receptor transgenic T cells, reduced blood glucose concentrations and delayed diabetes, suggesting islet effects rather than immune modulation by ALT. Indeed, ALT did not reduce interferon-γ production by leukocytes from ovalbumin-pre-immunised NODShiLt mice and NODscid recipients given diabetogenic ALT treated NOD splenocytes were not protected against T1D. To elucidate β-cell effects, NOD-derived MIN6N8 β-cell major histocompatibility complex (MHC) Class Ia surface antigens were examined using immunopeptidomics. Overall, no major changes in the immunopeptidome were observed during the various treatments with all peptides exhibiting allele specific consensus binding motifs. As expected, longer MHC Class Ia peptides were captured bound to H-2Db than H-2Kb under all conditions. Moreover, more 10–12 mer peptides were isolated from H-2Db after AGE modified bovine serum albumin (AGE-BSA) treatment, compared with bovine serum albumin (BSA) or AGE-BSA+ALT treatment. Proteomics of MIN6N8 cells showed enrichment of processes associated with catabolism, the immune system, cell cycling and presynaptic endocytosis with AGE-BSA compared with BSA treatments. These data show that short-term ALT intervention, given prediabetes, does not arrest experimental T1D but transiently impacts β-cell function.
Secreted HLA recapitulates the immunopeptidome and allows in-depth coverage of HLA A*02:01 ligands.
Publisher: Elsevier BV
Date: 06-2012
DOI: 10.1016/J.MOLIMM.2012.02.117
Abstract: HLA molecules are cell-surface glycoproteins that present peptides, derived from intracellular protein antigens, for surveillance by T lymphocytes. Secreted HLA (sHLA) technology is a powerful approach for studying these peptides, since it facilitates large-scale production of HLA-bound peptides. We compared secreted and membrane-bound forms of HLA A2 in terms of intracellular trafficking and their bound peptide repertoire (termed the immunopeptidome). We demonstrate that sHLA and membrane bound HLA (mHLA) negotiate intracellular compartments with similar maturation kinetics. Moreover, mass spectrometry revealed a substantial overlap in the immunopeptidome was observed when HLA A2-bound peptides were purified from various sources of sHLA and mHLA. By combining machine based algorithms with manual validation, we identified 1266 non-redundant peptides. Analysis of these peptides revealed a number bearing post-translational modifications, although some of these may arise spontaneously others represent modifications performed within the cell that survive antigen processing. Peptides bearing some of these modifications have not previously been described for HLA ligands, therefore, this compendium of 1266 non-redundant peptide sequences adds greatly to the existing database of HLA A2 ligands. Peptides from all sources displayed comparable HLA A2 consensus binding motifs, peptide lengths, predicted HLA A2 binding affinities and putative source antigens. We conclude that sHLA is a valid and useful technique for studying the immunopeptidome.
The use of post-source decay in matrix-assisted laser desorption/ionisation mass spectrometry to delineate T cell determinants
Publisher: Elsevier BV
Date: 03-2001
DOI: 10.1016/S0022-1759(00)00361-6
Abstract: The identification of naturally processed peptides presented by molecules of the major histocompatibility complex (MHC) has progressed significantly over the past decade. The elution of peptides from immunoaffinity purified complexes of MHC class I or class II molecules has provided highly specific biochemical information regarding the nature of endogenous peptides capable of binding to and being presented by particular MHC alleles. Whilst Edman chemistry is sufficient for the identification of abundant or homogeneous immunodominant peptides contained in s les of fractionated peptides, mass spectrometry has proved more powerful for sequencing less abundant species present in the typically heterogeneous fractions of eluted peptides. This review focuses on the characterisation of T cell determinants by matrix-assisted laser desorption/ionisation (MALDI)-time-of-flight (TOF) mass spectrometry (MS). We demonstrate, with specific ex les, the utility of post-source decay in MALDI-TOF MS for the characterisation of the amino acid sequences of both native and modified T cell determinants. The potential advantages and pitfalls of this technique relative to the more commonly used forms of tandem mass spectrometry in electrospray and ion spray modes of ionisation as well as hybrid quadrupole-quadrupole-TOF instruments are discussed. We highlight the complementarity between these techniques and discuss the advantages in the combined use of both MALDI- and electrospray-based instrumentation in epitope identification strategies.
Overlapping Peptides Elicit Distinct CD8 T Cell Responses following Influenza A Virus Infection
Publisher: The American Association of Immunologists
Date: 10-2020
Abstract: The presentation of pathogen-derived peptides on MHC class I molecules is essential for the initiation of adaptive CD8+ T cell immunity, which in turn is critical for effective control of many significant human infections. The identification of immunogenic pathogen-derived epitopes and a detailed understanding of how they are recognized by TCRs is essential for the design of effective T cell–based vaccines. In this study, we have characterized the T cell recognition and immune responses in mice to two naturally presented influenza A virus–derived peptides previously identified from virally infected cells via mass spectrometry. These neuraminidase-derived peptides, NA181–190 (SGPDNGAVAV) and NA181–191 (SGPDNGAVAVL), are completely overlapping with the exception of a 1 aa extension at the C terminus of the longer peptide. This minor peptidic difference results in the induction of two completely independent and non–cross-reactive T cell populations that show distinct functional characteristics after influenza A virus infection of B6 mice. We show that the unique TCR reactivity to the overlapping peptides is present in the naive repertoire prior to immune expansion in B6 mice. Moreover, we provide a structural explanation underlying the distinct CD8+ T cell reactivities, which reinforces the concept that peptide length is a key determinant of Ag specificity in CD8+ T cell responses.
MHC-I peptides get out of the groove and enable a novel mechanism of HIV-1 escape
Publisher: Springer Science and Business Media LLC
Date: 20-02-2017
DOI: 10.1038/NSMB.3381
Supplementary Table S1 from Spliced Peptides and Cytokine-Driven Changes in the Immunopeptidome of Melanoma
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/2326-6066.22542564
Abstract: Table S1
An open-source computational and data resource to analyze digital maps of immunopeptidomes
Publisher: eLife Sciences Publications, Ltd
Date: 08-07-2015
DOI: 10.7554/ELIFE.07661
Abstract: We present a novel mass spectrometry-based high-throughput workflow and an open-source computational and data resource to reproducibly identify and quantify HLA-associated peptides. Collectively, the resources support the generation of HLA allele-specific peptide assay libraries consisting of consensus fragment ion spectra, and the analysis of quantitative digital maps of HLA peptidomes generated from a range of biological sources by SWATH mass spectrometry (MS). This study represents the first community-based effort to develop a robust platform for the reproducible and quantitative measurement of the entire repertoire of peptides presented by HLA molecules, an essential step towards the design of efficient immunotherapies.
The Use of Imaging Mass Spectrometry to Study Peptide Toxin Distribution in Australian Sea Anemones
Publisher: CSIRO Publishing
Date: 2017
DOI: 10.1071/CH17228
A Structural Basis for the Selection of Dominant αβ T Cell Receptors in Antiviral Immunity
Publisher: Elsevier BV
Date: 2003
DOI: 10.1016/S1074-7613(02)00513-7
Abstract: We have examined the basis for immunodominant or "public" TCR usage in an antiviral CTL response. Residues encoded by each of the highly selected genetic elements of an immunodominant clonotype recognizing Epstein-Barr virus were critical to the antigen specificity of the receptor. Upon recognizing antigen, the immunodominant TCR undergoes extensive conformational changes in the complementarity determining regions (CDRs), including the disruption of the canonical structures of the germline-encoded CDR1alpha and CDR2alpha loops to produce an enhanced fit with the HLA-peptide complex. TCR ligation induces conformational changes in the TCRalpha constant domain thought to form part of the docking site for CD3epsilon. These findings indicate that TCR immunodominance is associated with structural properties conferring receptor specificity and suggest a novel structural link between TCR ligation and intracellular signaling.
Combined proteomic and transcriptomic interrogation of the venom gland of Conus geographus uncovers novel components and functional compartmentalization.
Publisher: Elsevier BV
Date: 04-2014
Structural and regulatory diversity shape HLA-C protein expression levels
Publisher: Springer Science and Business Media LLC
Date: 26-06-2017
DOI: 10.1038/NCOMMS15924
Abstract: Expression of HLA-C varies widely across in iduals in an allele-specific manner. This variation in expression can influence efficacy of the immune response, as shown for infectious and autoimmune diseases. MicroRNA binding partially influences differential HLA-C expression, but the additional contributing factors have remained undetermined. Here we use functional and structural analyses to demonstrate that HLA-C expression is modulated not just at the RNA level, but also at the protein level. Specifically, we show that variation in exons 2 and 3, which encode the α1/α2 domains, drives differential expression of HLA-C allomorphs at the cell surface by influencing the structure of the peptide-binding cleft and the ersity of peptides bound by the HLA-C molecules. Together with a phylogenetic analysis, these results highlight the ersity and long-term balancing selection of regulatory factors that modulate HLA-C expression.
Reply to Roudier et al.: HLA-DRB1 polymorphism, anti-citrullinated protein antibodies, and rheumatoid arthritis
Publisher: Elsevier BV
Date: 05-2018
Supplementary Table S2 from Spliced Peptides and Cytokine-Driven Changes in the Immunopeptidome of Melanoma
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/2326-6066.22542561
Abstract: Table S2
Stable isotope shifted matrices enable the use of low mass ion precursor scanning for targeted metabolite identification
Publisher: Springer Science and Business Media LLC
Date: 17-01-2011
Abstract: We describe a method to identify metabolites of proteins that eliminates endogenous background by using stable isotope labeled matrices. This technique allows selective screening of the intact therapeutic molecule and all metabolites using a modified precursor ion scan that monitors low molecular weight fragment ions produced during MS/MS. This distinct set of low mass ions differs between isotopically labeled and natural isotope containing species allowing excellent discrimination between endogenous compounds and target analytes. All compounds containing amino acids that consist of naturally abundant isotopes can be selected using this scanning technique for further analysis, including metabolites of the parent molecule. The sensitivity and selectivity of this technique is discussed with specific ex les of insulin metabolites identified within a complex matrix using a range of different validated low mass target ions.
Lack of prominent peptide-major histocompatibility complex features limits repertoire diversity in virus-specific CD8+ T cell populations.
Publisher: Springer Science and Business Media LLC
Date: 27-02-2005
DOI: 10.1038/NI1175
Abstract: Using both 'reverse genetics' and structural analysis, we have examined the in vivo relationship between antigenicity and T cell receptor (TCR) repertoire ersity. Influenza A virus infection of C57BL/6 mice induces profoundly different TCR repertoires specific for the nucleoprotein peptide of amino acids 366-374 (NP366) and the acid polymerase peptide of amino acids 224-233 (PA224) presented by H-2D(b). Here we show the H-2D(b)-NP366 complex with a 'featureless' structure selected a limited TCR repertoire characterized by 'public' TCR usage. In contrast, the prominent H-2D(b)-PA224 complex selected erse, in idually 'private' TCR repertoires. Substitution of the arginine at position 7 of PA224 with an alanine reduced the accessible side chains of the epitope. Infection with an engineered virus containing a mutation at the site encoding the exposed arginine at position 7 of PA224 selected a restricted TCR repertoire similar in ersity to that of the H-2D(b)-NP366-specific response. Thus, the lack of prominent features in an antigenic complex of peptide and major histocompatibility complex class I is associated with a diminished spectrum of TCR usage.
Natural HLA class I polymorphism controls the pathway of antigen presentation and susceptibility to viral evasion
Publisher: Rockefeller University Press
Date: 28-06-2004
DOI: 10.1084/JEM.20031680
Abstract: HLA class I polymorphism creates ersity in epitope specificity and T cell repertoire. We show that HLA polymorphism also controls the choice of Ag presentation pathway. A single amino acid polymorphism that distinguishes HLA-B*4402 (Asp116) from B*4405 (Tyr116) permits B*4405 to constitutively acquire peptides without any detectable incorporation into the transporter associated with Ag presentation (TAP)-associated peptide loading complex even under conditions of extreme peptide starvation. This mode of peptide capture is less susceptible to viral interference than the conventional loading pathway used by HLA-B*4402 that involves assembly of class I molecules within the peptide loading complex. Thus, B*4402 and B*4405 are at opposite extremes of a natural spectrum in HLA class I dependence on the PLC for Ag presentation. These findings unveil a new layer of MHC polymorphism that affects the generic pathway of Ag loading, revealing an unsuspected evolutionary trade-off in selection for optimal HLA class I loading versus effective pathogen evasion.
Benchmarking Bioinformatics Pipelines in Data-Independent Acquisition Mass Spectrometry for Immunopeptidomics
Publisher: Elsevier BV
Date: 04-2023
Enhancing tumor vaccines
Publisher: Informa UK Limited
Date: 02-2010
DOI: 10.1586/ERV.09.159
Abstract: Peptide vaccination strategies aimed at inducing CD4(+) T-cell responses may be h ered due to the poorly receptive nature of MHC class II molecules to exogenous antigen. It has recently been reported that the small organic molecule adamantane ethanol, when included as an adjuvant in peptide vaccination, is capable of enhancing ligand exchange and markedly augmenting the subsequent antigen-specific CD4(+) T-cell response. These results highlight a novel adjuvant strategy tested in vivo, which opens a further doorway to improving the efficacy of peptide vaccination and continuing the push towards clinical trials.
The use of proteomics to understand antiviral immunity
Publisher: Elsevier BV
Date: 12-2018
DOI: 10.1016/J.SEMCDB.2017.12.002
Abstract: Viruses are intracellular pathogens that cause a vast array of diseases, which are often severe and typified by high morbidity and mortality rates. Viral infections continue to be a global health burden and effective vaccines and therapeutics are constantly sought to prevent and treat these infections. The development of such treatments generally relies on understanding the mechanisms that underpin efficient host antiviral immune responses. This review summarises recent developments in our understanding of antiviral adaptive immunity and in particular, highlights the use of mass spectrometry to elucidate viral antigens and their processing and presentation to T cells and other immune effectors. These processed peptides serve as potential vaccine candidates or may facilitate clinical monitoring, diagnosis and immunotherapy of infectious diseases.
Diamond integrated quantum photonics
Publisher: Elsevier BV
Date: 09-2008
Comparative Metabolomics and Transcriptomics Reveal Multiple Pathways Associated with Polymyxin Killing in Pseudomonas aeruginosa
Publisher: American Society for Microbiology
Date: 26-02-2019
DOI: 10.1128/MSYSTEMS.00149-18
Abstract: Pseudomonas aeruginosa has been highlighted by the recent WHO Global Priority Pathogen List due to multidrug resistance. Without new antibiotics, polymyxins remain a last-line therapeutic option for this difficult-to-treat pathogen. The emergence of polymyxin resistance highlights the growing threat to our already very limited antibiotic armamentarium and the urgency to understand the exact mechanisms of polymyxin activity and resistance. Integration of the correlative metabolomics and transcriptomics results in the present study discovered that polymyxin treatment caused significant perturbations in the biosynthesis of lipids, lipopolysaccharide, and peptidoglycan, central carbon metabolism, and oxidative stress. Importantly, lipid A modifications were surprisingly rapid in response to polymyxin treatment at clinically relevant concentrations. This is the first study to reveal the dynamics of polymyxin-induced cellular responses at the systems level, which highlights that combination therapy should be considered to minimize resistance to the last-line polymyxins. The results also provide much-needed mechanistic information which potentially benefits the discovery of new-generation polymyxins.
A Systems Approach to Understand Antigen Presentation and the Immune Response
Publisher: Springer New York
Date: 24-12-2015
DOI: 10.1007/978-1-4939-3341-9_14
Abstract: The mammalian immune system has evolved to respond to pathogenic, environmental, and cellular changes in order to maintain the health of the host. These responses include the comparatively primitive innate immune response, which represents a rapid and relatively nonspecific reaction to challenge by pathogens and the more complex cellular adaptive immune response. This adaptive response evolves with the pathogenic challenge, involves the cross talk of several cell types, and is highly specific to the pathogen due to the liberation of peptide antigens and their presentation on the surface of affected cells. Together these two forms of immunity provide a surveillance mechanism for the system-wide scrutiny of cellular function, environment, and health. As such the immune system is best understood at a systems biology level, and studies that combine gene expression, protein expression, and liberation of peptides for antigen presentation can be combined to provide a detailed understanding of immunity. This chapter details our experience in identifying peptide antigens and combining this information with more traditional proteomics approaches to understand the generation of immune responses on a holistic level.
The allopurinol metabolite, oxypurinol, drives oligoclonal expansions of drug‐reactive T cells in resolved hypersensitivity cases and drug‐naïve healthy donors
Publisher: Wiley
Date: 14-07-2023
DOI: 10.1111/ALL.15814
Abstract: Allopurinol (ALP) is a successful drug used in the treatment of gout. However, this drug has been implicated in hypersensitivity reactions that can cause severe to life‐threatening reactions such as Stevens–Johnson syndrome (SJS) and toxic epidermal necrolysis (TEN). In iduals who carry the human leukocyte antigen (HLA)‐B*58:01 allotype are at higher risk of experiencing a hypersensitivity reaction (odds ratios ranging from 5.62 to 580.3 for mild to severe reactions, respectively). In addition to the parent drug, the metabolite oxypurinol (OXP) is implicated in triggering T cell‐mediated immunopathology via a labile interaction with HLA‐B*58:01. To date, there has been limited information regarding the T‐cell receptor (TCR) repertoire usage of reactive T cells in patients with ALP‐induced SJS or TEN and, in particular, there are no reports examining paired αβTCRs. Here, using in vitro drug‐treated PBMCs isolated from both resolved ALP‐induced SJS/TEN cases and drug‐naïve healthy donors, we show that OXP is the driver of CD8 + T cell‐mediated responses and that drug‐exposed memory T cells can exhibit a proinflammatory immunophenotype similar to T cells described during active disease. Furthermore, this response supported the pharmacological interaction with immune receptors (p‐i) concept by showcasing (i) the labile metabolite interaction with peptide/HLA complexes, (ii) immunogenic complex formation at the cell surface, and (iii) lack of requirement for antigen processing to elicit drug‐induced T cell responsiveness. Examination of paired OXP‐induced αβTCR repertoires highlighted an oligoclonal and private clonotypic profile in both resolved ALP‐induced SJS/TEN cases and drug‐naïve healthy donors.
Amyloid Self‐Assembly of hIAPP8‐20 via the Accumulation of Helical Oligomers, α‐Helix to β‐Sheet Transition, and Formation of β‐Barrel Intermediates
Publisher: Wiley
Date: 25-03-2019
Hard wiring of T cell receptor specificity for the major histocompatibility complex is underpinned by TCR adaptability
Publisher: Proceedings of the National Academy of Sciences
Date: 18-05-2010
Abstract: αβ T cell receptors (TCRs) are genetically restricted to corecognize peptide antigens bound to self-major histocompatibility complex (pMHC) molecules however, the basis for this MHC specificity remains unclear. Despite the current dogma, evaluation of the TCR–pMHC-I structural database shows that the nongermline-encoded complementarity-determining region (CDR)-3 loops often contact the MHC-I, and the germline-encoded CDR1 and -2 loops frequently participate in peptide-mediated interactions. Nevertheless, different TCRs adopt a roughly conserved docking mode over the pMHC-I, in which three MHC-I residues (65, 69, and 155) are invariably contacted by the TCR in one way or another. Nonetheless, the impact of mutations at these three positions, either in idually or together, was not uniformly detrimental to TCR recognition of pHLA-B*0801 or pHLA-B*3508. Moreover, when TCR–pMHC-I recognition was impaired, this could be partially restored by expression of the CD8 coreceptor. The structure of a TCR–pMHC-I complex in which these three (65, 69, and 155) MHC-I positions were all mutated resulted in shifting of the TCR footprint relative to the cognate complex and formation of compensatory interactions. Collectively, our findings reveal the inherent adaptability of the TCR in maintaining peptide recognition while accommodating changes to the central docking site on the pMHC-I.
Employing proteomics in the study of antigen presentation: an update
Publisher: Informa UK Limited
Date: 03-08-2018
DOI: 10.1080/14789450.2018.1509000
Abstract: Our immune system discriminates self from non-self by examining the peptide cargo of human leukocyte antigen (HLA) molecules displayed on the cell surface. Successful recognition of HLA-bound non-self peptides can induce T cell responses leading to, for ex le, the destruction of infected cells. Today, largely due to advances in technology, we have an unprecedented capability to identify the nature of these presented peptides and unravel the true complexity of antigen presentation. Areas covered: In addition to conventional linear peptides, HLA molecules also present post-translationally modified sequences comprising a wealth of chemical and structural modifications, including a novel class of noncontiguous spliced peptides. This review focuses on these emerging themes in antigen presentation and how mass spectrometry in particular has contributed to a new view of the antigenic landscape that is presented to the immune system. Expert Commentary: Advances in the sensitivity of mass spectrometers and use of hybrid fragmentation technologies will provide more information-rich spectra of HLA bound peptides leading to more definitive identification of T cell epitopes. Coupled with improvements in s le preparation and new informatics workflows, studies will access novel classes of peptide antigen and allow interrogation of rare and clinically relevant s les.
The peptide-loading complex and ligand selection during the assembly of HLA class I molecules
Publisher: Elsevier BV
Date: 06-2000
DOI: 10.1016/S0161-5890(00)00075-4
Abstract: The identification and characterisation of the class I peptide loading complex has resulted in an appreciation of the co-ordinated and multifaceted nature of HLA class I assembly in the lumen of the endoplasmic reticulum. This loading complex consists of the assembling class I heterodimer in association with a number of molecular chaperones. These chaperones can be classified as generic to the folding of most glycoproteins in the endoplasmic reticulum or specific to the class I loading pathway. The functions of the various components of the loading complex in class I molecule assembly are reviewed. A critical component of the class I loading complex is the specialised chaperone tapasin. The role of tapasin in the stabilisation and retention of empty or suboptimally loaded class I molecules and the facilitation of the loading of these molecules with more appropriate ligands is discussed. As such, it is proposed that tapasin is a major determinant of peptide repertoire selection for class I-restricted presentation in normal antigen presenting cells. The potential implications in vaccine design and autoimmunity are discussed.
Supplementary Figure and Table Legends from Spliced Peptides and Cytokine-Driven Changes in the Immunopeptidome of Melanoma
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/2326-6066.22542576
Abstract: Supplementary figure legends and tables descriptors
Discovery by proteogenomics and characterization of an RF-amide neuropeptide from cone snail venom
Publisher: Elsevier BV
Date: 2015
HLA class II immunopeptidomics reveals that co‐inherited HLA‐allotypes within an extended haplotype can improve proteome coverage for immunosurveillance
Publisher: Wiley
Date: 17-08-2021
Abstract: Human leucocyte antigen (HLA) class II molecules in humans are encoded by three different loci, HLA‐DR, ‐DQ, and ‐DP. These molecules share approximately 70% sequence similarity and all present peptide ligands to circulating T cells. While the peptide repertoires of numerous HLA‐DR, ‐DQ, and ‐DP allotypes have been examined, there have been few reports on the combined repertoire of these co‐inherited molecules expressed in a single cell as an extended HLA haplotype. Here we describe the endogenous peptide repertoire of a human B lymphoblastoid cell line (C1R) expressing the class II haplotype HLA‐DR12/DQ7/DP4. We have identified 71350 unique naturally processed peptides presented collectively by HLA‐DR12, HLA‐DQ7, or HLA‐DP4. The resulting “haplodome” is complemented by the cellular proteome defined by standard LC‐MS/MS approaches. This large dataset has shed light on properties of these class II ligands especially the preference for membrane and extracellular source proteins. Our data also provides insights into the co‐evolution of these conserved haplotypes of closely linked and co‐inherited HLA molecules which together increase sequence coverage of cellular proteins for immune surveillance with minimal overlap between each co‐inherited HLA‐class II allomorph.
Supplementary Table S1 from Spliced Peptides and Cytokine-Driven Changes in the Immunopeptidome of Melanoma
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/2326-6066.22542564.V1
Abstract: Table S1
The production, purification and crystallization of a soluble heterodimeric form of a highly selected T-cell receptor in its unliganded and liganded state
Publisher: International Union of Crystallography (IUCr)
Date: 26-11-2002
DOI: 10.1107/S0907444902015482
Abstract: T-cell antigen receptors (TcRs) are heterodimeric cell-surface receptors that play a pivotal role in the cellular immune response. The TcR interacts specifically with a peptide-laden major histocompatability complex (pMHC). A human TcR has been characterized that interacts with an immunodominant epitope, FLRGRAYGL, from the Epstein-Barr virus, a ubiquitous human pathogen, in complex with HLA-B8. Despite the vast TcR repertoire, this TcR is found in up to 10% of the total T-cell population in seropositive HLA-B8+ in iduals. In this report, this highly selected TcR is characterized by expressing in Escherichia coli, refolding, purifying and crystallizing the receptor. In addition, the HLA-B8-FLRGRAYGL complex has been expressed in E. coli, refolded and shown to be functionally active. Using native gel electrophoresis, the refolded TcR is shown to be capable of binding specifically to the refolded HLA-B8-FLRGRAYGL and this TcR has been crystallized in complex with the pMHC. The crystals of the unliganded and liganded TcR diffract to 1.5 and 2.5 A, respectively.
Defective Proinsulin Handling Modulates the MHC I Bound Peptidome and Activates the Inflammasome in β-Cells
Publisher: MDPI AG
Date: 30-03-2022
DOI: 10.3390/BIOMEDICINES10040814
Abstract: How immune tolerance is lost to pancreatic β-cell peptides triggering autoimmune type 1 diabetes is enigmatic. We have shown that loss of the proinsulin chaperone glucose-regulated protein (GRP) 94 from the endoplasmic reticulum (ER) leads to mishandling of proinsulin, ER stress, and activation of the immunoproteasome. We hypothesize that inadequate ER proinsulin folding capacity relative to biosynthetic need may lead to an altered β-cell major histocompatibility complex (MHC) class-I bound peptidome and inflammasome activation, sensitizing β-cells to immune attack. We used INS-1E cells with or without GRP94 knockout (KO), or in the presence or absence of GRP94 inhibitor PU-WS13 (GRP94i, 20 µM), or exposed to proinflammatory cytokines interleukin (IL)-1β or interferon gamma (IFNγ) (15 pg/mL and 10 ng/mL, respectively) for 24 h. RT1.A (rat MHC I) expression was evaluated using flow cytometry. The total RT1.A-bound peptidome analysis was performed on cell lysates fractionated by reverse-phase high-performance liquid chromatography (RP-HPLC), followed by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). The nucleotide-binding oligomerization domain, leucine rich repeat and pyrin domain containing protein (NLRP1), nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor alpha (IκBα), and (pro) IL-1β expression and secretion were investigated by Western blotting. GRP94 KO increased RT1.A expression in β-cells, as did cytokine exposure compared to relevant controls. Immunopeptidome analysis showed increased RT1.A-bound peptide repertoire in GRP94 KO/i cells as well as in the cells exposed to cytokines. The GRP94 KO/cytokine exposure groups showed partial overlap in their peptide repertoire. Notably, proinsulin-derived peptide ersity increased among the total RT1.A peptidome in GRP94 KO/i along with cytokines exposure. NLRP1 expression was upregulated in GRP94 deficient cells along with decreased IκBα content while proIL-1β cellular levels declined, coupled with increased secretion of mature IL-1β. Our results suggest that limiting β-cell proinsulin chaperoning enhances RT1.A expression alters the MHC-I peptidome including proinsulin peptides and activates inflammatory pathways, suggesting that stress associated with impeding proinsulin handling may sensitize β-cells to immune-attack.
Data from Spliced Peptides and Cytokine-Driven Changes in the Immunopeptidome of Melanoma
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/2326-6066.C.6549978
Abstract: Abstract Antigen recognition by CD8 sup + /sup T cells is governed by the pool of peptide antigens presented on the cell surface in the context of HLA class I complexes. Studies have shown not only a high degree of plasticity in the immunopeptidome, but also that a considerable fraction of all presented peptides is generated through proteasome-mediated splicing of noncontiguous regions of proteins to form novel peptide antigens. Here, we used high-resolution mass spectrometry combined with new bioinformatic approaches to characterize the immunopeptidome of melanoma cells in the presence or absence of IFNγ. In total, we identified more than 60,000 peptides from a single patient-derived cell line (LM-MEL-44) and demonstrated that IFNγ induced changes in the peptidome, with an overlap of only approximately 50% between basal and treated cells. Around 6% to 8% of the peptides were identified as i cis /i -spliced peptides, and 2,213 peptides (1,827 linear and 386 i cis /i -spliced peptides) were derived from known melanoma-associated antigens. These peptide antigens were equally distributed between the constitutive- and IFNγ-induced peptidome. We next examined additional HLA-matched patient-derived cell lines to investigate how frequently these peptides were identified and found that a high proportion of both linear and spliced peptides was conserved between in idual patient tumors, drawing on data amassing to more than 100,000 peptide sequences. Several of these peptides showed i in vitro /i immunogenicity across multiple patients with melanoma. These observations highlight the breadth and complexity of the repertoire of immunogenic peptides that can be exploited therapeutically and suggest that spliced peptides are a major class of tumor antigens. /
Supplementary Figures from Spliced Peptides and Cytokine-Driven Changes in the Immunopeptidome of Melanoma
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/2326-6066.22542573
Abstract: Supplementary Figures S1-S10
Mass spectral identification of Vc1.1 and differential distribution of conopeptides in the venom duct of conus victoriae. effect of post-translational modifications and disulfide isomerisation on bioactivity
Publisher: Springer Science and Business Media LLC
Date: 07-03-2009
High-performance liquid chromatography of amino acids, peptides and proteins
Publisher: Elsevier BV
Date: 1989
DOI: 10.1016/S0021-9673(01)93861-0
Abstract: The gradient elution behaviour of eight synthetic peptides encompassing residues [6-13] of human growth hormone, i.e. Leu1-Ser-Arg-Leu-Phe-Asp-Asn-Ala8, has been investigated, by using an octadecylsilica, a butylsilica, and a polymeric fluorocarbon as stationary phases. Quantitative expressions, derived from the linear-solvent-strength theory and the general plate-height theory, were used to assess the influence of gradient time on the relative retention and bandwidths of these peptides. It was demonstrated that the chromatographic properties of the cyclised imide form involving Asp6 are consistent with the formation of a highly stabilised hipathic helix, while the open-chain alpha- and beta-rearranged forms eluted as less rigid structures. The putative hydrophobic contact region consists of two leucine residues and one phenylalanine residue. From an analysis of the retention and bandwidth data obtained at pH 9, a surface-induced molecular reorientation of the beta-linked peptides was observed, in which the repulsion of the aspartyl carboxyl group from the hydrophobic stationary phase directs the C-terminal moiety away from the sorbent surface. Furthermore, the fluorocarbon sorbent exhibited characteristics favourable for use in preparative purification of these peptides. The present results demonstrate the sensitivity of reversed-phase high-performance liquid chromatography (RP-HPLC) to monitor small changes in the interactive behaviour of peptides with hydrocarbonaceous ligands and aquo-organic solvent combinations in reversed-phase systems. These observations further illustrate the general utility of HPLC for investigating the conformational behaviour of peptides at solid-liquid interfaces.
Pancreatic beta cells are highly susceptible to oxidative and ER stresses during the development of diabetes.
Publisher: American Chemical Society (ACS)
Date: 08-12-2015
DOI: 10.1021/PR500643H
Abstract: The complex interplay of many cell types and the temporal heterogeneity of pancreatic islet composition obscure the direct role of resident alpha and beta cells in the development of Type 1 diabetes. Therefore, in addition to studying islets isolated from non-obese diabetic mice, we analyzed homogeneous cell populations of murine alpha (αTC-1) and beta (NIT-1) cell lines to understand the role and differential survival of these two predominant islet cell populations. A total of 56 proteins in NIT-1 cells and 50 in αTC-1 cells were differentially expressed when exposed to proinflammatory cytokines. The major difference in the protein expression between cytokine-treated NIT-1 and αTC-1 cells was free radical scavenging enzymes. A similar observation was made in cytokine-treated whole islets, where a comprehensive analysis of subcellular fractions revealed that 438 unique proteins were differentially expressed under inflammatory conditions. Our data indicate that beta cells are relatively susceptible to ER and oxidative stress and reveal key pathways that are dysregulated in beta cells during cytokine exposure. Additionally, in the islets, inflammation also leads to enhanced antigen presentation, which completes a three-way insult on beta cells, rendering them targets of infiltrating T lymphocytes.
Identification of Conus Peptidylprolyl Cis-Trans Isomerases (PPIases) and Assessment of Their Role in the Oxidative Folding of Conotoxins
Publisher: Elsevier BV
Date: 04-2010
Diversity of Conotoxin Gene Superfamilies in the Venomous Snail, Conus victoriae
Publisher: Public Library of Science (PLoS)
Date: 05-02-2014
High-performance liquid chromatography of amino acids, peptides, and proteins. 123. Dynamics of peptides in reversed-phase high-performance liquid chromatography
Publisher: American Chemical Society (ACS)
Date: 11-1993
DOI: 10.1021/AC00069A016
Abstract: The dynamics of several peptides in reversed-phase high-performance liquid chromatography (RP-HPLC) have been investigated on both n-octadecyl (C18) silica and n-butyl (C4) silica sorbents. In particular, the conformational interconversions and the relative rates of chromatographic relaxation of bombesin, glucagon, and beta-endorphin on both C18 and C4 n-alkylsilicas were monitored by examining changes in the experimental bandwidths of these peptides as a function of temperature and column residence time under linear gradient elution RP-HPLC conditions. The observed band-broadening trends were correlated with previously derived retention parameters and thermodynamic descriptors of the association process determined for bombesin, beta-endorphin, glucagon, and a control peptide, penta-L-phenylalanine. This study confirms that bandwidth measurements can be used as an integral experimental component to study the effect of the secondary structure of peptidic solutes on their RP-HPLC retention behavior. Further, the data demonstrate the utility of RP-HPLC as a tool to examine peptide conformational dynamics at hydrophobic surfaces. The relevance of these results to the general phenomenon of peptide-lipid interactions is discussed in terms of the associated evidence for lipid-induced changes in the conformation of these three bioactive peptides.
The Protein Import Channel in the Outer Mitosomal Membrane of Giardia intestinalis
Publisher: Oxford University Press (OUP)
Date: 16-06-2009
Benchmarking predictions of MHC class I restricted T cell epitopes in a comprehensively studied model system
Publisher: Public Library of Science (PLoS)
Date: 26-05-2020
Natural micropolymorphism in human leukocyte antigens provides a basis for genetic control of antigen recognition
Publisher: Rockefeller University Press
Date: 12-01-2009
DOI: 10.1084/JEM.20082136
Abstract: Human leukocyte antigen (HLA) gene polymorphism plays a critical role in protective immunity, disease susceptibility, autoimmunity, and drug hypersensitivity, yet the basis of how HLA polymorphism influences T cell receptor (TCR) recognition is unclear. We examined how a natural micropolymorphism in HLA-B44, an important and large HLA allelic family, affected antigen recognition. T cell–mediated immunity to an Epstein-Barr virus determinant (EENLLDFVRF) is enhanced when HLA-B*4405 was the presenting allotype compared with HLA-B*4402 or HLA-B*4403, each of which differ by just one amino acid. The micropolymorphism in these HLA-B44 allotypes altered the mode of binding and dynamics of the bound viral epitope. The structure of the TCR–HLA-B*4405EENLLDFVRF complex revealed that peptide flexibility was a critical parameter in enabling preferential engagement with HLA-B*4405 in comparison to HLA-B*4402/03. Accordingly, major histocompatibility complex (MHC) polymorphism can alter the dynamics of the peptide-MHC landscape, resulting in fine-tuning of T cell responses between closely related allotypes.
TAP genes and immunity
Publisher: Elsevier BV
Date: 10-2004
RHybridFinder: An R package to process immunopeptidomic data for putative hybrid peptide discovery
Publisher: Elsevier BV
Date: 12-2021
A sensitive method for detecting proliferation of rare autoantigen-specific human T cells
Publisher: Elsevier BV
Date: 12-2003
DOI: 10.1016/J.JIM.2003.09.004
Abstract: The ability to measure proliferation of rare antigen-specific T cells among many bystanders is critical for the evaluation of cellular immune function in health and disease. T-cell proliferation in response to antigen has been measured almost exclusively by 3H-thymidine incorporation. This method does not directly identify the phenotype of the proliferating cells and is frequently not sufficiently sensitive to detect rare autoantigen-specific T cells. To overcome these problems, we developed a novel assay for antigen-specific human T-cell proliferation. Peripheral blood mononuclear cells (PBMC) were labelled with the fluorescent dye 5,6-carboxylfluorescein diacetate succinimidyl ester (CFSE) and cells that proliferated in response to antigen, with resultant reduction in CFSE intensity, were measured directly by flow cytometry. This assay was more sensitive than 3H-thymidine incorporation and detected the proliferation of rare antigen-specific CD4(+) T cells at 10-fold lower antigen concentrations. It also allowed the phenotype of the proliferating cells to be directly determined. Using the CFSE assay we were able to measure directly the proliferation of human CD4(+) T cells from healthy donors in response to the type 1 diabetes autoantigens glutamic acid decarboxylase (GAD) and proinsulin (PI).
Human Leukocyte Antigen Class I-Restricted Activation of CD8+ T Cells Provides the Immunogenetic Basis of a Systemic Drug Hypersensitivity
Publisher: Elsevier BV
Date: 06-2008
DOI: 10.1016/J.IMMUNI.2008.04.020
Abstract: The basis for strong immunogenetic associations between particular human leukocyte antigen (HLA) class I allotypes and inflammatory conditions like Behçet's disease (HLA-B51) and ankylosing spondylitis (HLA-B27) remain mysterious. Recently, however, even stronger HLA associations are reported in drug hypersensitivities to the reverse-transcriptase inhibitor abacavir (HLA-B57), the gout prophylactic allopurinol (HLA-B58), and the antiepileptic carbamazepine (HLA-B*1502), providing a defined disease trigger and suggesting a general mechanism for these associations. We show that systemic reactions to abacavir were driven by drug-specific activation of cytokine-producing, cytotoxic CD8+ T cells. Recognition of abacavir required the transporter associated with antigen presentation and tapasin, was fixation sensitive, and was uniquely restricted by HLA-B*5701 and not closely related HLA allotypes with polymorphisms in the antigen-binding cleft. Hence, the strong association of HLA-B*5701 with abacavir hypersensitivity reflects specificity through creation of a unique ligand as well as HLA-restricted antigen presentation, suggesting a basis for the strong HLA class I-association with certain inflammatory disorders.
CD8+ T cell landscape in Indigenous and non-Indigenous people restricted by influenza mortality-associated HLA-A*24:02 allomorph
Publisher: Springer Science and Business Media LLC
Date: 18-05-2021
DOI: 10.1038/S41467-021-23212-X
Abstract: Indigenous people worldwide are at high risk of developing severe influenza disease. HLA-A*24:02 allele, highly prevalent in Indigenous populations, is associated with influenza-induced mortality, although the basis for this association is unclear. Here, we define CD8 + T-cell immune landscapes against influenza A (IAV) and B (IBV) viruses in HLA-A*24:02-expressing Indigenous and non-Indigenous in iduals, human tissues, influenza-infected patients and HLA-A*24:02-transgenic mice. We identify immunodominant protective CD8 + T-cell epitopes, one towards IAV and six towards IBV, with A24/PB2 550–558 -specific CD8 + T cells being cross-reactive between IAV and IBV. Memory CD8 + T cells towards these specificities are present in blood (CD27 + CD45RA − phenotype) and tissues (CD103 + CD69 + phenotype) of healthy in iduals, and effector CD27 − CD45RA − PD-1 + CD38 + CD8 + T cells in IAV/IBV patients. Our data show influenza-specific CD8 + T-cell responses in Indigenous Australians, and advocate for T-cell-mediated vaccines that target and boost the breadth of IAV/IBV-specific CD8 + T cells to protect high-risk HLA-A*24:02-expressing Indigenous and non-Indigenous populations from severe influenza disease.
Inability to detect cross-reactive memory T cells challenges the frequency of heterologous immunity among common viruses
Publisher: The American Association of Immunologists
Date: 15-06-2018
Abstract: Human memory T cells that cross-react with epitopes from unrelated viruses can potentially modulate immune responses to subsequent infections by a phenomenon termed heterologous immunity. However, it is unclear whether similarities in structure rather than sequence underpin heterologous T cell cross-reactivity. In this study, we aimed to explore the mechanism of heterologous immunity involving immunodominant epitopes derived from common viruses restricted to high-frequency HLA allotypes (HLA-A*02:01, -B*07:02, and -B*08:01). We examined EBV-specific memory T cells for their ability to cross-react with CMV or influenza A virus–derived epitopes. Following T cell immunoassays to determine phenotype and function, complemented with biophysical and structural investigations of peptide/HLA complexes, we did not detect cross-reactivity of EBV-specific memory T cells toward either CMV or influenza A virus epitopes presented by any of the selected HLA allomorphs. Thus, despite the ubiquitous nature of these human viruses and the dominant immune response directed toward the selected epitopes, heterologous virus-specific T cell cross-reactivity was not detected. This suggests that either heterologous immunity is not as common as previously reported, or that it requires a very specific biological context to develop and be clinically relevant.
Human Leukocyte Antigen (HLA) B27 Allotype-Specific Binding and Candidate Arthritogenic Peptides Revealed through Heuristic Clustering of Data-independent Acquisition Mass Spectrometry (DIA-MS) Data
Publisher: Elsevier BV
Date: 06-2016
The CDR3 regions of an immunodominant T cell receptor dictate the 'energetic landscape' of peptide-MHC recognition
Publisher: Springer Science and Business Media LLC
Date: 09-01-2005
DOI: 10.1038/NI1155
Abstract: The energetic bases of T cell recognition are unclear. Here, we studied the 'energetic landscape' of peptide-major histocompatibility complex (pMHC) recognition by an immunodominant alphabeta T cell receptor (TCR). We quantified and evaluated the effect of natural and systematic substitutions in the complementarity-determining region (CDR) loops on ligand binding in the context of the structural detail of each component of the immunodominant TCR-pMHC complex. The CDR1 and CDR2 loops contributed minimal energy through direct recognition of the antigen and instead had a chief function in stabilizing the ligated CDR3 loops. The underlying energetic basis for recognition lay in the CDR3 loops. Therefore the energetic burden of the CDR loops in the TCR-pMHC interaction is variable among TCRs, reflecting the inherent adaptability of the TCR in ligating different ligands.
C-terminal Src kinase-homologous kinase (CHK), a unique inhibitor inactivating multiple active conformations of Src family tyrosine kinases
Publisher: Elsevier BV
Date: 2015
Mouse and human microglial phenotypes in Alzheimer’s disease are controlled by amyloid plaque phagocytosis through Hif1α
Publisher: Cold Spring Harbor Laboratory
Date: 17-05-2019
DOI: 10.1101/639054
Abstract: The important role of microglia, the brain’s resident immune cells, in Alzheimer’s disease (AD) is now well recognized, however their molecular and functional ersity and underlying mechanisms still remain controversial. To transcriptionally and functionally characterize the ersity of microglia in AD and aging, we isolated the amyloid plaque-containing (XO4 + ) and non-containing (XO4 − ) microglia from an AD mouse model. Transcriptomics analysis unveiled independent transcriptional trajectories in ageing and AD. XO4 + microglial transcriptomes linked plaque phagocytosis to altered expression of bona fide late onset AD genetic risk factors. We further revealed that the XO4 + transcriptional program is present in a subset of human microglia from AD patients and is a direct and reversible consequence of Aβ plaque phagocytosis. Conversely, XO4 − microglia in AD displayed an accelerated ageing signature and contained more intracellular post synaptic material than plaque-containing microglia, despite reduced active synaptosome phagocytosis. Mechanistically, we predicted HIF1α as a core regulator of the XO4 − /XO4 + axis, and further validated the mechanism in vitro using human stem cell-derived microglia like cells and primary human microglia. Together these findings unveiled the molecular mechanism underpinning the functional ersity of microglia in AD, providing opportunities to develop treatments targeted at subset specific manipulation of the microglial niche.
Protein secretion and outer membrane assembly inAlphaproteobacteria
Publisher: Oxford University Press (OUP)
Date: 11-2008
Supplementary Table S6 from Spliced Peptides and Cytokine-Driven Changes in the Immunopeptidome of Melanoma
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/2326-6066.22542549
Abstract: Table S6
C-terminal Src kinase-homologous kinase (CHK), a unique inhibitor inactivating multiple active conformations of Src family tyrosine kinases
Publisher: Elsevier BV
Date: 11-2006
Supplementary Table S5 from Spliced Peptides and Cytokine-Driven Changes in the Immunopeptidome of Melanoma
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/2326-6066.22542552
Abstract: Table S5
The immunogenicity of a viral cytotoxic T cell epitope is controlled by its MHC-bound conformation
Publisher: Rockefeller University Press
Date: 07-11-2005
DOI: 10.1084/JEM.20050864
Abstract: Thousands of potentially antigenic peptides are encoded by an infecting pathogen however, only a small proportion induce measurable CD8+ T cell responses. To investigate the factors that control peptide immunogenicity, we have examined the cytotoxic T lymphocyte (CTL) response to a previously undefined epitope (77APQPAPENAY86) from the BZLF1 protein of Epstein-Barr virus (EBV). This peptide binds well to two human histocompatibility leukocyte antigen (HLA) allotypes, HLA-B*3501 and HLA-B*3508, which differ by a single amino acid at position 156 (156Leucine vs. 156Arginine, respectively). Surprisingly, only in iduals expressing HLA-B*3508 show evidence of a CTL response to the 77APQPAPENAY86 epitope even though EBV-infected cells expressing HLA-B*3501 process and present similar amounts of peptide for CTL recognition, suggesting that factors other than peptide presentation levels are influencing immunogenicity. Functional and structural analysis revealed marked conformational differences in the peptide, when bound to each HLA-B35 allotype, that are dictated by the polymorphic HLA residue 156 and that directly affected T cell receptor recognition. These data indicate that the immunogenicity of an antigenic peptide is influenced not only by how well the peptide binds to major histocompatibility complex (MHC) molecules but also by its bound conformation. It also illustrates a novel mechanism through which MHC polymorphism can further ersify the immune response to infecting pathogens.
Supplementary Table S4 from Spliced Peptides and Cytokine-Driven Changes in the Immunopeptidome of Melanoma
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/2326-6066.22542555
Abstract: Table S4
Supplementary Table S3 from Spliced Peptides and Cytokine-Driven Changes in the Immunopeptidome of Melanoma
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/2326-6066.22542558
Abstract: Table S3
Application of quantitative proteomics technologies to the biomarker discovery pipeline for multiple sclerosis
Publisher: Wiley
Date: 2013
Abstract: Multiple sclerosis is an inflammatory-mediated demyelinating disorder most prevalent in young Caucasian adults. The various clinical manifestations of the disease present several challenges in the clinic in terms of diagnosis, monitoring disease progression and response to treatment. Advances in MS-based proteomic technologies have revolutionized the field of biomarker research and paved the way for the identification and validation of disease-specific markers. This review focuses on the novel candidates discovered by the application of quantitative proteomics to relevant disease-affected tissues in both the human context and within the animal model of the disease known as experimental autoimmune encephalomyelitis. The role of targeted MS approaches for biomarker validation studies, such as multiple reaction monitoring will also be discussed.
Probing the Binding Behavior and Conformational States of Globular Proteins in Reversed-Phase High-Performance Liquid Chromatography
Publisher: American Chemical Society (ACS)
Date: 22-05-1999
DOI: 10.1021/AC9808369
Abstract: Reversed-phase high-performance liquid chromatography (RP-HPLC) is a widely used technique for the separation of proteins under low pH aquo-organic solvent gradient elution conditions, typically carried out at ambient temperatures. These conditions can however induce conformational effects with proteins as evident from changes in their biological or immunological activities. By monitoring the influence of temperature on the retention and band-broadening characteristics of proteins, the role of conformational processes in these lipophilic environments can be examined. These processes can then be interpreted in terms of a two-state model involving a native (N) and a fully unfolded species (U) or more complex folding/unfolding models. In the present study, the gradient elution RP-HPLC behavior of sperm whale myoglobin (SWMYO) and hen egg white lysozyme (HEWL) has been investigated at temperatures between 5 and 85 degrees C with n-octadecyl (C18)- and n-butyl (C4)-silica reversed-phase sorbents. The interaction of these proteins with these reversed-phase sorbents has also been examined in terms of the contributions that the heme prosthetic group of SWMYO and the disulfide bonds in HEWL make to the stabilization of the native conformation of these proteins in these hydrophobic environments. The observed interconversions of multiple peak zones of SWMYO and HEWL in the presence of C18 and C4 ligands have been subsequently analyzed in terms of the unfolding processes that these proteins can undergo at low pH and at elevated temperatures. The ability of hydrocarbonaceous ligands to trap ensemblies of partially unfolded conformational intermediates of proteins in these perturbing environments has been examined. Pseudo-first-order rate constants have been derived for these processes from analysis of the dependencies on time of the concentration of the different protein species at specified temperatures. The relationship of these processes to the conformational transitions that these proteins can undergo via molten globule-like intermediates (i.e., compact denatured states with a significant amount of residual secondary structure) in solution has also been examined. This study thus further documents an experimental strategy to assess the folding/unfolding behavior of globular proteins in the presence of hydrophobic surfaces and aquo-organic solvents, whereby the system parameters can potentially affect the preservation of native conformations, and thus the function, of the protein under these conditions.
Disparate thermodynamics governing T cell receptor-MHC-1 interactions implicate extrinsic factors in guiding MHC restriction
Publisher: Proceedings of the National Academy of Sciences
Date: 25-04-2006
Abstract: The underlying basis of major histocompatibility complex (MHC) restriction is unclear. Nevertheless, current data suggest that a common thermodynamic signature dictates αβ T cell receptor (TcR) ligation. To evaluate whether this thermodynamic signature defines MHC restriction, we have examined the thermodynamic basis of a highly characterized immunodominant TcR interacting with its cognate peptide–MHC-I ligand. Surprisingly, we observed this interaction to be governed by favorable enthalpic and entropic forces, which is in contrast to the prevailing generality, namely, enthalpically driven interactions combined with markedly unfavorable entropic forces. We conclude that extrinsic molecular factors, such as coreceptor ligation, conformational adjustments involved in TcR signaling, or constraints dictated by higher-order arrangement of ligated TcRs, might play a greater role in guiding MHC restriction than appreciated previously.
Discovery of novel disease-specific and membrane-associated candidate markers in a mouse model of multiple sclerosis
Publisher: Elsevier BV
Date: 03-2014
Protective efficacy of cross-reactive CD8+ T cells recognising mutant viral epitopes depends on peptide-MHC-I structural interactions and T cell activation threshold.
Publisher: Public Library of Science (PLoS)
Date: 12-08-2010
High-performance liquid chromatography of amino acids, peptides and proteins
Publisher: Elsevier BV
Date: 1989
DOI: 10.1016/S0021-9673(01)93862-2
Abstract: The influence of temperature on the gradient elution properties of synthetic peptides related to residues [6-13] of human growth hormone, e.g., Leu1-Ser-Arg-Leu-Phe-Asp-Asn-Ala8, has been studied by using both an octadecylsilica and a polymeric fluorocarbon stationary phase. Correlation of changes in the solute hydrophobic contact area and affinity for the stationary phase, as given by S and log k0 values respectively, revealed that the alpha- and imide forms are more conformationally stable than the beta-linked peptide. In addition, negative values of the standard entropy change, delta S0 assoc, for the transfer of the solute to the stationary phase, were observed for both alpha- and beta-linked peptides. These results are indicative of an increased ordering of the system upon solute adsorption and implies that the open-chain peptides exist in solution in more flexible conformations, while the helical structure of the cyclised imide is more rigid and constrained. The implications of the relative conformational stability of these peptides in their role as insulin-potentiating agents is also discussed.
Isolation of T cell receptors targeting recurrent neoantigens in hematological malignancies
Publisher: BMJ
Date: 13-07-2018
A novel strategy for the targeted analysis of protein and peptide metabolites.
Publisher: Wiley
Date: 15-12-2011
Prevention of Cytotoxic T Cell Escape Using a Heteroclitic Subdominant Viral T Cell Determinant
Publisher: Public Library of Science (PLoS)
Date: 24-10-2008
Drugs and drug-like molecules can modulate the function of mucosal-associated invariant T cells
Publisher: Springer Science and Business Media LLC
Date: 06-02-2017
DOI: 10.1038/NI.3679
Abstract: The major-histocompatibility-complex-(MHC)-class-I-related molecule MR1 can present activating and non-activating vitamin-B-based ligands to mucosal-associated invariant T cells (MAIT cells). Whether MR1 binds other ligands is unknown. Here we identified a range of small organic molecules, drugs, drug metabolites and drug-like molecules, including salicylates and diclofenac, as MR1-binding ligands. Some of these ligands inhibited MAIT cells ex vivo and in vivo, while others, including diclofenac metabolites, were agonists. Crystal structures of a T cell antigen receptor (TCR) from a MAIT cell in complex with MR1 bound to the non-stimulatory and stimulatory compounds showed distinct ligand orientations and contacts within MR1, which highlighted the versatility of the MR1 binding pocket. The findings demonstrated that MR1 was able to capture chemically erse structures, spanning mono- and bicyclic compounds, that either inhibited or activated MAIT cells. This indicated that drugs and drug-like molecules can modulate MAIT cell function in mammals.
Structural and functional correlates of enhanced antiviral immunity generated by heteroclitic CD8 T cell epitopes
Publisher: The American Association of Immunologists
Date: 06-2014
Abstract: Peptides that bind poorly to MHC class I molecules often elicit low–functional avidity T cell responses. Peptide modification by altering the anchor residue facilitates increased binding affinity and may elicit T cells with increased functional avidity toward the native epitope (“heteroclitic”). This augmented MHC binding is likely to increase the half-life and surface density of the heteroclitic complex, but precisely how this enhanced T cell response occurs in vivo is not known. Furthermore, the ideal heteroclitic epitope will elicit T cell responses that completely cross-react with the native epitope, maximizing protection and minimizing undesirable off-target effects. Such epitopes have been difficult to identify. In this study, using mice infected with a murine coronavirus that encodes epitopes that elicit high (S510, CSLWNGPHL)– and low (S598, RCQIFANI)–functional avidity responses, we show that increased expression of peptide S598 but not S510 generated T cells with enhanced functional avidity. Thus, immune responses can be augmented toward T cell epitopes with low functional avidity by increasing Ag density. We also identified a heteroclitic epitope (RCVIFANI) that elicited a T cell response with nearly complete cross-reactivity with native epitope and demonstrated increased MHC eptide abundance compared with native S598. Structural and thermal melt analyses indicated that the Q600V substitution enhanced stability of the peptide/MHC complex without greatly altering the antigenic surface, resulting in highly cross-reactive T cell responses. Our data highlight that increased peptide/MHC complex display contributes to heteroclitic epitope efficacy and describe parameters for maximizing immune responses that cross-react with the native epitope.
Immunopeptidomic Analysis Reveals That Deamidated HLA-bound Peptides Arise Predominantly from Deglycosylated Precursors
Publisher: Elsevier BV
Date: 07-2020
The molecular basis for peptide repertoire selection in the human leukocyte antigen (HLA) C*06:02 molecule
Publisher: Elsevier BV
Date: 10-2017
T cell receptor recognition of a 'super-bulged' major histocompatibility complex class I–bound peptide
Publisher: Springer Science and Business Media LLC
Date: 25-09-2005
DOI: 10.1038/NI1257
Abstract: Unusually long major histocompatibility complex (MHC) class I-restricted epitopes are important in immunity, but their 'bulged' conformation represents a potential obstacle to alphabeta T cell receptor (TCR)-MHC class I docking. To elucidate how such recognition is achieved while still preserving MHC restriction, we have determined here the structure of a TCR in complex with HLA-B(*)3508 presenting a peptide 13 amino acids in length. This complex was atypical of TCR-peptide-MHC class I interactions, being dominated at the interface by peptide-mediated interactions. The TCR assumed two distinct orientations, swiveling on top of the centrally bulged, rigid peptide such that only limited contacts were made with MHC class I. Although the TCR-peptide recognition resembled an antibody-antigen interaction, the TCR-MHC class I contacts defined a minimal 'generic footprint' of MHC-restriction. Thus our findings simultaneously demonstrate the considerable adaptability of the TCR and the 'shape' of MHC restriction.
Supplementary Table S4 from Spliced Peptides and Cytokine-Driven Changes in the Immunopeptidome of Melanoma
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/2326-6066.22542555.V1
Abstract: Table S4
The Beta Cell Immunopeptidome
Publisher: Elsevier
Date: 2014
A natural product compound inhibits coronaviral replication in vitro by binding to the conserved Nsp9 SARS-CoV-2 protein
Publisher: Elsevier BV
Date: 12-2021
The three-dimensional structure of an H-superfamily conotoxin reveals a granulin fold arising from a common ICK cysteine framework
Publisher: Elsevier BV
Date: 05-2019
T Cell Allorecognition via Molecular Mimicry
Publisher: Elsevier BV
Date: 12-2009
DOI: 10.1016/J.IMMUNI.2009.09.025
Abstract: T cells often alloreact with foreign human leukocyte antigens (HLA). Here we showed the LC13 T cell receptor (TCR), selected for recognition on self-HLA-B( *)0801 bound to a viral peptide, alloreacts with B44 allotypes (HLA-B( *)4402 and HLA-B( *)4405) bound to two different allopeptides. Despite extensive polymorphism between HLA-B( *)0801, HLA-B( *)4402, and HLA-B( *)4405 and the disparate sequences of the viral and allopeptides, the LC13 TCR engaged these peptide-HLA (pHLA) complexes identically, accommodating mimicry of the viral peptide by the allopeptide. The viral and allopeptides adopted similar conformations only after TCR ligation, revealing an induced-fit mechanism of molecular mimicry. The LC13 T cells did not alloreact against HLA-B( *)4403, and the single residue polymorphism between HLA-B( *)4402 and HLA-B( *)4403 affected the plasticity of the allopeptide, revealing that molecular mimicry was associated with TCR specificity. Accordingly, molecular mimicry that is HLA and peptide dependent is a mechanism for human T cell alloreactivity between disparate cognate and allogeneic pHLA complexes.
Repertoire of Classical MHC Class I and Class II Molecules
Publisher: Elsevier
Date: 2016
In Immunopeptidomics We Need a Sniper Instead of a Shotgun
Publisher: Wiley
Date: 07-03-2018
Abstract: Immunopeptidomics employs the use of mass spectrometry to identify and quantify peptides presented on the surface of cells by major histocompatibility complex (MHC human leukocyte antigen [HLA], in humans) molecules, an essential component of adaptive immunity. Currently, immunopeptidomics follows the same or similar workflows as the more established field of shotgun proteomics, yet inherent differences between these two fields create significant drawbacks for the former. In this viewpoint, we would like to highlight such technical issues and provide suggestions for novel workflows that would increase peptide sequencing coverage, depth, and confidence, collectively enhancing the capabilities of the field of immunopeptidomics.
HLA and Drug Hypersensitivity
Publisher: Elsevier
Date: 2016
Expression and purification of the minor histocompatibility antigen, HA-1H generated in Escherichia coli.
Publisher: Elsevier BV
Date: 07-2007
DOI: 10.1016/J.PEP.2007.02.011
Abstract: The minor histocompatibility antigen HA-1H is a potential immunotherapeutic molecule. It can be used as a target for graft versus leukaemia reactions to eliminate residual HA-1H expressing leukaemic cells in patients following haemopoietic stem cell transplantation, whereby HA-1H primed donor cells can be transferred into a patient via adoptive immunotherapy. However, thus far only synthetic peptides corresponding to a HLA-A *0201 restricted HA-1H epitope have been used to generate HA-1H specific T cells. We are the first laboratory to clone, express and purify a region of HA-1H using an Escherichia coli expression system. The recombinant HA-1H protein was purified under denaturing conditions and the affinity purification tag removed using thrombin to remove non-specific amino acids. The 92 amino acid recombinant protein was characterised by mass spectrometry. Our rationale is that by using a recombinant HA-1H protein rather than peptide, HA-1H specific T cells may be generated from presentation of multiple HA-1H epitopes complexed in different HLA molecules. A multi-epitope approach may have wider applicability and maybe more effective at leukaemia control. The recombinant HA-1H protein may also be used as a research tool to identify novel CD4(+) helper T cell and CD8(+) cytotoxic T cell epitopes.
Antagonism of Antiviral and Allogeneic Activity of a Human Public CTL Clonotype by a Single Altered Peptide Ligand: Implications for Allograft Rejection
Publisher: The American Association of Immunologists
Date: 05-2005
DOI: 10.4049/JIMMUNOL.174.9.5593
Abstract: Alloreactive T lymphocytes are central mediators of graft-versus-host disease and allograft rejection. A public CTL clonotype with specificity for the alloantigens HLA-B*4402 and B*4405 is often expanded to large numbers in healthy HLA-B*0801+ in iduals, driven by cross-reactive stimulation with the common, persistent herpesvirus EBV. Since such alloreactive memory CTL expansions have the potential to influence transplantation outcome, altered peptide ligands (APLs) of the target HLA-B*0801-binding EBV peptide, FLRGRAYGL, were screened as specific antagonists for this immunodominant clonotype. One APL, FLRGRFYGL, exerted powerful antagonism of a prototypic T cell clone expressing this immunodominant TCR when costimulated with target cells presenting HLA-B*0801FLRGRAYGL. Significantly, this APL also reduced the lysis of allogeneic target cells expressing HLA-B*4402 by up to 99%. The affinities of the agonist and antagonist complexes for the public TCR, measured using solution and solid-phase assays, were 8 and 138 μM, respectively. Surprisingly, the half-life of the agonist and antagonist complexes was similar, yet the association rate for the antagonist complex was significantly slower. These observations were further supported by structural studies that suggested a large conformational hurdle was required to ligate the immunodominant TCR to the HLA-B*0801 antagonist complex. By defining an antagonist APL against an immunodominant alloreactive TCR, these findings raise the prospect of exploiting such peptides to inhibit clinical alloreactivity, particularly against clonal T cell expansions that react with alloantigens.
HLA-A∗11:01-restricted CD8+ T cell immunity against influenza A and influenza B viruses in Indigenous and non-Indigenous people
Publisher: Public Library of Science (PLoS)
Date: 07-03-2022
DOI: 10.1371/JOURNAL.PPAT.1010337
Abstract: HLA-A*11:01 is one of the most prevalent human leukocyte antigens (HLAs), especially in East Asian and Oceanian populations. It is also highly expressed in Indigenous people who are at high risk of severe influenza disease. As CD8 + T cells can provide broadly cross-reactive immunity to distinct influenza strains and subtypes, including influenza A, B and C viruses, understanding CD8 + T cell immunity to influenza viruses across prominent HLA types is needed to rationally design a universal influenza vaccine and generate protective immunity especially for high-risk populations. As only a handful of HLA-A*11:01-restricted CD8 + T cell epitopes have been described for influenza A viruses (IAVs) and epitopes for influenza B viruses (IBVs) were still unknown, we embarked on an epitope discovery study to define a CD8 + T cell landscape for HLA-A*11:01-expressing Indigenous and non-Indigenous Australian people. Using mass-spectrometry, we identified IAV- and IBV-derived peptides presented by HLA-A*11:01 during infection. 79 IAV and 57 IBV peptides were subsequently screened for immunogenicity in vitro with peripheral blood mononuclear cells from HLA-A*11:01-expressing Indigenous and non-Indigenous Australian donors. CD8 + T cell immunogenicity screening revealed two immunogenic IAV epitopes (A11/PB2 320-331 and A11/PB2 323-331 ) and the first HLA-A*11:01-restricted IBV epitopes (A11/M 41-49 , A11/NS1 186-195 and A11/NP 511-520 ). The immunogenic IAV- and IBV-derived peptides were % conserved among their respective influenza viruses. Identification of novel immunogenic HLA-A*11:01-restricted CD8 + T cell epitopes has implications for understanding how CD8 + T cell immunity is generated towards IAVs and IBVs. These findings can inform the development of rationally designed, broadly cross-reactive influenza vaccines to ensure protection from severe influenza disease in HLA-A*11:01-expressing in iduals.
Proteomic interrogation of venom delivery in marine cone snails: Novel insights into the role of the venom bulb
Publisher: American Chemical Society (ACS)
Date: 29-09-2010
DOI: 10.1021/PR100431X
Abstract: Cone snails of the genus Conus are predatory marine gastropods mainly found in the shallow waters of the tropics and warm temperate seas. To prey on other marine organisms including fish, cone snails have evolved complex venoms synthesized and delivered by a highly sophisticated venom apparatus. Upon prey discovery, the venom is perfused through a harpoon-like radula tooth and rapidly injected into the prey to cause paralysis. While the venom components of cone snails have been intensively characterized, the mechanism of venom translocation and loading prior to and during injection remains elusive. The involvement of the venom bulb, a muscular dilation of the venom gland has been suggested, however evidence is sparse. Here, we use a combination of proteomics, molecular biology, and morphological examination to elucidate the potential role of the venom bulb in venom translocation and delivery. Analysis of the venom bulb proteome clearly demonstrated a function of this organ in muscular movement and, more interestingly, in burst muscle contraction. Morphological examination revealed high structural similarities to the mantle muscle of squids, animals known for their rapid escape response. We sequenced and further characterized arginine kinase, a key protein of rapid muscular movement in invertebrates and show high concentrations of this enzyme in the bulb when compared to the venom gland and the foot muscle. Proteins characteristic for venom biosynthesis were low in abundance. On the basis of our findings, we suggest that the bulb of cone snails is a highly specialized organ of venom translocation. Delivery of venom is driven by burst contractions of the bulb rapidly forcing the venom through the radula tooth into the prey.
CD1a on Langerhans cells controls inflammatory skin disease
Publisher: Springer Science and Business Media LLC
Date: 22-08-2016
DOI: 10.1038/NI.3523
The Choice of HLA‐Associated Peptide Enrichment and Purification Strategy Affects Peptide Yields and Creates a Bias in Detected Sequence Repertoire
Publisher: Wiley
Date: 06-2020
Response to Comment on a subset of HLA-I peptides are not genomically templated: Evidence for cis- And trans-spliced peptide ligands
Publisher: American Association for the Advancement of Science (AAAS)
Date: 02-08-2019
DOI: 10.1126/SCIIMMUNOL.AAW8457
Abstract: This is our response to the Technical Comment by Rolfs et al. where we point out errors in their reanalysis of our data.
Mismatch in epitope specificities between IFNγ inflamed and uninflamed conditions leads to escape from T lymphocyte killing in melanoma
Publisher: BMJ
Date: 16-02-2016
Reactivity with dichotomous determinants of Ro 60 stratifies autoantibody responses in lupus and primary Sjögren's syndrome.
Publisher: Wiley
Date: 29-04-2010
DOI: 10.1002/ART.27370
Optimization of the MHC Class I Peptide Cargo Is Dependent on Tapasin
Publisher: Elsevier BV
Date: 04-2002
DOI: 10.1016/S1074-7613(02)00304-7
Abstract: The loading of MHC class I molecules with their peptide cargo is undertaken by a multimolecular peptide loading complex within the endoplasmic reticulum. We show that MHC class I molecules can optimize their peptide repertoire over time and that this process is dependent on tapasin. Optimization of the peptide repertoire is both quantitatively and qualitatively improved by tapasin. The extent of optimization is maximal when MHC class I molecules are allowed to load within the fully assembled peptide loading complex. Finally, we identify a single natural polymorphism (116D>Y) in HLA-B*4402 that permits tapasin-independent loading of HLA-B*4405 (116Y). In the presence of tapasin, the tapasin-independent allele B*4405 (116Y) acquires a repertoire of peptides that is less optimal than the tapasin-dependent allele B*4402 (116D).
Supplementary Table S6 from Spliced Peptides and Cytokine-Driven Changes in the Immunopeptidome of Melanoma
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/2326-6066.22542549.V1
Abstract: Table S6
Predisposition to abacavir hypersensitivity conferred by HLA-B*5701 and a haplotypic Hsp70-Hom variant
Publisher: Proceedings of the National Academy of Sciences
Date: 15-03-2004
Abstract: Susceptibility to a clinically significant drug hypersensitivity syndrome associated with abacavir use seems to have a strong genetic component. We have previously shown that the presence of HLA-B * 5701 strongly predicts abacavir hypersensitivity and have identified a potential susceptibility locus within a 300-kb region between the MEGT1 and C4A6 loci in the central MHC. We now report the results of fine recombinant genetic mapping in an expanded patient population of 248 consecutive, fully ascertained, abacavir-exposed in iduals in the Western Australian HIV Cohort Study, in which 18 cases of definite abacavir hypersensitivity (7.3%) and 230 tolerant controls were identified. Haplotype mapping within patients with allelic markers of the 57.1 ancestral haplotype suggests a susceptibility locus within the 14-kb Hsp70 gene cluster. HLA-B * 5701 was present in 94.4% of hypersensitive cases compared with 1.7% of controls (odds ratio, 960 P 0.00001). A haplotypic nonsynonymous polymorphism of Hsp70-Hom ( HspA1L , resulting from the substitution of residue M493T in the peptide-binding subunit) was found in combination with HLA-B * 5701 in 94.4% of hypersensitive cases and 0.4% of controls (odds ratio, 3,893 P 0.00001). In iduals with abacavir hypersensitivity demonstrated increased monocyte tumor necrosis factor expression in response to ex vivo abacavir stimulation, which was abrogated with CD8 + T cell depletion. These data indicate that the concurrence of HLA-B * 5701 and Hsp70-Hom M493T alleles is necessary for the development of abacavir hypersensitivity, which is likely to be mediated by an HLA-B * 5701 -restricted immune response to abacavir.
New Approaches to Investigate Drug-Induced Hypersensitivity.
Publisher: American Chemical Society (ACS)
Date: 15-11-2017
DOI: 10.1021/ACS.CHEMRESTOX.6B00333
Abstract: The workshop on "New Approaches to Investigate Drug-Induced Hypersensitivity" was held on June 5, 2014 at the Foresight Center, University of Liverpool. The aims of the workshop were to (1) discuss our current understanding of the genetic, clinical, and chemical basis of small molecule drug hypersensitivity, (2) highlight the current status of assays that might be developed to predict potential drug immunogenicity, and (3) identify the limitations, knowledge gaps, and challenges that limit the use of these assays and utilize the knowledge gained from the workshop to develop a pathway to establish new and improved assays that better predict drug-induced hypersensitivity reactions during the early stages of drug development. This perspective reviews the clinical and immunological bases of drug hypersensitivity and summarizes various experts' views on the different topics covered during the meeting.
β-Amino acid-containing hybrid peptides—new opportunities in peptidomimetics
Publisher: Royal Society of Chemistry (RSC)
Date: 2007
DOI: 10.1039/B708507A
Abstract: Hybrid peptides consisting of alpha-amino acids with judiciously placed beta-amino acids show great promise as peptidomimetics in an increasing range of therapeutic applications. This reflects a combination of increased stability, high specificity and relative ease of synthesis.
The production and purification of the human T-cell receptors, the CD3∊γ and CD3∊δ heterodimers: complex formation and crystallization with OKT3, a therapeutic monoclonal antibody
Publisher: International Union of Crystallography (IUCr)
Date: 21-07-2004
Assembly of the secretion pores GspD , W za and CsgG into bacterial outer membranes does not require the O mp85 proteins BamA or TamA
Publisher: Wiley
Date: 06-06-2015
DOI: 10.1111/MMI.13055
Abstract: In Gram-negative bacteria, β-barrel proteins are integrated into the outer membrane by the β-barrel assembly machinery, with key components of the machinery being the Omp85 family members BamA and TamA. Recent crystal structures and cryo-electron microscopy show a erse set of secretion pores in Gram-negative bacteria, with α-helix (Wza and GspD) or β-strand (CsgG) transmembrane segments in the outer membrane. We developed assays to measure the assembly of three distinct secretion pores that mediate protein (GspD), curli fibre (CsgG) and capsular polysaccharide (Wza) secretion by bacteria and show that depletion of BamA and TamA does not diminish the assembly of Wza, GspD or CsgG. Like the well characterised pilotins for GspD and other secretins, small periplasmic proteins enhance the assembly of the CsgG β-barrel. We discuss a model for integral protein assembly into the bacterial outer membrane, focusing on the commonalities and differences in the assembly of Wza, GspD and CsgG.
The self-peptide repertoire plays a critical role in transplant tolerance induction
Publisher: American Society for Clinical Investigation
Date: 11-2021
DOI: 10.1172/JCI146771
Kinetics of Antigen Expression and Epitope Presentation during Virus Infection
Publisher: Public Library of Science (PLoS)
Date: 31-01-2013
A Modular BAM Complex in the Outer Membrane of the α-Proteobacterium Caulobacter crescentus
Publisher: Public Library of Science (PLoS)
Date: 08-01-2010
Constraints within major histocompatibility complex class I restricted peptides: presentation and consequences for T-cell recognition
Publisher: Proceedings of the National Academy of Sciences
Date: 08-03-2010
Abstract: Residues within processed protein fragments bound to major histocompatibility complex class I (MHC-I) glycoproteins have been considered to function as a series of “independent pegs” that either anchor the peptide (p) to the MHC-I and/or interact with the spectrum of αβ-T-cell receptors (TCRs) specific for the pMHC-I epitope in question. Mining of the extensive pMHC-I structural database established that many self- and viral peptides show extensive and direct interresidue interactions, an unexpected finding that has led us to the idea of “constrained” peptides. Mutational analysis of two constrained peptides (the HLA B44 restricted self-peptide (B44DPα–EEFGRAFSF) and an H2-D b restricted influenza peptide (D b PA, SSLENFRAYV) demonstrated that the conformation of the prominently exposed arginine in both peptides was governed by interactions with MHC-I-orientated flanking residues from the peptide itself. Using reverse genetics in a murine influenza model, we revealed that mutation of an MHC-I-orientated residue (SSLEN F RAYV → SSLEN A RAYV) within the constrained PA peptide resulted in a diminished cytotoxic T lymphocyte (CTL) response and the recruitment of a limited pMHC-I specific TCR repertoire. Interactions between in idual peptide positions can thus impose fine control on the conformation of pMHC-I epitopes, whereas the perturbation of such constraints can lead to a previously unappreciated mechanism of viral escape.
Spreading of the immune response from 52 kDaRo and 60 kDaRo to calreticulin in experimental autoimmunity
Publisher: SAGE Publications
Date: 1998
DOI: 10.1191/096120398678919606
Abstract: Calreticulin (CR) is widely recognized as a new human autoantigen but there are conflicting data concerning its relationship with the Ro(SS-A) ribonucleoprotein (RNP). Recent evidence suggests that CR binds to 52 kDaRo (Ro52) by a protein rotein interaction and binds to hY RNA and rubella virus RNA. Other studies have shown that initiation of immunity to either Ro52 or 60 kDaRo (Ro60) can lead to reciprocal spreading of autoimmunity to Ro60 or Ro52, respectively, and induce anti-La autoantibodies in some strains of mice. These findings support a physical association of these polypeptides in Ro/La complexes. To test the hypothesis that CR is physically associated with Ro52 and/or Ro60 we examined the sera of Ro52-, Ro60 and La-immunized mice for intermolecular spreading to CR. Immune sera from BALB/c and C3H/HeJ mice immunized with recombinant 6xHis-mouse Ro52, 6xHis-human Ro60 or 6xHis-human La were tested for reactivity by ELISA and immunoblotting with a full-length human CR protein expressed as a soluble maltose binding protein fusion protein (CR-MBP). Five of the six Ro52-immunized C3H/HeJ mice sera and all six Ro60-immunized C3H/HeJ mice sera reacted with the CR-MBP (but not a MBP control) on ELISA. In the BALB/c group, the responder rate was lower with one in six of the Ro52-immunized and one in five of the Ro60-immunized mice spreading to CR. In contrast, none of the BALB/corC3H/HeJ mice which was immunized with La showed evidence of a recruited anti-CR antibody response. Immunoblotting of the different recombinant proteins with immune sera from the C3H/HeJ mice confirmed the specificity of the initial and recruited antibody responses. The spreading of immunity from Ro52 and Ro60 to CR in Ro-immunized mice suggests that a subpopulation of CR or CR-like molecules must associate under certain circumstances with Ro52 and Ro60 polypeptides in vivo, possibly as Ro/CR complexes concentrated in surface membrane blebs of apoptotic cells. The lack of spreading to CR in Laimmunized mice suggests that CR may be associated with a subpopulation of Ro particles from which La has already dissociated.
Discovery of an archetypal protein transport system in bacterial outer membranes
Publisher: Springer Science and Business Media LLC
Date: 04-2012
DOI: 10.1038/NSMB.2261
Abstract: Bacteria have mechanisms to export proteins for erse purposes, including colonization of hosts and pathogenesis. A small number of archetypal bacterial secretion machines have been found in several groups of bacteria and mediate a fundamentally distinct secretion process. Perhaps erroneously, proteins called 'autotransporters' have long been thought to be one of these protein secretion systems. Mounting evidence suggests that autotransporters might be substrates to be secreted, not an autonomous transporter system. We have discovered a new translocation and assembly module (TAM) that promotes efficient secretion of autotransporters in proteobacteria. Functional analysis of the TAM in Citrobacter rodentium, Salmonella enterica and Escherichia coli showed that it consists of an Omp85-family protein, TamA, in the outer membrane and TamB in the inner membrane of erse bacterial species. The discovery of the TAM provides a new target for the development of therapies to inhibit colonization by bacterial pathogens.
Immune self-reactivity triggered by drug-modified HLA-peptide repertoire
Publisher: Springer Science and Business Media LLC
Date: 23-05-2012
DOI: 10.1038/NATURE11147
Abstract: Human leukocyte antigens (HLAs) are highly polymorphic proteins that initiate immunity by presenting pathogen-derived peptides to T cells. HLA polymorphisms mostly map to the antigen-binding cleft, thereby ersifying the repertoire of self-derived and pathogen-derived peptide antigens selected by different HLA allotypes. A growing number of immunologically based drug reactions, including abacavir hypersensitivity syndrome (AHS) and carbamazepine-induced Stevens-Johnson syndrome (SJS), are associated with specific HLA alleles. However, little is known about the underlying mechanisms of these associations, including AHS, a prototypical HLA-associated drug reaction occurring exclusively in in iduals with the common histocompatibility allele HLA-B*57:01, and with a relative risk of more than 1,000 (refs 6, 7). We show that unmodified abacavir binds non-covalently to HLA-B*57:01, lying across the bottom of the antigen-binding cleft and reaching into the F-pocket, where a carboxy-terminal tryptophan typically anchors peptides bound to HLA-B*57:01. Abacavir binds with exquisite specificity to HLA-B*57:01, changing the shape and chemistry of the antigen-binding cleft, thereby altering the repertoire of endogenous peptides that can bind HLA-B*57:01. In this way, abacavir guides the selection of new endogenous peptides, inducing a marked alteration in 'immunological self'. The resultant peptide-centric 'altered self' activates abacavir-specific T-cells, thereby driving polyclonal CD8 T-cell activation and a systemic reaction manifesting as AHS. We also show that carbamazepine, a widely used anti-epileptic drug associated with hypersensitivity reactions in HLA-B*15:02 in iduals, binds to this allotype, producing alterations in the repertoire of presented self peptides. Our findings simultaneously highlight the importance of HLA polymorphism in the evolution of pharmacogenomics and provide a general mechanism for some of the growing number of HLA-linked hypersensitivities that involve small-molecule drugs.
The enantioselective total synthesis and unambiguous proof of the absolute stereochemistry of pervilleine C
Publisher: Georg Thieme Verlag KG
Date: 08-2008
Experimental autoimmune encephalomyelitis (EAE) IN C57Bl/6 mice is not associated with astrogliosis
Publisher: Elsevier BV
Date: 03-2011
DOI: 10.1016/J.JNEUROIM.2010.10.006
Abstract: The C57Bl/6 mouse is the preferred host for the maintenance of gene deletion mutants and holds a unique place in investigations of cytokine/chemokine networks in neuroinflammation. It is also susceptible to experimental autoimmune encephalomyelitis (EAE), a multiple sclerosis (MS)-like disease commonly used to assess potential MS therapies. Investigations of glial reactivity in EAE have revealed hitherto undescribed astroglial responses in this model, characterized by progressively diminishing glial fibrillary acidic protein and aquaporin-4 immunostaining, from early disease. These observations show that astrocyte responses vary with the EAE paradigm and are an important pathological criterion for disease mapping and therapy evaluation.
The complexity of T cell–mediated penicillin hypersensitivity reactions
Publisher: Wiley
Date: 06-07-2020
DOI: 10.1111/ALL.14355
Abstract: Penicillin refers to a group of beta‐lactam antibiotics that are the first‐line treatment for a range of infections. However, they also possess the ability to form novel antigens, or neoantigens, through haptenation of proteins and can stimulate a range of immune‐mediated adverse reactions—collectively known as drug hypersensitivity reactions (DHRs). IgE‐mediated reactions towards these neoantigens are well studied however, IgE‐independent reactions are less well understood. These reactions usually manifest in a delayed manner as different forms of cutaneous eruptions or liver injury consistent with priming of an immune response. Ex vivo studies have confirmed the infiltration of T cells into the site of inflammation, and the subsets of T cells involved appear dependent on the nature of the reaction. Here, we review the evidence that has led to our current understanding of these immune‐mediated reactions, discussing the nature of the lesional T cells, the characterization of drug‐responsive T cells isolated from patient blood, and the potential mechanisms by which penicillins enter the antigen processing and presentation pathway to stimulate these deleterious responses. Thus, we highlight the need for a more comprehensive understanding of the underlying genetic and molecular basis of penicillin‐induced DHRs.
The Accessibility of the Prosthetic Group Biotin during Monomeric Avidin Affinity Chromatography
Publisher: Elsevier BV
Date: 07-1996
Abstract: Valvular heart disease (VHD) is a common heart disease that affects blood flow. It usually requires heart surgery. Valvular heart disease complicated with pulmonary artery hypertension (VHD-PAH) may be lethal due to heart failure that results from increased heart burden. It is important for these patients to seek early treatment in order to minimize the heart damage. However, there is no reliable diagnosis method in VHD. In this study, we found DNA methylation was increased at the promoter of
Mitigating Human IAPP Amyloidogenesis In Vivo with Chiral Silica Nanoribbons
Publisher: Wiley
Date: 07-10-2018
Quantification of epitope abundance reveals the effect of direct and cross-presentation on influenza CTL responses
Publisher: Springer Science and Business Media LLC
Date: 28-06-2019
DOI: 10.1038/S41467-019-10661-8
Abstract: The magnitude of T cell responses to infection is a function of the naïve T cell repertoire combined with the context and duration of antigen presentation. Using mass spectrometry, we identify and quantify 21 class 1 MHC-restricted influenza A virus (IAV)-peptides following either direct or cross-presentation. All these peptides, including seven novel epitopes, elicit T cell responses in infected C57BL/6 mice. Directly presented IAV epitopes maintain their relative abundance across distinct cell types and reveal a broad range of epitope abundances. In contrast, cross-presented epitopes are more uniform in abundance. We observe a clear disparity in the abundance of the two key immunodominant IAV antigens, wherein direct infection drives optimal nucleoprotein (NP) 366–374 presentation, while cross-presentation is optimal for acid polymerase (PA) 224–233 presentation. The study demonstrates how assessment of epitope abundance in both modes of antigen presentation is necessary to fully understand the immunogenicity and response magnitude to T cell epitopes.
Anthelmintic closantel enhances bacterial killing of polymyxin B against multidrug-resistant Acinetobacter baumannii
Publisher: Springer Science and Business Media LLC
Date: 16-12-2015
DOI: 10.1038/JA.2015.127
Noncontiguous T cell epitopes in autoimmune diabetes: From mice to men and back again
Publisher: Elsevier BV
Date: 07-2021
Nonprecipitating anti-La(SS-B) autoantibodies in primary Sjogren's syndrome
Publisher: Elsevier BV
Date: 06-1996
Abstract: Anti-La(SS-B) precipitin-negative sera show a restricted epitope recognition and can be easily overlooked in routine laboratory testing. We have therefore determined the prevalence of nonprecipitating anti-La(SS-B) antibodies in patients with primary Sjögren's syndrome and studied their clinical and immunological associations. Clinical details were obtained from 68 patients with primary Sjögren's syndrome, and serum s les were examined by enzyme-linked immunosorbent assay using purified recombinant La, 60-kDa Ro, and 52-kDa Ro proteins and by counterimmunoelectrophoresis. Thirteen patients (19%) were identified with anti-La antibodies which were nonprecipitating. These patients had similar clinical findings to other groups of patients with Sjogren's syndrome, but had significantly lower rheumatoid factor and serum IgG levels than patients with anti-La precipitins. None of the patients with nonprecipitating anti-La antibodies had previously contained anti-La precipitins in their sera. Furthermore, they tended to have lower levels of antibodies directed against denatured 60-kDa Ro (but not 52-kDa Ro) compared with anti-La precipitin-positive patients. Patients with Sjögren's syndrome associated with nonprecipitating anti-La antibodies represent a stable serological and clinical subset in which there appears to be limited ersification of the autoimmune response to the Ro60 and La proteins of the Ro/La ribonucleoprotein.
Molecular markers of preterm labor in the choriodecidua
Publisher: Springer Science and Business Media LLC
Date: 03-2010
Data from Spliced Peptides and Cytokine-Driven Changes in the Immunopeptidome of Melanoma
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/2326-6066.C.6549978.V1
Abstract: Abstract Antigen recognition by CD8 sup + /sup T cells is governed by the pool of peptide antigens presented on the cell surface in the context of HLA class I complexes. Studies have shown not only a high degree of plasticity in the immunopeptidome, but also that a considerable fraction of all presented peptides is generated through proteasome-mediated splicing of noncontiguous regions of proteins to form novel peptide antigens. Here, we used high-resolution mass spectrometry combined with new bioinformatic approaches to characterize the immunopeptidome of melanoma cells in the presence or absence of IFNγ. In total, we identified more than 60,000 peptides from a single patient-derived cell line (LM-MEL-44) and demonstrated that IFNγ induced changes in the peptidome, with an overlap of only approximately 50% between basal and treated cells. Around 6% to 8% of the peptides were identified as i cis /i -spliced peptides, and 2,213 peptides (1,827 linear and 386 i cis /i -spliced peptides) were derived from known melanoma-associated antigens. These peptide antigens were equally distributed between the constitutive- and IFNγ-induced peptidome. We next examined additional HLA-matched patient-derived cell lines to investigate how frequently these peptides were identified and found that a high proportion of both linear and spliced peptides was conserved between in idual patient tumors, drawing on data amassing to more than 100,000 peptide sequences. Several of these peptides showed i in vitro /i immunogenicity across multiple patients with melanoma. These observations highlight the breadth and complexity of the repertoire of immunogenic peptides that can be exploited therapeutically and suggest that spliced peptides are a major class of tumor antigens. /
A comprehensive analysis of constitutive naturally processed and presented HLA ‐C*04:01 (Cw4) – specific peptides
Publisher: Wiley
Date: 07-01-2014
DOI: 10.1111/TAN.12282
Abstract: The human B lymphoblastoid cell line C1R is widely regarded as human leukocyte antigen-A (HLA-A)/HLA-B negative and is therefore frequently exploited as a recipient cell line to study HLA class I functions. However, the normal levels of HLA-C*04:01 often h er the investigation of introduced HLA class I allomorphs, which is particularly evident in sensitive applications such as mass spectrometry. Here we describe the comprehensive analysis of endogenous HLA-C*04:01 ligands expressed on the surface of C1R cells to (i) define a large sequence dataset of HLA-C*04:01 ligands, to (ii) refine the HLA-C*04:01 peptide-binding motif and (iii) to provide a resource that allows discrimination between peptides bound to introduced HLA class I subtypes and to the endogenous HLA-C*04:01 molecules.
Quokka: a comprehensive tool for rapid and accurate prediction of kinase family-specific phosphorylation sites in the human proteome
Publisher: Oxford University Press (OUP)
Date: 27-06-2018
DOI: 10.1093/BIOINFORMATICS/BTY522
Abstract: Kinase-regulated phosphorylation is a ubiquitous type of post-translational modification (PTM) in both eukaryotic and prokaryotic cells. Phosphorylation plays fundamental roles in many signalling pathways and biological processes, such as protein degradation and protein-protein interactions. Experimental studies have revealed that signalling defects caused by aberrant phosphorylation are highly associated with a variety of human diseases, especially cancers. In light of this, a number of computational methods aiming to accurately predict protein kinase family-specific or kinase-specific phosphorylation sites have been established, thereby facilitating phosphoproteomic data analysis. In this work, we present Quokka, a novel bioinformatics tool that allows users to rapidly and accurately identify human kinase family-regulated phosphorylation sites. Quokka was developed by using a variety of sequence scoring functions combined with an optimized logistic regression algorithm. We evaluated Quokka based on well-prepared up-to-date benchmark and independent test datasets, curated from the Phospho.ELM and UniProt databases, respectively. The independent test demonstrates that Quokka improves the prediction performance compared with state-of-the-art computational tools for phosphorylation prediction. In summary, our tool provides users with high-quality predicted human phosphorylation sites for hypothesis generation and biological validation. The Quokka webserver and datasets are freely available at Supplementary data are available at Bioinformatics online.
HIVed, a knowledgebase for differentially expressed human genes and proteins during HIV infection, replication and latency
Publisher: Springer Science and Business Media LLC
Date: 30-04-2917
DOI: 10.1038/SREP45509
Abstract: Measuring the altered gene expression level and identifying differentially expressed genes roteins during HIV infection, replication and latency is fundamental for broadening our understanding of the mechanisms of HIV infection and T-cell dysfunction. Such studies are crucial for developing effective strategies for virus eradication from the body. Inspired by the availability and enrichment of gene expression data during HIV infection, replication and latency, in this study, we proposed a novel compendium termed HIVed (HIV expression database hivlatency.erc.monash.edu/ ) that harbours comprehensive functional annotations of proteins, whose genes have been shown to be dysregulated during HIV infection, replication and latency using different experimental designs and measurements. We manually curated a variety of third-party databases for structural and functional annotations of the protein entries in HIVed. With the goal of benefiting HIV related research, we collected a number of biological annotations for all the entries in HIVed besides their expression profile, including basic protein information, Gene Ontology terms, secondary structure, HIV-1 interaction and pathway information. We hope this comprehensive protein-centric knowledgebase can bridge the gap between the understanding of differentially expressed genes and the functions of their protein products, facilitating the generation of novel hypotheses and treatment strategies to fight against the HIV pandemic.
Assembly of the type II secretion system such as found in Vibrio cholerae depends on the novel Pilotin AspS.
Publisher: Public Library of Science (PLoS)
Date: 10-01-2013
Allelic polymorphism in the T cell receptor and its impact on immune responses
Publisher: Rockefeller University Press
Date: 21-06-2010
DOI: 10.1084/JEM.20100603
Abstract: In comparison to human leukocyte antigen (HLA) polymorphism, the impact of allelic sequence variation within T cell receptor (TCR) loci is much less understood. Particular TCR loci have been associated with autoimmunity, but the molecular basis for this phenomenon is undefined. We examined the T cell response to an HLA-B*3501–restricted epitope (HPVGEADYFEY) from Epstein-Barr virus (EBV), which is frequently dominated by a TRBV9*01+ public TCR (TK3). However, the common allelic variant TRBV9*02, which differs by a single amino acid near the CDR2β loop (Gln55→His55), was never used in this response. The structure of the TK3 TCR, its allelic variant, and a nonnaturally occurring mutant (Gln55→Ala55) in complex with HLA-B*3501HPVGEADYFEY revealed that the Gln55→His55 polymorphism affected the charge complementarity at the TCR–peptide-MHC interface, resulting in reduced functional recognition of the cognate and naturally occurring variants of this EBV peptide. Thus, polymorphism in the TCR loci may contribute toward variability in immune responses and the outcome of infection.
HLA-class II specificity assessed by high-density peptide microarray interactions
Publisher: Cold Spring Harbor Laboratory
Date: 02-03-2020
DOI: 10.1101/2020.02.28.969667
Abstract: The ability to predict and/or identify MHC binding peptides is an essential component of T cell epitope discovery something that ultimately should benefit the development of vaccines and immunotherapies. In particular, MHC class I (MHC-I) prediction tools have matured to a point where accurate selection of optimal peptide epitopes is possible for virtually all MHC-I allotypes in comparison, current MHC class II (MHC-II) predictors are less mature. Since MHC-II restricted CD4+ T cells control and orchestrate most immune responses, this shortcoming severely h ers the development of effective immunotherapies. The ability to generate large panels of peptides and subsequently large bodies of peptide-MHC-II interaction data is key to the solution of this problem a solution that also will support the improvement of bioinformatics predictors, which critically relies on the availability of large amounts of accurate, erse and representative data. Here, we have used recombinant HLA-DRB1*01:01 and HLA-DRB1*03:01 molecules to interrogate high-density peptide arrays, in casu containing 70,000 random peptides in triplicates. We demonstrate that the binding data acquired contains systematic and interpretable information reflecting the specificity of the HLA-DR molecules investigated. Collectively, with a cost per peptide reduced to a few cents combined with the flexibility of recombinant HLA technology, this poses an attractive strategy to generate vast bodies of MHC-II binding data at an unprecedented speed and for the benefit of generating peptide-MHC-II binding data as well as improving MHC-II prediction tools.
A Charged Amino Acid Residue in the Transmembrane/Cytoplasmic Region of Tapasin Influences MHC Class I Assembly and Maturation
Publisher: The American Association of Immunologists
Date: 15-01-2005
DOI: 10.4049/JIMMUNOL.174.2.962
Abstract: Tapasin influences the quantity and quality of MHC eptide complexes at the cell surface however, little is understood about the structural features that underlie its effects. Because tapasin, MHC class I, and TAP are transmembrane proteins, the tapasin transmembrane/cytoplasmic region has the potential to affect interactions at the endoplasmic reticulum membrane. In this study, we have assessed the influence of a conserved lysine at position 408, which lies in the tapasin transmembrane/cytoplasmic domain. We found that substitutions at position K408 in tapasin affected the expression of MHC class I molecules at the cell surface, and down-regulated tapasin stabilization of TAP. In addition to affecting TAP interaction with tapasin, the substitution of alanine, but not tryptophan, for the lysine at tapasin position 408 increased the amount of tapasin found in association with the open, peptide-free form of the HLA-B8 H chain. Tapasin K408A was also associated with more folded, β2-microglobulin-assembled HLA-B8 molecules than wild-type tapasin. Consistent with our observation of a large pool of tapasin K408A-associated HLA-B8 molecules, the rate at which HLA-B8 migrated from the endoplasmic reticulum was slower in tapasin K408A-expressing cells than in wild-type tapasin-expressing cells. Thus, the alanine substitution at position 408 in tapasin may interfere with the stable acquisition by MHC class I molecules of peptides that are sufficiently optimal to allow MHC class I release from tapasin.
The molecular basis of how buried human leukocyte antigen polymorphism modulates natural killer cell function
Publisher: Proceedings of the National Academy of Sciences
Date: 13-05-2020
Abstract: Micropolymorphisms within human leukocyte antigen (HLA) class I molecules can change the architecture of the peptide-binding cleft, leading to differences in peptide presentation and T cell recognition. The impact of such HLA variation on natural killer (NK) cell recognition remains unclear. Given the differential association of HLA-B*57:01 and HLA-B*57:03 with the control of HIV, recognition of these HLA-B57 allomorphs by the killer cell immunoglobulin-like receptor (KIR) 3DL1 was compared. Despite differing by only two polymorphic residues, both buried within the peptide-binding cleft, HLA-B*57:01 more potently inhibited NK cell activation. Direct-binding studies showed KIR3DL1 to preferentially recognize HLA-B*57:01, particularly when presenting peptides with positively charged position (P)Ω-2 residues. In HLA-B*57:01, charged PΩ-2 residues were oriented toward the peptide-binding cleft and away from KIR3DL1. In HLA-B*57:03, the charged PΩ-2 residues protruded out from the cleft and directly impacted KIR3DL1 engagement. Accordingly, KIR3DL1 recognition of HLA class I ligands is modulated by both the peptide sequence and conformation, as determined by the HLA polymorphic framework, providing a rationale for understanding differences in clinical associations.
The Evolving Landscape of Autoantigen Discovery and Characterization in Type 1 Diabetes
Publisher: American Diabetes Association
Date: 16-04-2019
DOI: 10.2337/DBI18-0066
Abstract: Type 1 diabetes (T1D) is an autoimmune disease that is caused, in part, by T cell–mediated destruction of insulin-producing β-cells. High risk for disease, in those with genetic susceptibility, is predicted by the presence of two or more autoantibodies against insulin, the 65-kDa form of glutamic acid decarboxylase (GAD65), insulinoma-associated protein 2 (IA-2), and zinc transporter 8 (ZnT8). Despite this knowledge, we still do not know what leads to the breakdown of tolerance to these autoantigens, and we have an incomplete understanding of T1D etiology and pathophysiology. Several new autoantibodies have recently been discovered using innovative technologies, but neither their potential utility in monitoring disease development and treatment nor their role in the pathophysiology and etiology of T1D has been explored. Moreover, neoantigen generation (through posttranslational modification, the formation of hybrid peptides containing two distinct regions of an antigen or antigens, alternative open reading frame usage, and translation of RNA splicing variants) has been reported, and autoreactive T cells that target these neoantigens have been identified. Collectively, these new studies provide a conceptual framework to understand the breakdown of self-tolerance, if such modifications occur in a tissue- or disease-specific context. A recent workshop sponsored by the National Institute of Diabetes and Digestive and Kidney Diseases brought together investigators who are using new methods and technologies to identify autoantigens and characterize immune responses toward these proteins. Researchers with erse expertise shared ideas and identified resources to accelerate antigen discovery and the detection of autoimmune responses in T1D. The application of this knowledge will direct strategies for the identification of improved biomarkers for disease progression and treatment response monitoring and, ultimately, will form the foundation for novel antigen-specific therapeutics. This Perspective highlights the key issues that were addressed at the workshop and identifies areas for future investigation.
Epitope-specific TCRbeta repertoire diversity imparts no functional advantage on the CD8+ T cell response to cognate viral peptides.
Publisher: Proceedings of the National Academy of Sciences
Date: 12-02-2008
Abstract: TCR repertoire ersity has been convincingly shown to facilitate responsiveness of CD8 + T cell populations to mutant virus peptides, thereby safeguarding against viral escape. However, the impact of repertoire ersity on the functionality of the CD8 + T cell response to cognate peptide-MHC class I complex (pMHC) recognition remains unclear. Here, we have compared TCRβ chain repertoires of three influenza A epitope-specific CD8 + T cell responses in C57BL/6 (B6) mice: D b NP 366–374 , D b PA 224–233 , and a recently described epitope derived from the +1 reading frame of the influenza viral polymerase B subunit (residues 62–70) (D b PB1-F2 62 ). Corresponding to the relative antigenicity of the respective pMHCs, and irrespective of the location of prominent residues, the D b PA 224 - and D b PB1-F2 62 -specific repertoires were similarly erse, whereas the D b NP 366 population was substantially narrower. Importantly, parallel analysis of response magnitude, cytotoxicity, TCR avidity, and cytokine production for the three epitope-specific responses revealed no obvious functional advantage conferred by increased T cell repertoire ersity. Thus, whereas a erse repertoire may be important for recognition of epitope variants, its effect on the response to cognate pMHC recognition appears minimal.
Preemptive priming readily overcomes structure-based mechanisms of virus escape
Publisher: Proceedings of the National Academy of Sciences
Date: 14-03-2013
Abstract: A reverse-genetics approach has been used to probe the mechanism underlying immune escape for influenza A virus-specific CD8 + T cells responding to the immunodominant D b NP 366 epitope. Engineered viruses with a substitution at a critical residue (position 6, P6M) all evaded recognition by WT D b NP 366 -specific CD8 + T cells, but only the NPM6I and NPM6T mutants altered the topography of a key residue (His155) in the MHC class I binding site. Following infection with the engineered NPM6I and NPM6T influenza viruses, both mutations were associated with a substantial “hole” in the naïve T-cell receptor repertoire, characterized by very limited T-cell receptor ersity and minimal primary responses to the NPM6I and NPM6T epitopes. Surprisingly, following respiratory challenge with a serologically distinct influenza virus carrying the same mutation, preemptive immunization against these escape variants led to the generation of secondary CD8 + T-cell responses that were comparable in magnitude to those found for the WT NP epitope. Consequently, it might be possible to generate broadly protective T-cell immunity against commonly occurring virus escape mutants. If this is generally true for RNA viruses (like HIV, hepatitis C virus, and influenza) that show high mutation rates, priming against predicted mutants before an initial encounter could function to prevent the emergence of escape variants in infected hosts. That process could be a step toward preserving immune control of particularly persistent RNA viruses and may be worth considering for future vaccine strategies.
β2 -glycoprotein I and protection from anti-SSA/Ro60- associated cardiac manifestations of neonatal lupus
Publisher: The American Association of Immunologists
Date: 07-2011
Abstract: One mechanism to molecularly explain the strong association of maternal anti-Ro60 Abs with cardiac disease in neonatal lupus (NL) is that these Abs initiate injury by binding to apoptotic cardiomyocytes in the fetal heart. Previous studies have demonstrated that β2-glycoprotein I (β2GPI) interacts with Ro60 on the surface of apoptotic Jurkat cells and prevents binding of anti-Ro60 IgG. Accordingly, the current study was initiated to test two complementary hypotheses, as follows: 1) competition between β2GPI and maternal anti-Ro60 Abs for binding apoptotic induced surface-translocated Ro60 occurs on human fetal cardiomyocytes and 2) circulating levels of β2GPI influence injury in anti-Ro60–exposed fetuses. Initial flow cytometry experiments conducted on apoptotic human fetal cardiomyocytes demonstrated dose-dependent binding of β2GPI. In competitive inhibition experiments, β2GPI prevented opsonization of apoptotic cardiomyocytes by maternal anti-Ro60 IgG. ELISA was used to quantify β2GPI in umbilical cord blood from 97 neonates exposed to anti-Ro60 Abs, 53 with cardiac NL and 44 with no cardiac disease. β2GPI levels were significantly lower in neonates with cardiac NL. Plasmin-mediated cleavage of β2GPI prevented binding to Ro60 and promoted the formation of pathogenic anti-Ro60 IgG-apoptotic cardiomyocyte complexes. In aggregate these data suggest that intact β2GPI in the fetal circulation may be a novel cardioprotective factor in anti-Ro60–exposed pregnancies.
Use of proteomics to define targets of T-cell immunity
Publisher: Informa UK Limited
Date: 06-2005
Abstract: The mammalian immune system has evolved to display peptides derived from microbial antigens to immune effector cells. Liberated from the intact antigens through distinct proteolytic mechanisms, these peptides are subsequently transported to the cell surface while bound to chaperone-like receptors known as major histocompatibility complex molecules. These complexes are then scrutinized by T-cells that express receptors with specificity for specific major histocompatibility complex-peptide complexes. In normal uninfected cells, this process of antigen processing and presentation occurs continuously, with the resultant array of self-antigen-derived peptides displayed on the surface of these cells. Changes in this cellular peptide array alert the immune system to changes in the intracellular environment that may be associated with infection, oncogenesis or other abnormal cellular processes, resulting in a cascade of events that result in the elimination of the abnormal cell. Since peptides play such an essential role in informing the immune system of infection with viral or microbial pathogens and the transformation of cells in malignancy, the tools of proteomics, in particular mass spectrometry, are ideally suited to study these immune responses at a molecular level. Recent advances in studies of immune responses that have utilized mass spectrometry and associated technologies are reviewed. The authors gaze into the future and look at current challenges and where proteomics will impact in immunology over the next 5 years.
Proinsulin-Specific, HLA-DQ8, and HLA-DQ8-Transdimer–Restricted CD4+ T Cells Infiltrate Islets in Type 1 Diabetes
Publisher: American Diabetes Association
Date: 25-08-2014
DOI: 10.2337/DB14-0858
Abstract: Type 1 diabetes (T1D) develops when insulin-secreting β-cells, found in the pancreatic islets of Langerhans, are destroyed by infiltrating T cells. How human T cells recognize β-cell-derived antigens remains unclear. Genetic studies have shown that HLA and insulin alleles are the most strongly associated with risk of T1D. These long-standing observations implicate CD4+ T-cell responses against (pro)insulin in the pathogenesis of T1D. To dissect the autoimmune T-cell response against human β-cells, we isolated and characterized 53 CD4+ T-cell clones from within the residual pancreatic islets of a deceased organ donor who had T1D. These 53 clones expressed 47 unique clonotypes, 8 of which encoded proinsulin-specific T-cell receptors. On an in idual clone basis, 14 of 53 CD4+ T-cell clones (26%) recognized 6 distinct but overlapping epitopes in the C-peptide of proinsulin. These clones recognized C-peptide epitopes presented by HLA-DQ8 and, notably, HLA-DQ8 transdimers that form in HLA-DQ2/-DQ8 heterozygous in iduals. Responses to these epitopes were detected in the peripheral blood mononuclear cells of some people with recent-onset T1D but not in HLA-matched control subjects. Hence, proinsulin-specific, HLA-DQ8, and HLA-DQ8-transdimer–restricted CD4+ T cells are strongly implicated in the autoimmune pathogenesis of human T1D.
T cell receptor cross-reactivity between gliadin and bacterial peptides in celiac disease.
Publisher: Springer Science and Business Media LLC
Date: 23-12-2020
DOI: 10.1038/S41594-019-0353-4
Abstract: The human leukocyte antigen (HLA) locus is strongly associated with T cell-mediated autoimmune disorders. HLA-DQ2.5-mediated celiac disease (CeD) is triggered by the ingestion of gluten, although the relative roles of genetic and environmental risk factors in CeD is unclear. Here we identify microbially derived mimics of gliadin epitopes and a parental bacterial protein that is naturally processed by antigen-presenting cells and activated gliadin reactive HLA-DQ2.5-restricted T cells derived from CeD patients. Crystal structures of T cell receptors in complex with HLA-DQ2.5 bound to two distinct bacterial peptides demonstrate that molecular mimicry underpins cross-reactivity toward the gliadin epitopes. Accordingly, gliadin reactive T cells involved in CeD pathogenesis cross-react with ubiquitous bacterial peptides, thereby suggesting microbial exposure as a potential environmental factor in CeD.
Comparison between the isocratic and gradient retention behaviour of polypeptides in reversed-phase liquid chromatographic environments
Publisher: Elsevier BV
Date: 08-1999
DOI: 10.1016/S0021-9673(99)00440-9
Abstract: The isocratic and gradient elution behaviour of beta-endorphin and glucagon, two polypeptides known to exist in hipathic alpha-helical conformations in lipophilic environments, have been examined under reversed-phase high-performance liquid chromatographic (RP-HPLC) conditions with low pH, aquo-acetonitrile mobile phases. The effects of changes in the volume fraction, psi, of the organic solvent modifier and temperature, T, on the magnitudes of the S and log k(o) values of these two polypeptides, obtained from the plots of logarithmic capacity factor (log k') vs. psi using isocratic elution conditions have been determined. These data have then been compared to the corresponding S and log k(o) values, obtained from the plots of logarithmic median capacity factor (log k) versus the median volume fraction of the organic solvent modifier (psi) derived from the linear gradient elution data, using the same n-butyl silica sorbent and related aquo-acetonitrile mobile phase conditions. As apparent from these studies, substantial differences occur in the temperature-dependent trends and magnitudes of the corresponding S and S values, or the log k(o) and log k(o) values, when these parameters are derived from experimental data acquired by these two different elution methods. Moreover, when gradient elution data for beta-endorphin and glucagon are utilised, the extrapolated values of the intercept and slope of the plots of log k vs. 1/T (corresponding to an apparent change in the median enthalpy of association, deltaH(o)assoc, or an apparent change in the median entropy of association, deltaS(o)assoc) substantially deviated from the values obtained for the thermodynamic parameters, deltaH(o)assoc and deltaS(o)assoc, derived from the log k' vs. 1/T plots using the corresponding isocratic data. These findings thus have important implications for biophysical and thermodynamic investigations when gradient elution data are employed to assess the molecular basis of the interaction of polypeptides with non-polar ligates.
Human T-cells recognise N-terminally Fmoc-modified peptide
Publisher: Elsevier BV
Date: 09-2003
DOI: 10.1016/S0264-410X(03)00402-X
Abstract: We aimed to generate T-cell clones specific for human pre-proinsulin. An HLA DQ8, CD4+ T-cell clone that recognised a 10mer (C65-A9) peptide from pre-proinsulin was isolated. Further analysis revealed that the clone responded neither to recombinant proinsulin nor to re-synthesised C65-A9 peptide. Analysis of the original peptide revealed minor contamination (<0.5%) with an N-terminal Fmoc adduct. This peptide was synthesised and shown to stimulate the clone. Thus, Fmoc-modified peptides, which are common contaminants in synthetic peptides, can stimulate human CD4+ T-cells. This finding has important implications for the use of synthetic peptides in screening and epitope mapping studies and their use as vaccines in humans.
The shaping of T cell receptor recognition by self-tolerance
Publisher: Elsevier BV
Date: 02-2009
DOI: 10.1016/J.IMMUNI.2008.11.011
Abstract: During selection of the T cell repertoire, the immune system navigates the subtle distinction between self-restriction and self-tolerance, yet how this is achieved is unclear. Here we describe how self-tolerance toward a trans-HLA (human leukocyte antigen) allotype shapes T cell receptor (TCR) recognition of an Epstein-Barr virus (EBV) determinant (FLRGRAYGL). The recognition of HLA-B8-FLRGRAYGL by two archetypal TCRs was compared. One was a publicly selected TCR, LC13, that is alloreactive with HLA-B44 the other, CF34, lacks HLA-B44 reactivity because it arises when HLA-B44 is coinherited in trans with HLA-B8. Whereas the alloreactive LC13 TCR docked at the C terminus of HLA-B8-FLRGRAYGL, the CF34 TCR docked at the N terminus of HLA-B8-FLRGRAYGL, which coincided with a polymorphic region between HLA-B8 and HLA-B44. The markedly contrasting footprints of the LC13 and CF34 TCRs provided a portrait of how self-tolerance shapes the specificity of TCRs selected into the immune repertoire.
The A-chain of insulin is a hot-spot for CD4 + T cell epitopes in human type 1 diabetes
Publisher: Oxford University Press (OUP)
Date: 11-02-2009
DOI: 10.1111/J.1365-2249.2009.03907.X
Abstract: Type 1 diabetes (T1D) is caused by T cell-mediated destruction of the pancreatic insulin-producing β cells. While the role of CD4+ T cells in the pathogenesis of T1D is accepted widely, the epitopes recognized by pathogenic human CD4+ T cells remain poorly defined. None the less, responses to the N-terminal region of the insulin A-chain have been described. Human CD4+ T cells from the pancreatic lymph nodes of subjects with T1D respond to the first 15 amino acids of the insulin A-chain. We identified a human leucocyte antigen-DR4-restricted epitope comprising the first 13 amino acids of the insulin A-chain (A1-13), dependent upon generation of a vicinal disulphide bond between adjacent cysteines (A6–A7). Here we describe the analysis of a CD4+ T cell clone, isolated from a subject with T1D, which recognizes a new HLR-DR4-restricted epitope (KRGIVEQCCTSICS) that overlaps the insulin A1-13 epitope. This is a novel epitope, because the clone responds to proinsulin but not to insulin, T cell recognition requires the last two residues of the C-peptide (Lys, Arg) and recognition does not depend upon a vicinal disulphide bond between the A6 and A7 cysteines. The finding of a further CD4+ T cell epitope in the N-terminal A-chain region of human insulin underscores the importance of this region as a target of CD4+ T cell responses in human T1D.
The shared susceptibility epitope of HLA-DR4 binds citrullinated self-antigens and the TCR
Publisher: American Association for the Advancement of Science (AAAS)
Date: 02-04-0005
DOI: 10.1126/SCIIMMUNOL.ABE0896
Abstract: Citrullination of a fibrinogen peptide influences HLA-DR4 binding and TCR usage by neoepitope-reactive T cells in rheumatoid arthritis.
T Cell Determinants Incorporating β-Amino Acid Residues Are Protease Resistant and Remain Immunogenic In Vivo
Publisher: The American Association of Immunologists
Date: 15-09-2005
DOI: 10.4049/JIMMUNOL.175.6.3810
Abstract: A major hurdle in designing successful epitope-based vaccines resides in the delivery, stability, and immunogenicity of the peptide immunogen. The short-lived nature of unmodified peptide-based vaccines in vivo limits their therapeutic application in the immunotherapy of cancers and chronic viral infections as well as their use in generating prophylactic immunity. The incorporation of β-amino acids into peptides decreases proteolysis, yet its potential application in the rational design of T cell mimotopes is poorly understood. To address this, we have replaced each residue of the SIINFEKL epitope in idually with the corresponding β-amino acid and examined the resultant efficacy of these mimotopes. Some analogs displayed similar MHC binding and superior protease stability compared with the native epitope. Importantly, these analogs were able to generate cross-reactive CTLs in vivo that were capable of lysing tumor cells that expressed the unmodified epitope as a surrogate tumor Ag. Structural analysis of peptides in which anchor residues were substituted with β-amino acids revealed the basis for enhanced MHC binding and retention of immunogenicity observed for these analogs and paves the way for future vaccine design using β-amino acids. We conclude that the rational incorporation of β-amino acids into T cell determinants is a powerful alternative to the traditional homologous substitution of randomly chosen naturally occurring α-amino acids, and these mimotopes may prove particularly useful for inclusion in epitope-based vaccines.
Functional and Structural Characteristics of NY-ESO-1-related HLA A2-restricted Epitopes and the Design of a Novel Immunogenic Analogue
Publisher: Elsevier BV
Date: 05-2004
Downregulation of MHC class I expression by influenza A and B viruses
Publisher: Frontiers Media SA
Date: 29-05-2019
Supplementary Figures from Spliced Peptides and Cytokine-Driven Changes in the Immunopeptidome of Melanoma
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/2326-6066.22542573.V1
Abstract: Supplementary Figures S1-S10
Conserved Motifs Reveal Details of Ancestry and Structure in the Small TIM Chaperones of the Mitochondrial Intermembrane Space
Publisher: Oxford University Press (OUP)
Date: 13-02-2007
Abstract: The mitochondrial inner and outer membranes are composed of a variety of integral membrane proteins, assembled into the membranes posttranslationally. The small translocase of the inner mitochondrial membranes (TIMs) are a group of approximately 10 kDa proteins that function as chaperones to ferry the imported proteins across the mitochondrial intermembrane space to the outer and inner membranes. In yeast, there are 5 small TIM proteins: Tim8, Tim9, Tim10, Tim12, and Tim13, with equivalent proteins reported in humans. Using hidden Markov models, we find that many eukaryotes have proteins equivalent to the Tim8 and Tim13 and the Tim9 and Tim10 subunits. Some eukaryotes provide "snapshots" of evolution, with a single protein showing the features of both Tim8 and Tim13, suggesting that a single progenitor gene has given rise to each of the small TIMs through duplication and modification. We show that no "Tim12" family of proteins exist, but rather that variant forms of the cognate small TIMs have been recently duplicated and modified to provide new functions: the yeast Tim12 is a modified form of Tim10, whereas in humans and some protists variant forms of Tim9, Tim8, and Tim13 are found instead. Sequence motif analysis reveals acidic residues conserved in the Tim10 substrate-binding tentacles, whereas more hydrophobic residues are found in the equivalent substrate-binding region of Tim13. The substrate-binding region of Tim10 and Tim13 represent structurally independent domains: when the acidic domain from Tim10 is attached to Tim13, the Tim8-Tim13(10) complex becomes essential and the Tim9-Tim10 complex becomes dispensable. The conserved features in the Tim10 and Tim13 subunits provide distinct binding surfaces to accommodate the broad range of substrate proteins delivered to the mitochondrial inner and outer membranes.
Association of stress proteins with autoantigens: a possible mechanism for triggering autoimmunity?
Publisher: Oxford University Press (OUP)
Date: 16-04-2003
DOI: 10.1046/J.1365-2249.2003.02153.X
Abstract: Patterns of autoantibody production are diagnostic of many autoimmune disorders the recent observation of additional autospecificities towards stress-induced proteins may also provide insight into the mechanisms by which such responses arise. Grp78 (also known as BiP) is a target of autoaggressive B and T cell responses in our murine model of anti-Ro (SS-A) autoimmunity and also in rheumatoid arthritis. In this report we demonstrate reciprocal intermolecular spreading occurs between Ro52 and Grp78 in immunized mice, reflecting physiological association of these molecules in vivo. Moreover, we provide direct biochemical evidence that Grp78 associates with the clinically relevant autoantigen, Ro52 (SS-A). Due to the discrete compartmentalization of Ro52 (nucleocytoplasmic) and Grp78 (endoplasmic reticulum ER) we propose that association of these molecules occurs either in apoptotic cells, where they have been demonstrated indirectly to co-localize in discrete apoptotic bodies, or in B cells themselves where both Ro52 and Grp78 are known to bind to immunoglobulin heavy chains. Tagging of molecules by association with Grp78 may facilitate receptor mediated phagocytotsis of the complex we show evidence that exogenous Grp78 can associate with cell surface receptors on a subpopulation of murine splenocytes. Given the likelihood that Grp78 will associate with viral glycoproteins in the ER it is possible that it may become a bystander target of the spreading antiviral immune response. Thus, we propose a model whereby immunity elicited towards Grp78 leads to the selection of responses towards the Ro polypeptides and the subsequent cascade of responses observed in human disease.
Direct identification of HLA‐presented CD8 T cell epitopes from transmitted founder HIV‐1 variants
Publisher: Wiley
Date: 08-2021
Abstract: Cytotoxic T lymphocytes (CTLs) are a critical arm of the immune response to viral infections. The activation and expansion of antigen specific CTL requires recognition of peptide antigens presented on class I major histocompatibility complex molecules (MHC‐1) of infected cells. Methods to identify presented peptide antigens that do not rely on the pre‐existence of antigen specific CTL are critical to the development of new vaccines. We infected activated CD4+ T cells with two HIV‐1 transmitted founder (TF) isolates and used high‐resolution mass spectrometry (MS) to identify HIV peptides bound on MHC‐1. Using this approach, we identified 14 MHC‐1 bound peptides from across the two TF isolates. Assessment of predicted binding thresholds revealed good association of the identified peptides to the shared HLA alleles between the HIV+ donors and the naïve PBMC s le with three peptides identified through peptide sequencing inducing a CD8 T‐cell response ( p 0.05). Direct infection of naïve CD4 cells by HIV TF isolates and sequencing of MHC‐I presented peptides by HPLC‐MS/MS enables identification of novel peptides that may be missed by alternative epitope mapping strategies and can provide valuable insight in to the first peptides presented by an HIV‐infected CD4 cell in the first few days post infection.
The SysteMHC Atlas project
Publisher: Oxford University Press (OUP)
Date: 29-07-2017
DOI: 10.1093/NAR/GKX664
Allelic association with ankylosing spondylitis fails to correlate with human leukocyte antigen B27 homodimer formation
Publisher: Elsevier BV
Date: 12-2019
A Structural and Immunological Basis for the Role of Human Leukocyte Antigen DQ8 in Celiac Disease
Publisher: Elsevier BV
Date: 07-2007
DOI: 10.1016/J.IMMUNI.2007.05.015
Abstract: The risk of celiac disease is strongly associated with human leukocyte antigen (HLA) DQ2 and to a lesser extent with HLA DQ8. Although the pathogenesis of HLA-DQ2-mediated celiac disease is established, the underlying basis for HLA-DQ8-mediated celiac disease remains unclear. We showed that T helper 1 (Th1) responses in HLA-DQ8-associated celiac pathology were indeed HLA DQ8 restricted and that multiple, mostly deamidated peptides derived from protease-sensitive sites of gliadin were recognized. This pattern of reactivity contrasted with the more absolute deamidation dependence and relative protease resistance of the dominant gliadin peptide in DQ2-mediated disease. We provided a structural basis for the selection of HLA-DQ8-restricted, deamidated gliadin peptides. The data established that the molecular mechanisms underlying HLA-DQ8-mediated celiac disease differed markedly from the HLA-DQ2-mediated form of the disease. Accordingly, nondietary therapeutic interventions in celiac disease might need to be tailored to the genotype of the in idual.
Immunopeptidomics Special Issue.
Publisher: Wiley
Date: 06-2018
Using mass spectrometry to monitor drug induced changes in antigen presentation by the human leukocyte antigen
Publisher: Wiley
Date: 18-07-2014
Conformational effects in reversed-phase high-performance liquid chromatography of polypeptides I. Resolution of insulin variants
Publisher: Elsevier BV
Date: 09-1995
DOI: 10.1016/0021-9673(95)00241-E
Abstract: In order to further characterise the role of conformation in the retention behaviour of polypeptides and proteins in reversed-phase high-performance liquid chromatography (RP-HPLC), the chromatographic properties of four different insulins have been studied as a function of temperature (over the range 5-85 degrees C) and column residence time (over the range 10-60 min). The role of the ligand structure was also investigated by comparing results obtained with a n-octadecyl (C18) and a n-butyl (C4) ligand immobilised to the same porous silica. Comparative structure-retention-stability relationships were determined from an examination of the influence of temperature on a number of chromatographic parameters including the chromatographic contact area, the affinity constant and the experimental band width. The results demonstrated that variations in temperature can be used to affect significant changes in selectivity between the different insulins despite their very high degree of sequence homology. These observations have permitted specific amino acid residues, and in particular those residues encompassing the region A8-A10, to be proposed to be directly involved in the chromatographic contact area of the insulin molecules. Overall, the analysis of the changes in various chromatographic parameters in response to variation of the amino acid sequence, temperature and other experimental parameters provides a powerful tool to elucidate the structural basis for the interfacial stability and the role of conformation on the retention behaviour of polypeptides and proteins in RP-HPLC.
Identification of an I‐Ad restricted peptide on the 65‐kilodalton heat shock protein of Mycobacterium avium
Publisher: Wiley
Date: 12-2002
DOI: 10.1046/J.1440-1711.2002.01128.X
Abstract: The 65 kilodalton heat shock protein (Hsp65) from mycobacterial species elicits immune responses and in some cases protective immunity. Here we have used a DNA sublibrary approach to identify antigenic fragments of Mycobacterium avium Hsp65 and a synthetic peptide approach to delineate CD4+ T cell determinants. A panel of Hsp65 reactive CD4+ T cell clones was established from lymph node cells obtained from BALB/c mice immunized with recombinant Hsp65. The clones were tested for proliferative reactivity against the products of the DNA sublibrary of the hsp65 gene. A T cell epitope, restricted by the I-Ad molecule, was identified within the C-terminal region of Hsp65 and the minimal epitope (amino acid residues 489-503) delineated using overlapping peptides spanning the C-terminal fragment. Additionally, the CD4+ T cell clone recognizing this epitope also responded to native Hsp65 present in M. avium lysates by both proliferation and cytokine production, indicating that the epitope was present and processed similarly both in the native and the recombinant forms of Hsp65. This sequence identified in BALB/c mice (Hsp65 489-503) is identical in other mycobacteria, notably M. tuberculosis, M. bovis and M. leprae, suggesting the epitope may have wider application in murine models of other mycobacterial infections.
Responses against islet antigens in NOD mice are prevented by tolerance to proinsulin but not IGRP
Publisher: American Society for Clinical Investigation
Date: 12-2006
DOI: 10.1172/JCI29602
Identification of a dominant self‐ligand bound to three HLA B44 alleles and the preliminary crystallographic analysis of recombinant forms of each complex
Publisher: Wiley
Date: 08-08-2002
DOI: 10.1016/S0014-5793(02)03149-6
Abstract: A naturally processed and presented ligand that is shared by human leukocyte antigen (HLA) B*4402, B*4403 and B*4405 molecules has been identified in peptides isolated from immunoaffinity purified HLA B44 complexes. This peptide derived from HLA DPalpha residues 46-54, an endogenous product of HLA DP expressed in the cell line Hmy2.C1R, is a prominent peptide in the mass spectra of species isolated as bound peptides from each allele when the three HLA B44 subtypes were introduced as transfected gene products. Recombinant truncated forms of HLA B*4405(1-276), HLA B*4403(1-276), HLA B*4402(1-276) and beta(2)-microglobulin have been prepared as inclusion bodies in Escherichia coli and refolded in the presence of the DPalpha(46-54) peptide and purified by a combination of size exclusion and anion exchange chromatography. This material was determined to be correctly folded based on detection of a conformational epitope recognized by the W6/32 monoclonal antibody. Large, plate-like crystals of the three complexes were produced using polyethylene glycol as the precipitant. All the crystals belong to the space group P2(1)2(1)2(1) with unit cell dimensions of approximately a=51, b=82, c=110 A. The crystals of three B44/DPalpha complexes diffracted to a resolution of 1.9 A or better. For the first time, using this natural, high abundance ligand of the HLA B44 molecules we have successfully expressed and refolded the three HLA B44 molecules and produced crystals amenable to structural studies.
Citrullinated peptide dendritic cell immunotherapy in HLA risk genotype–positive rheumatoid arthritis patients
Publisher: American Association for the Advancement of Science (AAAS)
Date: 03-06-2015
DOI: 10.1126/SCITRANSLMED.AAA9301
Abstract: Citrullinated peptide-exposed DCs induced immune regulatory effects in HLA risk genotype–positive RA patients.
Tumors reveal their secrets to cytotoxic T cells
Publisher: Proceedings of the National Academy of Sciences
Date: 03-10-2006
Membrane proteomics by high performance liquid chromatography–tandem mass spectrometry: Analytical approaches and challenges
Publisher: Wiley
Date: 21-01-2013
Abstract: Membrane proteins (MPs) play erse biologically important structural and functional roles including molecular transport, cell communication, and signal transduction. The dysfunctions of many are linked to deleterious human diseases and thus are of utmost importance in drug discovery. MPs comprise approximately 20-30% of all open reading frames (ORFs), however they are typically under-represented in many LC-MS proteomics experiments due to their low abundance and poor solubility. To address these analytical challenges, various MP enrichment, solubilization, digestion, and fractionation strategies have been employed to further improve the coverage of the membrane systems while maintaining compatibility with MS detection. This review discusses both established and emerging high-throughput gel-free analytical workflows in membrane proteomics, and the inherent advantages, disadvantages, and orthogonality of the various approaches. The issues of critical importance for successful LC-MS/MS detection such as detergent selection and minimizing ion suppression in detergent-based workflows are discussed in detail. Recent studies comparing the performance of different analytical strategies are highlighted in order to provide practical insight into the choice of the most appropriate method for membrane-centric applications ranging from cell surface biomarker discovery to MP interaction network mapping.
Defining the Substrate Specificity Determinants Recognized by the Active Site of C-Terminal Src Kinase-Homologous Kinase (CHK) and Identification of β-Synuclein as a Potential CHK Physiological Substr
Publisher: American Chemical Society (ACS)
Date: 18-07-2011
DOI: 10.1021/BI2001938
The energetic basis underpinning T-cell receptor recognition of a super-bulged peptide bound to a major histocompatibility complex class I molecule
Publisher: Elsevier BV
Date: 04-2012
Ingestion of oats and barley in patients with celiac disease mobilizes cross-reactive T cells activated by avenin peptides and immuno-dominant hordein peptides
Publisher: Elsevier BV
Date: 2015
DOI: 10.1016/J.JAUT.2014.10.003
Abstract: Celiac disease (CD) is a common CD4(+) T cell mediated enteropathy driven by gluten in wheat, rye, and barley. Whilst clinical feeding studies generally support the safety of oats ingestion in CD, the avenin protein from oats can stimulate intestinal gluten-reactive T cells isolated from some CD patients in vitro. Our objective was to establish whether ingestion of oats or other grains toxic in CD stimulate an avenin-specific T cell response in vivo. We fed participants a meal of oats (100 g/day over 3 days) to measure the in vivo polyclonal avenin-specific T cell responses to peptides contained within comprehensive avenin peptide libraries in 73 HLA-DQ2.5(+) CD patients. Grain cross-reactivity was investigated using oral challenge with wheat, barley, and rye. Avenin-specific responses were observed in 6/73 HLA-DQ2.5(+) CD patients (8%), against four closely related peptides. Oral barley challenge efficiently induced cross-reactive avenin/hordein-specific T cells in most CD patients, whereas wheat or rye challenge did not. In vitro, immunogenic avenin peptides were susceptible to digestive endopeptidases and showed weak HLA-DQ2.5 binding stability. Our findings indicate that CD patients possess T cells capable of responding to immuno-dominant hordein epitopes and homologous avenin peptides ex vivo, but the frequency and consistency of these T cells in blood is substantially higher after oral challenge with barley compared to oats. The low rates of T cell activation after a substantial oats challenge (100 g/d) suggests that doses of oats commonly consumed are insufficient to cause clinical relapse, and supports the safety of oats demonstrated in long-term feeding studies.
Mass spectrometry–based identification of MHC-bound peptides for immunopeptidomics
Publisher: No publisher found
Date: 2019
DOI: 10.1038/S41596-019-0133-Y
Abstract: Peptide antigens bound to molecules encoded by the major histocompatibility complex (MHC) and presented on the cell surface form the targets of T lymphocytes. This critical arm of the adaptive immune system facilitates the eradication of pathogen-infected and cancerous cells, as well as the production of antibodies. Methods to identify these peptide antigens are critical to the development of new vaccines, for which the goal is the generation of effective adaptive immune responses and long-lasting immune memory. Here, we describe a robust protocol for the identification of MHC-bound peptides from cell lines and tissues, using nano-ultra-performance liquid chromatography coupled to high-resolution mass spectrometry (nUPLC-MS/MS) and recent improvements in methods for isolation and characterization of these peptides. The protocol starts with the immunoaffinity capture of naturally processed MHC-peptide complexes. The peptides dissociate from the class I human leukocyte antigens (HLAs) upon acid denaturation. This peptide cargo is then extracted and separated into fractions by HPLC, and the peptides in these fractions are identified using nUPLC-MS/MS. With this protocol, several thousand peptides can be identified from a wide variety of cell types, including cancerous and infected cells and those from tissues, with a turnaround time of 2-3 d.
Molecular machinations of the MHC-I peptide loading complex
Publisher: Elsevier BV
Date: 02-2008
DOI: 10.1016/J.COI.2007.12.005
Abstract: The acquisition of an optimal peptide ligand by MHC class I molecules is crucial for the generation of immunity to viruses and tumors. This process is orchestrated by a molecular machine known as the peptide loading complex (PLC) that consists of specialized and general ER-resident molecules. These proteins collaborate to ensure the loading of an optimal peptide ligand into the antigen binding cleft of class I molecules. The surprising ersity of peptides bound to MHC class I molecules and recapitulation of class I assembly in vitro have provided new insights into the molecular machinations of peptide loading. Coupled with the extraordinary polymorphism of class I molecules and their differential dependence on various components of the PLC for cell surface expression, a picture of peptide loading at the molecular level has recently emerged and will be discussed herein.
Resourcing, annotating, and analysing synthetic peptides of SARS-CoV-2 for immunopeptidomics and other immunological studies.
Publisher: Wiley
Date: 14-04-2021
Abstract: SARS‐CoV‐2 has caused a significant ongoing pandemic worldwide. A number of studies have examined the T cell mediated immune responses against SARS‐CoV‐2, identifying potential T cell epitopes derived from the SARS‐CoV‐2 proteome. Such studies will aid in identifying targets for vaccination and immune monitoring. In this study, we applied tandem mass spectrometry and proteomic techniques to a library of ∼40,000 synthetic peptides, in order to generate a large dataset of SARS‐CoV‐2 derived peptide MS/MS spectra. On this basis, we built an online knowledgebase, termed virusMS ( virusms.erc.monash.edu/ ), to document, annotate and analyse these synthetic peptides and their spectral information. VirusMS incorporates a user‐friendly interface to facilitate searching, browsing and downloading the database content. Detailed annotations of the peptides, including experimental information, peptide modifications, predicted peptide‐HLA (human leukocyte antigen) binding affinities, and peptide MS/MS spectral data, are provided in virusMS.
Post-translationally modified T cell epitopes: immune recognition and immunotherapy
Publisher: Springer Science and Business Media LLC
Date: 08-09-2009
DOI: 10.1007/S00109-009-0526-4
Abstract: The functionality of proteins is greatly extended by a erse array of post-translational modifications (PTMs), many of which are recognized by the immune system. Notably, a significant proportion of peptides presented to T cells by the major histocompatibility complex in vivo are post-translationally modified. Since the cellular mechanisms that introduce and control protein modifications can differ between health and disease, the associated changes in antigen presentation have the potential to alter immune responses. A number of such situations have been implicated with infection, inflammation, autoimmune disease, and cancer, and the investigation of PTMs that affect antigen recognition has provided insight in disease progression as well as raising prospects for novel approaches in immunotherapy.
CTL Recognition of a Bulged Viral Peptide Involves Biased TCR Selection
Publisher: The American Association of Immunologists
Date: 15-09-2005
DOI: 10.4049/JIMMUNOL.175.6.3826
Abstract: MHC class I molecules generally present peptides of 8–10 aa long, forming an extended coil in the HLA cleft. Although longer peptides can also bind to class I molecules, they tend to bulge from the cleft and it is not known whether the TCR repertoire has sufficient plasticity to recognize these determinants during the antiviral CTL response. In this study, we show that unrelated in iduals infected with EBV generate a significant CTL response directed toward an HLA-B*3501-restricted, 11-mer epitope from the BZLF1 Ag. The 11-mer determinant adopts a highly bulged conformation with seven of the peptide side chains being solvent-exposed and available for TCR interaction. Such a complex potentially creates a structural challenge for TCR corecognition of both HLA-B*3501 and the peptide Ag. Surprisingly, unrelated B*3501 donors recognizing the 11-mer use identical or closely related αβ TCR sequences that share particular CDR3 motifs. Within the small number of dominant CTL clonotypes observed, each has discrete fine specificity for the exposed side chain residues of the peptide. The data show that bulged viral peptides are indeed immunogenic but suggest that the highly constrained TCR repertoire reflects a limit to TCR ersity when responding to some unusual MHC peptide ligands.
Quantitative proteomic profiling reveals novel region‐specific markers in the adult mouse brain
Publisher: Wiley
Date: 06-01-2014
Abstract: Despite major advances in neuroscience, a comprehensive understanding of the structural and functional components of the adult brain compartments remains to be fully elucidated at a quantitative molecular level. Indeed, over half of the soluble- and membrane-annotated proteins are currently unmapped within online digital brain atlases. In this study, two complementary approaches were used to assess the unique repertoire of proteins enriched within select regions of the adult mouse CNS, including the brain stem, cerebellum, and remaining brain hemispheres. Of the 1200 proteins visualized by 2D-DIGE, approximately 150 (including cytosolic and membrane proteins) were found to exhibit statistically significant changes in relative abundance thus representing putative region-specific brain markers. In addition to using a high-precision (18) O-labeling strategy for the quantitative LC-MS/MS mapping of membrane proteins isolated from myelin-enriched fractions, we have identified over 1000 proteins that have yet to be described in any other mammalian myelin proteome. A comparison of our myelin proteome was made to an existing transcriptome database containing mRNA abundance profiles during oligodendrocyte differentiation and has confirmed statistically significant abundance changes for ∼500 of these newly mapped proteins, thus revealing new roles in oligodendrocyte and myelin biology. These data offer a resource for the neuroscience community studying the molecular basis for specialized neuronal activities in the CNS and myelin-related disorders. The MS proteomics data associated with this manuscript have been deposited to the ProteomeXchange Consortium with the dataset identifier PXD000327 (ataset/PXD000327).
Conserved Features in the Structure, Mechanism, and Biogenesis of the Inverse Autotransporter Protein Family
Publisher: Oxford University Press (OUP)
Date: 10-05-2016
DOI: 10.1093/GBE/EVW112
Minimal Information About an Immuno‐Peptidomics Experiment (MIAIPE)
Publisher: Wiley
Date: 06-2018
Structural and Biological Basis of CTL Escape in Coronavirus-Infected Mice
Publisher: The American Association of Immunologists
Date: 15-03-2008
DOI: 10.4049/JIMMUNOL.180.6.3926
Abstract: Cytotoxic T lymphocyte escape occurs in many human infections, as well as mice infected with the JHM strain of mouse hepatitis virus, which exhibit CTL escape variants with mutations in a single epitope from the spike glycoprotein (S510). In all CTL epitopes prone to escape, only a subset of all potential variants is generally detected, even though many of the changes that are not selected would result in evasion of the T cell response. It is postulated that these unselected mutations significantly impair virus fitness. To define more precisely the basis for this preferential selection, we combine x-ray crystallographic studies of the MHC class I (Db)/S510 complexes with viral reverse genetics to identify a prominent TCR contact residue (tryptophan at position 4) prone to escape mutations. The data show that a mutation that is commonly detected in chronically infected mice (tryptophan to arginine) potently disrupts the topology of the complex, explaining its selection. However, other mutations at this residue, which also abrogate the CTL response, are never selected in vivo even though they do not compromise virus fitness in acutely infected animals or induce a significant de novo CTL response. Thus, while structural analyses of the S510/Db complex provide a strong basis for why some CTL escape variants are selected, our results also show that factors other than effects on virus fitness limit the ersification of CD8 T cell epitopes.
A structural basis for varied αβ TCR usage against an immunodominant EBV antigen restricted to a HLA-B8 molecule
Publisher: The American Association of Immunologists
Date: 2012
Abstract: EBV is a ubiquitous and persistent human pathogen, kept in check by the cytotoxic T cell response. In this study, we investigated how three TCRs, which differ in their T cell immunodominance hierarchies and gene usage, interact with the same EBV determinant (FLRGRAYGL), bound to the same Ag-presenting molecule, HLA-B8. We found that the three TCRs exhibit differing fine specificities for the viral Ag. Further, via structural and biophysical approaches, we demonstrated that the viral Ag provides the greatest energetic contribution to the TCR–peptide-HLA interaction, while focusing on a few adjacent HLA-based interactions to further tune fine-specificity requirements. Thus, the TCR engages the peptide-HLA with the viral Ag as the main glue, such that neighboring TCR–MHC interactions are recruited as a supportive adhesive. Collectively, we provide a portrait of how the host’s adaptive immune response differentially engages a common viral Ag.
Viral regulation of host cell biology by hijacking of the nucleolar DNA-damage response.
Publisher: Springer Science and Business Media LLC
Date: 03-08-2018
DOI: 10.1038/S41467-018-05354-7
Abstract: Recent studies indicate that nucleoli play critical roles in the DNA-damage response (DDR) via interaction of DDR machinery including NBS1 with nucleolar Treacle protein, a key mediator of ribosomal RNA (rRNA) transcription and processing. Here, using proteomics, confocal and single molecule super-resolution imaging, and infection under biosafety level-4 containment, we show that this nucleolar DDR pathway is targeted by infectious pathogens. We find that the matrix proteins of Hendra virus and Nipah virus, highly pathogenic viruses of the Henipavirus genus in the order Mononegavirales , interact with Treacle and inhibit its function, thereby silencing rRNA biogenesis, consistent with mimicking NBS1–Treacle interaction during a DDR. Furthermore, inhibition of Treacle expression/function enhances henipavirus production. These data identify a mechanism for viral modulation of host cells by appropriating the nucleolar DDR and represent, to our knowledge, the first direct intranucleolar function for proteins of any mononegavirus.
Supplementary Table S2 from Spliced Peptides and Cytokine-Driven Changes in the Immunopeptidome of Melanoma
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/2326-6066.22542561.V1
Abstract: Table S2
CD4+ T-Cell Deficiency in HIV Patients Responding to Antiretroviral Therapy Is Associated With Increased Expression of Interferon-Stimulated Genes in CD4+ T Cells
Publisher: Oxford University Press (OUP)
Date: 17-10-2011
Abstract: Most patients with human immunodeficiency virus (HIV) who remain CD4(+) T-cell deficient on antiretroviral therapy (ART) exhibit marked immune activation. As CD4(+) T-cell activation may be mediated by microbial translocation or interferon-alpha (IFN-α), we examined these factors in HIV patients with good or poor CD4(+) T-cell recovery on long-term ART. Messenger RNA levels for 3 interferon-stimulated genes were increased in CD4(+) T cells of patients with poor CD4(+) T-cell recovery, whereas levels in patients with good recovery did not differ from those in healthy controls. Poor CD4(+) T-cell recovery was also associated with CD4(+) T-cell expression of markers of activation, senescence, and apoptosis, and with increased serum levels of the lipopolysaccharide receptor and soluble CD14, but these were not significantly correlated with expression of the interferon-stimulated genes. Therefore, CD4(+) T-cell recovery may be adversely affected by the effects of IFN-α, which may be amenable to therapeutic intervention.
A naturally selected dimorphism within the HLA-B44 supertype alters class I structure, peptide repertoire, and T cell recognition
Publisher: Rockefeller University Press
Date: 25-08-2003
DOI: 10.1084/JEM.20030066
Abstract: HLA-B*4402 and B*4403 are naturally occurring MHC class I alleles that are both found at a high frequency in all human populations, and yet they only differ by one residue on the α2 helix (B*4402 Asp156→B*4403 Leu156). CTLs discriminate between HLA-B*4402 and B*4403, and these allotypes stimulate strong mutual allogeneic responses reflecting their known barrier to hemopoeitic stem cell transplantation. Although HLA-B*4402 and B*4403 share & % of their peptide repertoire, B*4403 presents more unique peptides than B*4402, consistent with the stronger T cell alloreactivity observed toward B*4403 compared with B*4402. Crystal structures of B*4402 and B*4403 show how the polymorphism at position 156 is completely buried and yet alters both the peptide and the heavy chain conformation, relaxing ligand selection by B*4403 compared with B*4402. Thus, the polymorphism between HLA-B*4402 and B*4403 modifies both peptide repertoire and T cell recognition, and is reflected in the paradoxically powerful alloreactivity that occurs across this “minimal” mismatch. The findings suggest that these closely related class I genes are maintained in erse human populations through their differential impact on the selection of peptide ligands and the T cell repertoire.
An emerging role for comprehensive proteome analysis in human pregnancy research
Publisher: Bioscientifica
Date: 06-2005
DOI: 10.1530/REP.1.00524
Abstract: Elucidation of underlying cellular and molecular mechanisms is pivotal to the comprehension of biological systems. The successful progression of processes such as pregnancy and parturition depends on the complex interactions between numerous biological molecules especially within the uterine microenvironment. The tissue- and stage-specific expression of these bio-molecules is intricately linked to and modulated by several endogenous and exogenous factors. Malfunctions may manifest as pregnancy disorders such as preterm labour, pre-ecl sia and fetal growth restriction that are major contributors to maternal and perinatal morbidity and mortality. Despite the immense amount of information available, our understanding of several aspects of these physiological processes remains incomplete. This translates into significant difficulties in the timely diagnosis and effective treatment of pregnancy-related complications. However, the emergence of powerful mass spectrometry-based proteomic techniques capable of identifying and characterizing multiple proteins simultaneously has added a new dimension to the field of biomedical research. Application of these high throughput methodologies with more conventional techniques in pregnancy-related research has begun to provide a novel perspective on the biochemical blueprint of pregnancy and its related disorders. Further, by enabling the identification of proteins specific to a disease process, proteomics is likely to contribute, not only to the comprehension of the underlying pathophysiologies, but also to the clinical diagnosis of multifactorial pregnancy disorders. Although the application of this technology to pregnancy research is in its infancy, characterization of the cellular proteome, unearthing of functional networks and the identification of disease biomarkers can be expected to significantly improve maternal healthcare in the future.
Novel strategy for identification of candidate cytotoxic T-cell epitopes from human preproinsulin.
Publisher: Wiley
Date: 13-10-2003
DOI: 10.1034/J.1399-0039.2003.00122.X
Abstract: We describe a strategy for identifying ligands of human leukocyte antigen (HLA) class I molecules based on a peptide library-mediated in vitro assembly of recombinant class I molecules. We established a microscale class I assembly assay and used a capture ELISA to quantify the assembled HLA-peptide complexes. The identity of the bound ligands was then deduced by mass spectrometry. In this method, HLA complexes assembled in vitro in the presence of components of a mixture of peptides were immunoprecipitated and the bound peptide(s) identified by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry. This process of epitope extraction is robust and can be used with complex mixtures containing in excess of 300 candidate ligands. A library of overlapping peptides representing all potential octamers, nonamers and decamers from human preproinsulin was synthesized using unique library chemistry. Peptides from the library were used to initiate assembly of recombinant HLA-B8, HLA-B15 and HLA-A2, facilitating the identification of candidate T-cell epitopes from preproinsulin.
Improved peptide-MHC class II interaction prediction through integration of eluted ligand and peptide affinity data
Publisher: Springer Science and Business Media LLC
Date: 10-06-2019
DOI: 10.1007/S00251-019-01122-Z
Abstract: Major histocompatibility complex (MHC) class II antigen presentation is a key component in eliciting a CD4+ T cell response. Precise prediction of peptide-MHC (pMHC) interactions has thus become a cornerstone in defining epitope candidates for rational vaccine design. Current pMHC prediction tools have, so far, primarily focused on inference from in vitro binding affinity. In the current study, we collate a large set of MHC class II eluted ligands generated by mass spectrometry to guide the prediction of MHC class II antigen presentation. We demonstrate that models developed on eluted ligands outperform those developed on pMHC binding affinity data. The predictive performance can be further enhanced by combining the eluted ligand and pMHC affinity data in a single prediction model. Furthermore, by including ligand data, the peptide length preference of MHC class II can be accurately learned by the prediction model. Finally, we demonstrate that our model significantly outperforms the current state-of-the-art prediction method, NetMHCIIpan, on an external dataset of eluted ligands and appears superior in identifying CD4+ T cell epitopes.
Human Leukocyte Antigen Class I-Restricted Activation of CD8+ T Cells Provides the Immunogenetic Basis of a Systemic Drug Hypersensitivity (DOI
Publisher: Elsevier BV
Date: 07-2008
Specialized insulin is used for chemical warfare by fish-hunting cone snails
Publisher: Proceedings of the National Academy of Sciences
Date: 20-01-2015
Abstract: The discovery and characterization of insulin, a key hormone of energy metabolism, provided a life-saving drug for diabetics. We show that insulin can be subverted for nefarious biological purposes: Venomous cone snails use specialized insulins to elicit hypoglycemic shock, facilitating capture of their fish prey. This finding extends our understanding of the chemical and functional ersity of venom components, such that the snail’s arsenal includes a erse set of neurotoxins that alters neuronal circuitry, as well as components that override glucose homeostasis. The highly expressed venom insulins are distinct from molluscan insulins and exhibit remarkable similarity to fish insulins. They are the smallest of all insulins characterized from any source, potentially providing new insights into structure-function elements of insulin action.
Positive-unlabelled learning of glycosylation sites in the human proteome
Publisher: Springer Science and Business Media LLC
Date: 06-03-2019
Linear ubiquitination prevents inflammation and regulates immune signalling
Publisher: Springer Science and Business Media LLC
Date: 03-2011
DOI: 10.1038/NATURE09816
Abstract: Members of the tumour necrosis factor (TNF) receptor superfamily have important functions in immunity and inflammation. Recently linear ubiquitin chains assembled by a complex containing HOIL-1 and HOIP (also known as RBCK1 and RNF31, respectively) were implicated in TNF signalling, yet their relevance in vivo remained uncertain. Here we identify SHARPIN as a third component of the linear ubiquitin chain assembly complex, recruited to the CD40 and TNF receptor signalling complexes together with its other constituents, HOIL-1 and HOIP. Mass spectrometry of TNF signalling complexes revealed RIP1 (also known as RIPK1) and NEMO (also known as IKKγ or IKBKG) to be linearly ubiquitinated. Mutation of the Sharpin gene (Sharpin(cpdm/cpdm)) causes chronic proliferative dermatitis (cpdm) characterized by inflammatory skin lesions and defective lymphoid organogenesis. Gene induction by TNF, CD40 ligand and interleukin-1β was attenuated in cpdm-derived cells which were rendered sensitive to TNF-induced death. Importantly, Tnf gene deficiency prevented skin lesions in cpdm mice. We conclude that by enabling linear ubiquitination in the TNF receptor signalling complex, SHARPIN interferes with TNF-induced cell death and, thereby, prevents inflammation. Our results provide evidence for the relevance of linear ubiquitination in vivo in preventing inflammation and regulating immune signalling.
Comparative Transcriptomics Highlights the Role of the Activator Protein 1 Transcription Factor in the Host Response to Ebolavirus
Publisher: American Society for Microbiology
Date: 12-2017
DOI: 10.1128/JVI.01174-17
Abstract: Ebolavirus and Marburgvirus comprise two genera of negative-sense single-stranded RNA viruses that cause severe hemorrhagic fevers in humans. Despite considerable research efforts, the molecular events following Ebola virus (EBOV) infection are poorly understood. With the view of identifying host factors that underpin EBOV pathogenesis, we compared the transcriptomes of EBOV-infected human, pig, and bat kidney cells using a transcriptome sequencing (RNA-seq) approach. Despite a significant difference in viral transcription/replication between the cell lines, all cells responded to EBOV infection through a robust induction of extracellular growth factors. Furthermore, a significant upregulation of activator protein 1 (AP1) transcription factor complex members FOS and JUN was observed in permissive cell lines. Functional studies focusing on human cells showed that EBOV infection induces protein expression, phosphorylation, and nuclear accumulation of JUN and, to a lesser degree, FOS. Using a luciferase-based reporter, we show that EBOV infection induces AP1 transactivation activity within human cells at 48 and 72 h postinfection. Finally, we show that JUN knockdown decreases the expression of EBOV-induced host gene expression. Taken together, our study highlights the role of AP1 in promoting the host gene expression profile that defines EBOV pathogenesis. IMPORTANCE Many questions remain about the molecular events that underpin filovirus pathophysiology. The rational design of new intervention strategies, such as postexposure therapeutics, will be significantly enhanced through an in-depth understanding of these molecular events. We believe that new insights into the molecular pathogenesis of EBOV may be possible by examining the transcriptomic response of taxonomically erse cell lines (derived from human, pig, and bat). We first identified the responsive pathways using an RNA-seq-based transcriptomics approach. Further functional and computational analysis focusing on human cells highlighted an important role for the AP1 transcription factor in mediating the transcriptional response to EBOV infection. Our study sheds new light on how host transcription factors respond to and promote the transcriptional landscape that follows viral infection.
Impact of glycans on T-cell tolerance to glycosylated self-antigens
Publisher: Wiley
Date: 15-07-2008
DOI: 10.1038/ICB.2008.48
Abstract: There is now substantial evidence that antigen post-translational modifications are recognized by T cells, and alterations in epitope modification has been linked to a number of autoimmune diseases. An estimated one third of the MHC ligands contain post-translational modification of epitopes. A common post-translational modification of proteins is glycosylation and it is predicted on theoretical grounds that approximately 1-5% of MHC ligands may bear a glycan. From numerous studies over the past 15 years it is clear that glycans can influence T cell responses either by contribution to the structure of the epitope or by influencing the profile of peptide epitopes presented by APCs. The influence of glycans on antigen processing and T cell recognition has particular relevance to the induction of tolerance to self-antigens. Here we discuss the potential impact of glycans on the profile of self-epitopes presented by APCs and the consequence of changes in glycosylation to generate neo self-epitopes resulting in the loss of tolerance and the development of autoimmune diseases. With the recent developments in profiling T cell epitopes, and with strategies for modulating glycosylation in vivo, it is now feasible to directly examine the global influence of glycans on self-tolerance and autoimmunity.
Simultaneous Quantification of Viral Antigen Expression Kinetics Using Data-Independent (DIA) Mass Spectrometry
Publisher: Elsevier BV
Date: 05-2015
Global metabolic analyses identify key differences in metabolite levels between polymyxin-susceptible and polymyxin-resistant Acinetobacter baumannii
Publisher: Springer Science and Business Media LLC
Date: 29-02-2016
DOI: 10.1038/SREP22287
Abstract: Multidrug-resistant Acinetobacter baumannii presents a global medical crisis and polymyxins are used as the last-line therapy. This study aimed to identify metabolic differences between polymyxin-susceptible and polymyxin-resistant A. baumannii using untargeted metabolomics. The metabolome of each A. baumannii strain was measured using liquid chromatography-mass spectrometry. Multivariate and univariate statistics and pathway analyses were employed to elucidate metabolic differences between the polymyxin-susceptible and -resistant A. baumannii strains. Significant differences were identified between the metabolic profiles of the polymyxin-susceptible and -resistant A. baumannii strains. The lipopolysaccharide (LPS) deficient, polymyxin-resistant 19606R showed perturbation in specific amino acid and carbohydrate metabolites, particularly pentose phosphate pathway (PPP) and tricarboxylic acid (TCA) cycle intermediates. Levels of nucleotides were lower in the LPS-deficient 19606R. Furthermore, 19606R exhibited a shift in its glycerophospholipid profile towards increased abundance of short-chain lipids compared to the parent polymyxin-susceptible ATCC 19606. In contrast, in a pair of clinical isolates 03–149.1 (polymyxin-susceptible) and 03–149.2 (polymyxin-resistant, due to modification of lipid A), minor metabolic differences were identified. Notably, peptidoglycan biosynthesis metabolites were significantly depleted in both of the aforementioned polymyxin-resistant strains. This is the first comparative untargeted metabolomics study to show substantial differences in the metabolic profiles of the polymyxin-susceptible and -resistant A. baumannii .
Phosphorylated self-peptides alter human leukocyte antigen class I-restricted antigen presentation and generate tumor-specific epitopes
Publisher: Proceedings of the National Academy of Sciences
Date: 24-02-2009
Abstract: Human leukocyte antigen (HLA) class I molecules present a variety of posttranslationally modified epitopes at the cell surface, although the consequences of such presentation remain largely unclear. Phosphorylation plays a critical cellular role, and deregulation in phosphate metabolism is associated with disease, including autoimmunity and tumor immunity. We have solved the high-resolution structures of 3 HLA A2-restricted phosphopeptides associated with tumor immunity and compared them with the structures of their nonphosphorylated counterparts. Phosphorylation of the epitope was observed to affect the structure and mobility of the bound epitope. In addition, the phosphoamino acid stabilized the HLA peptide complex in an epitope-specific manner and was observed to exhibit discrete flexibility within the antigen-binding cleft. Collectively, our data suggest that phosphorylation generates neoepitopes that represent demanding targets for T-cell receptor ligation. These findings provide insights into the mode of phosphopeptide presentation by HLA as well as providing a platform for the rational design of a generation of posttranslationally modified tumor vaccines.
Structural basis for enabling T-cell receptor diversity within biased virus-specific CD8+ T-cell responses.
Publisher: Proceedings of the National Academy of Sciences
Date: 23-05-2011
Abstract: Pathogen-specific responses are characterized by preferred profiles of peptide+class I MHC (pMHCI) glycoprotein-specific T-cell receptor (TCR) Variable (V)-region use. How TCRV-region bias impacts TCRαβ heterodimer selection and resultant ersity is unclear. The D b PA 224 –specific TCR repertoire in influenza A virus-infected C57BL/6J (B6) mice exhibits a preferred TCRV-region bias toward the TRBV29 gene segment and an optimal complementarity determining region (CDR3) β-length of 6 aa. Despite these restrictions, D b PA 224 -specific BV29 + T cells use a wide array of unique CDR3β sequences. Structural characterization of a single, TRBV29 + D b P A224 -specific TCRαβ-pMHCI complex demonstrated that CDR3α amino acid side chains made specific peptide interactions, but the CDR3β main chain exclusively contacted peptides. Thus, length but not amino acid sequence was key for recognition and flexibility in Vβ-region use. In support of this hypothesis, retrovirus expression of the D b PA 224 -specific TCRVα-chain was used to constrain pairing within a naive/immune epitope-specific repertoire. The retrogenic TCRVα paired with a ersity of CDR3βs in the context of a preferred TCRVβ spectrum. Overall, these data provide an explanation for the combination of TCRV region bias and ersity within selected repertoires, even as they maintain exquisite pMHCI specificity.
Quantifying epitope presentation using mass spectrometry
Publisher: Elsevier BV
Date: 12-2015
DOI: 10.1016/J.MOLIMM.2015.06.010
Abstract: Understanding the absolute quantities of MHC-bound epitopes (pMHC) presented on the surface of cells has long been a critical missing element in our knowledge of antigen presentation to T cells. Until recently, attaining such information has been restricted to the use of pMHC complex-specific monoclonal antibodies or T cell assays probing fractionated peptides eluted from cells. Although successful in a variety of cases, such approaches are limited in their scope and feasibility due to the nature of the reagents they are reliant upon. Here we report on the advancement of targeted mass spectrometry techniques to provide simultaneous and direct measurements of the relative and absolute levels of pMHC molecules and its potential for impact upon the field of antigen processing and presentation.
The insulin A-chain epitope recognized by human T cells is posttranslationally modified
Publisher: Rockefeller University Press
Date: 31-10-2005
DOI: 10.1084/JEM.20051251
Abstract: The autoimmune process that destroys the insulin-producing pancreatic β cells in type 1 diabetes (T1D) is targeted at insulin and its precursor, proinsulin. T cells that recognize the proximal A-chain of human insulin were identified recently in the pancreatic lymph nodes of subjects who had T1D. To investigate the specificity of proinsulin-specific T cells in T1D, we isolated human CD4+ T cell clones to proinsulin from the blood of a donor who had T1D. The clones recognized a naturally processed, HLA DR4–restricted epitope within the first 13 amino acids of the A-chain (A1–13) of human insulin. T cell recognition was dependent on the formation of a vicinal disulfide bond between adjacent cysteine residues at A6 and A7, which did not alter binding of the peptide to HLA DR4. CD4+ T cell clones that recognized this epitope were isolated from an HLA DR4+ child with autoantibodies to insulin, and therefore, at risk for T1D, but not from two healthy HLA DR4+ donors. We define for the first time a novel posttranslational modification that is required for T cell recognition of the insulin A-chain in T1D.
Restricted specificity of intermolecular spreading to endogenous La (SS-B) and 60 kDA Ro (SS-A) in experimental autoimmunity
Publisher: Wiley
Date: 18-07-2002
DOI: 10.1046/J.1365-3083.2002.01115.X
Abstract: Intermolecular spreading of humoral autoimmunity to different components of the Ro (SS-A) and La (SS-B) ribonucleoprotein (RNP) complex has been reported following immunization with a single component of the complex. Although the immune response to the immunizing antigen is polyclonal and ersified, little is known about the specificity of the recruited autoimmune responses to the endogenous Ro and La antigens which drive B-cell spreading. To determine the specificity of intermolecular spreading to La, we examined sera from 52 kDa Ro (Ro52)- and 60 kDa Ro (Ro60)-immunized C3H/HeJ mice for reactivity with recombinant fragments spanning endogenous mouse (m)La by enzyme-linked immunosorbent assay (ELISA) and immunoblotting. Sera from mice primed and boosted with recombinant Ro52 and Ro60 showed reactivity restricted to the COOH-terminal fragment of mLa (aa361-415). The recruited anti-La response was species-specific, cross-reacting weakly with the corresponding region on the human La molecule, and was abrogated by the preabsorption of the Ro-immune sera with mLa 361-415. Analogous experiments using recombinant mRo60 fragments spanning the mRo60 molecule revealed a similar pattern of oligoclonality in the specificity of anti-Ro60 autoimmunity following active immunization with La and Ro52. These results suggest that intermolecular-intrastructural T-B help is limiting in this model, and reveal unsuspected immunodominance of selected Ro-La epitopes in the spreading of the autoantibody response to these structures. The focusing of the recruited autoantibody response to these COOH-terminal regions of the Ro and La polypeptides may also reflect the surface accessibility of these regions in La-Ro RNP.
Huntingtin Inclusions Trigger Cellular Quiescence, Deactivate Apoptosis, and Lead to Delayed Necrosis
Publisher: Elsevier BV
Date: 05-2017
DOI: 10.1016/J.CELREP.2017.04.029
Abstract: Competing models exist in the literature for the relationship between mutant Huntingtin exon 1 (Httex1) inclusion formation and toxicity. In one, inclusions are adaptive by sequestering the proteotoxicity of soluble Httex1. In the other, inclusions compromise cellular activity as a result of proteome co-aggregation. Using a biosensor of Httex1 conformation in mammalian cell models, we discovered a mechanism that reconciles these competing models. Newly formed inclusions were composed of disordered Httex1 and ribonucleoproteins. As inclusions matured, Httex1 reconfigured into amyloid, and other glutamine-rich and prion domain-containing proteins were recruited. Soluble Httex1 caused a hyperpolarized mitochondrial membrane potential, increased reactive oxygen species, and promoted apoptosis. Inclusion formation triggered a collapsed mitochondrial potential, cellular quiescence, and deactivated apoptosis. We propose a revised model where sequestration of soluble Httex1 inclusions can remove the trigger for apoptosis but also co-aggregate other proteins, which curtails cellular metabolism and leads to a slow death by necrosis.
Defining the HLA class I‐associated viral antigen repertoire from HIV‐1‐infected human cells
Publisher: Wiley
Date: 04-11-2015
Quantitative and Qualitative Influences of Tapasin on the Class I Peptide Repertoire
Publisher: The American Association of Immunologists
Date: 15-01-2001
DOI: 10.4049/JIMMUNOL.166.2.1016
Abstract: Tapasin is critical for efficient loading and surface expression of most HLA class I molecules. The high level surface expression of HLA-B*2705 on tapasin-deficient 721.220 cells allowed the influence of this chaperone on peptide repertoire to be examined. Comparison of peptides bound to HLA-B*2705 expressed on tapasin-deficient and -proficient cells by mass spectrometry revealed an overall reduction in the recovery of B*2705-bound peptides isolated from tapasin-deficient cells despite similar yields of B27 heavy chain and β2-microglobulin. This indicated that a proportion of suboptimal ligands were associated with B27, and they were lost during the purification process. Notwithstanding this failure to recover these suboptimal peptides, there was substantial overlap in the repertoire and biochemical properties of peptides recovered from B27 complexes derived from tapasin-positive and -negative cells. Although many peptides were preferentially or uniquely isolated from B*2705 in tapasin-positive cells, a number of species were preferentially recovered in the absence of tapasin, and some of these peptide ligands have been sequenced. In general, these ligands did not exhibit exceptional binding affinity, and we invoke an argument based on lumenal availability and affinity to explain their tapasin independence. The differential display of peptides in tapasin-negative and -positive cells was also apparent in the reactivity of peptide-sensitive alloreactive CTL raised against tapasin-positive and -negative targets, demonstrating the functional relevance of the biochemical observation of changes in peptide repertoire in the tapasin-deficient APC. Overall, the data reveal that tapasin quantitatively and qualitatively influences ligand selection by class I molecules.
Rapid expansion of the protein disulfide isomerase gene family facilitates the folding of venom peptides
Publisher: Proceedings of the National Academy of Sciences
Date: 08-03-2016
Abstract: The majority of secreted proteins contain disulfide bonds that provide structural stability in the extracellular environment. The formation of correct disulfide bonds is assisted by the enzyme protein disulfide isomerase (PDI). Most secreted structural domains are ancient and widely distributed in all metazoans in contrast, erse sets of unique disulfide-rich structural domains have more recently evolved in venomous marine snails (superfamily Conoidea comprising ,000 species). We have discovered a previously undescribed gene family encoding PDIs of unprecedented ersity. We suggest that these enzymes constitute an important part of the supporting molecular infrastructure required for properly folding the plethora of structural domains expressed in the venoms of snails in different conoidean lineages.
More than one reason to rethink the use of peptides in vaccine design
Publisher: Springer Science and Business Media LLC
Date: 05-2007
DOI: 10.1038/NRD2224
Abstract: The use of peptides as therapeutics is experiencing renewed enthusiasm owing to advances in delivery, stability and design. Moreover, there is a growing emphasis on the use of peptides in vaccine design as insights into tissue-specific processing of the immunogenic epitopes of proteins and the discovery of unusually long cytotoxic T-lymphocyte epitopes broaden the range of targets and give clues to enhancing peptide immunogenicity. Peptides can also be synthesized with known post-translational modifications and/or deliberately introduced protease-resistant peptide bonds to regulate their processing independent of tissue-specific proteolysis and to stabilize these compounds in vivo. We discuss the potential of peptide-based vaccines for the treatment of chronic viral diseases and cancer, and review recent developments in the field of peptide-based vaccines.
A molecular basis for the association of the HLA-DRB1 locus, citrullination, and rheumatoid arthritis.
Publisher: Rockefeller University Press
Date: 04-11-2013
DOI: 10.1084/JEM.20131241
Abstract: Rheumatoid arthritis (RA) is strongly associated with the human leukocyte antigen (HLA)-DRB1 locus that possesses the shared susceptibility epitope (SE) and the citrullination of self-antigens. We show how citrullinated aggrecan and vimentin epitopes bind to HLA-DRB1*04:01/04. Citrulline was accommodated within the electropositive P4 pocket of HLA-DRB1*04:01/04, whereas the electronegative P4 pocket of the RA-resistant HLA-DRB1*04:02 allomorph interacted with arginine or citrulline-containing epitopes. Peptide elution studies revealed P4 arginine–containing peptides from HLA-DRB1*04:02, but not from HLA-DRB1*04:01/04. Citrullination altered protease susceptibility of vimentin, thereby generating self-epitopes that are presented to T cells in HLA-DRB1*04:01+ in iduals. Using HLA-II tetramers, we observed citrullinated vimentin- and aggrecan-specific CD4+ T cells in the peripheral blood of HLA-DRB1*04:01+ RA-affected and healthy in iduals. In RA patients, autoreactive T cell numbers correlated with disease activity and were deficient in regulatory T cells relative to healthy in iduals. These findings reshape our understanding of the association between citrullination, the HLA-DRB1 locus, and T cell autoreactivity in RA.
Physical and toxicological profiles of human IAPP amyloids and plaques
Publisher: Elsevier BV
Date: 2019
Refinement in the production and purification of recombinant HCMV IE1-pp65 protein for the generation of epitope-specific T cell immunity
Publisher: Elsevier BV
Date: 09-2008
DOI: 10.1016/J.PEP.2008.05.001
Abstract: Human cytomegalovirus (HCMV) remains one of the most common opportunistic infections causing disease following stem cell transplantation, despite the availability of anti-viral therapies. Adoptive immunotherapy has the potential to further aid in counteracting chronic viral reactivation and subsequent disease by restoring viral immunity through the transfer of virus-specific T cells from transplant donors to their recipients. Our study refines the production and purification of a recombinant HCMV protein containing two of the most immunodominant antigens (IE1 and pp65) for the generation of polyclonal HCMV-specific T cells. In doing so, a 6x His-tagged IE1-pp65 protein was generated using a serum-free baculovirus/insect cell expression system and soluble IE1-pp65 protein was subsequently purified using Ni-NTA affinity chromatography under stringent conditions to obtain a highly pure product. The ability of the recombinant IE1-pp65 protein to elicit a functional T cell mediated immune response was demonstrated by the vigorous reactivation and expansion of HLA-A2-restricted pp65(495-503)-specific CD8+ T cells. This recombinant IE1-pp65 protein can potentially generate a multitude of HLA-restricted HCMV-specific T cells, providing a better alternative to using costly overlapping peptides or HCMV lysates for expansion of T cells for use in adoptive immunotherapy strategies.
Human Plasma Protein Corona of Aβ Amyloid and Its Impact on Islet Amyloid Polypeptide Cross-Seeding
Publisher: American Chemical Society (ACS)
Date: 07-01-2020
Immunology by numbers: quantitation of antigen presentation completes the quantitative milieu of systems immunology!
Publisher: Elsevier BV
Date: 06-2016
DOI: 10.1016/J.COI.2016.03.007
Abstract: We review approaches to quantitate antigen presentation using a variety of biological and biochemical readouts and highlight the emerging role of mass spectrometry (MS) in defining and quantifying MHC-bound peptides presented at the cell surface. The combination of high mass accuracy in the determination of the molecular weight of the intact peptide of interest and its signature pattern of fragmentation during tandem MS provide an unambiguous and definitive identification. This is in contrast to the potential receptor cross-reactivity towards closely related peptides and variable dose responsiveness seen in biological readouts. In addition, we gaze into the not too distant future where big data approaches in MS can be accommodated to quantify whole immunopeptidomes both in vitro and in vivo.
Revisiting the Arthritogenic Peptide Theory: Quantitative Not Qualitative Changes in the Peptide Repertoire of HLA–B27 Allotypes
Publisher: Wiley
Date: 25-02-2015
DOI: 10.1002/ART.38963
Abstract: The association of HLA-B27 with spondyloarthropathy is one of the strongest documented for any autoimmune disease. A common hypothesis for this association is the arthritogenic peptide concept. This dictates that differences in the peptide binding preferences of disease-associated and non-disease-associated HLA-B27 allotypes underlie the presentation of bacterial and self-peptides, leading to cross-reactive T cell immunity and subsequent autoimmune attack of affected tissues. The aim of this study was to analyze and compare self-peptides from 8 HLA-B27 allotypes, to increase existing data sets of HLA-B27 ligands, to refine and compare their consensus-binding motifs, and to reveal similarities and differences in the peptide repertoire of the HLA-B27 subtypes. Qualitative differences in the peptides bound to the 8 most frequent HLA-B27 subtypes were determined by tandem mass spectrometry, and quantitative changes in allelic binding specificities were determined by highly sensitive and targeted multiple reaction monitoring mass spectrometry. We identified >7,500 major histocompatibility complex class I peptides derived from the 8 most common HLA-B27 allotypes (HLA-B*27:02 to HLA-B*27:09). We describe in idual binding motifs for these alleles for the 9-12-mer ligands. The peptide repertoires of these closely related alleles showed significant overlap. Allelic polymorphisms resulting in changes in the amino acid composition of the antigen-binding cleft manifested largely as quantitative changes in the peptide cargo of these molecules. Absolute binding preferences of HLA-B27 allotypes do not explain disease association. The arthritogenic peptide theory needs to be reassessed in terms of quantitative changes in self-peptide presentation, T cell selection, and altered conformation of bound peptides.
The self-peptide repertoire plays a critical role in transplant tolerance induction
Publisher: Cold Spring Harbor Laboratory
Date: 09-11-2020
DOI: 10.1101/2020.11.09.359968
Abstract: While direct allorecognition underpins both solid organ allograft rejection and tolerance induction, the specific molecular targets of most directly-alloreactive CD8 + T cells have not been defined. In this study, we used a combination of genetically-engineered MHC class I (MHC I) constructs, mice with a hepatocyte-specific mutation in the class I antigen-presentation pathway and immunopeptidomic analysis to provide definitive evidence for the contribution of the peptide cargo of allogeneic MHC I molecules to transplant tolerance induction. We established a systematic approach for the discovery of directly-recognised pMHC epitopes, and identified 17 strongly immunogenic H-2K b -associated peptides recognised by CD8 + T cells from B10.BR (H-2 k ) mice, 13 of which were also recognised by BALB/c (H-2 d ) mice. As few as five different tetramers used together were able to identify a high proportion of alloreactive T cells within a polyclonal population, suggesting that there are immunodominant allogeneic MHC-peptide complexes that can account for a large component of the alloresponse.
A T cell receptor flattens a bulged antigenic peptide presented by a major histocompatibility complex class I molecule
Publisher: Springer Science and Business Media LLC
Date: 28-01-2007
DOI: 10.1038/NI1432
Abstract: Plasticity of the T cell receptor (TCR) is a hallmark of major histocompatibility complex (MHC)-restricted T cell recognition. However, it is unclear whether interactions of TCR and peptide-MHC class I (pMHCI) always conform to this paradigm. Here we describe the structure of a TCR, ELS4, in its non-ligand-bound form and in complex with a prominent 'bulged' Epstein-Barr virus peptide bound to HLA-B(*)3501. This complex was atypical of previously characterized TCR-pMHCI interactions in that a rigid face of the TCR crumpled the bulged antigenic determinant. This peptide 'bulldozing' created a more featureless pMHCI determinant, allowing the TCR to maximize MHC class I contacts essential for MHC class I restriction of TCR recognition. Our findings represent a mechanism of antigen recognition whereby the plasticity of the T cell response is dictated mainly by adjustments in the MHC-bound peptide.
The primary immune response to Vaccinia virus vaccination includes cells with a distinct cytotoxic effector CD4 T-cell phenotype
Publisher: Elsevier BV
Date: 10-2016
DOI: 10.1016/J.VACCINE.2016.09.009
Abstract: Smallpox was eradicated by a global program of inoculation with Vaccinia virus (VV). Robust VV-specific CD4 T-cell responses during primary infection are likely essential to controlling VV replication. Although there is increasing interest in cytolytic CD4 T-cells across many viral infections, the importance of these cells during acute VV infection is unclear. We undertook a detailed functional and genetic characterization of CD4 T-cells during acute VV-infection of humans. VV-specific T-cells were identified by up-regulation of activation markers directly ex vivo and through cytokine and co-stimulatory molecule expression. At day-13-post primary inoculation with VV, CD38highCD45RO+ CD4 T-cells were purified by cell sorting, RNA isolated and analysed by microarray. Differential expression of up-regulated genes in activated CD4 T-cells was confirmed at the mRNA and protein levels. We compared analyses of VV-specific CD4 T-cells to studies on 12 subjects with primary HIV infection (PHI). VV-specific T-cells lines were established from PBMCs collected post vaccination and checked for cytotoxicity potential. A median 11.9% CD4 T-cells were CD38highCD45RO+ at day-13 post-VV inoculation, compared to 3.0% prior and 10.4% during PHI. Activated CD4 T-cells had an up-regulation of genes related to cytolytic function, including granzymes K and A, perforin, granulysin, TIA-1, and Rab27a. No difference was seen between CD4 T-cell expression of perforin or TIA-1 to VV and PHI, however granzyme k was more dominant in the VV response. At 25:1 effector to target ratio, two VV-specific T-cell lines exhibited 62% and 30% cytotoxicity respectively and CD107a degranulation. We show for the first time that CD4 CTL are prominent in the early response to VV. Understanding the role of CD4 CTL in the generation of an effective anti-viral memory may help develop more effective vaccines for diseases such as HIV.
The differentiation state of the Schwann cell progenitor drives phenotypic variation between two contagious cancers
Publisher: Public Library of Science (PLoS)
Date: 15-11-2021
DOI: 10.1371/JOURNAL.PPAT.1010033
Abstract: Contagious cancers are a rare pathogenic phenomenon in which cancer cells gain the ability to spread between genetically distinct hosts. Nine ex les have been identified across marine bivalves, dogs and Tasmanian devils, but the Tasmanian devil is the only mammalian species known to have given rise to two distinct lineages of contagious cancer, termed Devil Facial Tumour 1 (DFT1) and 2 (DFT2). Remarkably, DFT1 and DFT2 arose independently from the same cell type, a Schwann cell, and while their ultra-structural features are highly similar they exhibit variation in their mutational signatures and infection dynamics. As such, DFT1 and DFT2 provide a unique framework for investigating how a common progenitor cell can give rise to distinct contagious cancers. Using a proteomics approach, we show that DFT1 and DFT2 are derived from Schwann cells in different differentiation states, with DFT2 carrying a molecular signature of a less well differentiated Schwann cell. Under inflammatory signals DFT1 and DFT2 have different gene expression profiles, most notably involving Schwann cell markers of differentiation, reflecting the influence of their distinct origins. Further, DFT2 cells express immune cell markers typically expressed during nerve repair, consistent with an ability to manipulate their extracellular environment, facilitating the cell’s ability to transmit between in iduals. The emergence of two contagious cancers in the Tasmanian devil suggests that the inherent plasticity of Schwann cells confers a vulnerability to the formation of contagious cancers.
Hormone-like peptides in the venoms of marine cone snails
Publisher: Elsevier BV
Date: 04-2017
Optimisation of peptide-based cytotoxic T-cell determinants using non-natural amino acids
Publisher: Springer Science and Business Media LLC
Date: 09-2003
DOI: 10.1007/BF02442589
Correction: Article on Vaccination Reveals a Long Dominant CTL Epitope
Publisher: American Association for Cancer Research (AACR)
Date: 15-05-2009
Conformational effects in reversed-phase high-performance liquid chromatography of polypeptides II. The role of insulin A and B chains in the chromatographic behaviour of insulin
Publisher: Elsevier BV
Date: 09-1995
DOI: 10.1016/0021-9673(95)00228-F
Abstract: The contribution of the insulin A- and B-chain to the retention and bandwidth behaviour of bovine insulin has been investigated. The influence of temperature and residence time on the logarithmic capacity factor (log k) versus the mole fraction of organic modifier psi, i.e. the effect of temperature and ligand residency on the S and log k0 values of the in idual peptide chains, were assessed at temperatures between 5 and 85 degrees C and elution times between 30 to 90 min with an n-octadecyl (C18) and an n-butyl (C4) sorbent. Analysis of these log k versus psi dependencies revealed that the insulin A-chain exhibits retention behaviour significantly different to the intact insulin molecule whilst the B-chain exhibits retention behaviour which is remarkably similar to the parent protein. However, in terms of kinetic processes, the A-chain exhibited a peak-splitting phenomenon at higher temperatures which was similar to the behaviour of the intact insulin molecule, whilst only bandbroadening with no peak splitting was apparent for the B-chain. Overall, the similarity of the retention behaviour of the insulin B-chain and the intact insulin molecule with regard to their temperature and residency dependencies suggests that the insulin B-chain makes a significant contribution to the chromatographic contact region of the insulin molecule when this polypeptide is exposed to hydrocarbonaceous ligands at low to intermediate temperatures due to the progressive unfolding of the molecule and greater accessibility of the previously buried B-chain residues.(ABSTRACT TRUNCATED AT 250 WORDS)
T Cell Cross-Reactivity between a Highly Immunogenic EBV Epitope and a Self-Peptide Naturally Presented by HLA-B*18:01+ Cells
Publisher: The American Association of Immunologists
Date: 15-05-2015
Abstract: T cell cross-reactivity underpins the molecular mimicry hypothesis in which microbial peptides sharing structural features with host peptides stimulate T cells that cross-react with self-peptides, thereby initiating and/or perpetuating autoimmune disease. EBV represents a potentially important factor in the pathogenesis of several T cell–mediated autoimmune disorders, with molecular mimicry a likely mechanism. In this study, we describe a human self-peptide (DELEIKAY) that is a homolog of a highly immunogenic EBV T cell epitope (SELEIKRY) presented by HLA-B*18:01. This self-peptide was shown to bind stably to HLA-B*18:01, and peptide elution/mass spectrometric studies showed it is naturally presented by this HLA molecule on the surface of human cells. A significant proportion of CD8+ T cells raised from some healthy in iduals against this EBV epitope cross-reacted with the self-peptide. A erse array of TCRs was expressed by the cross-reactive T cells, with variable functional avidity for the self-peptide, including some T cells that appeared to avoid autoreactivity by a narrow margin, with only 10-fold more of the self-peptide required for equivalent activation as compared with the EBV peptide. Structural studies revealed that the self-peptide–HLA-B*18:01 complex is a structural mimic of the EBV peptide–HLA-B*18:01 complex, and that the strong antiviral T cell response is primarily dependent on the alanine/arginine mismatch at position 7. To our knowledge, this is the first report confirming the natural presentation of a self-peptide cross-recognized in the context of self-HLA by EBV-reactive CD8+ T cells. These results illustrate how aberrant immune responses and immunopathological diseases could be generated by EBV infection.
High molecular weight components of the injected venom of fish-hunting cone snails target the vascular system
Publisher: Elsevier BV
Date: 10-2013
DOI: 10.1016/J.JPROT.2013.07.007
Abstract: The venom of marine cone snails is a rich source of pharmacotherapeutic compounds with striking target specificity and functional ersity. Small, disulfide-rich peptide toxins are the most well characterized active compounds in cone snail venom. However, reports on the presence of larger polypeptides have recently emerged. The majority of these studies have focused on the content of the dissected venom gland rather than the injected venom itself. Recent breakthroughs in the sensitivity of protein and nucleotide sequencing techniques allow for the exploration of the proteomic ersity of injected venom. Using mass spectrometric analysis of injected venoms of the two fish-hunting cone snails Conus purpurascens and Conus ermineus, we demonstrate the presence of angiotensin-converting enzyme-1 (ACE-1) and endothelin converting enzyme-1 (ECE-1), metalloproteases that activate potent vasoconstrictive peptides. ACE activity was confirmed in the venom of C. purpurascens and was significantly reduced in venom preincubated with the ACE inhibitor captopril. Reverse-transcription PCR demonstrated that these enzymes are expressed in the venom glands of other cone snail species with different prey preferences. These findings strongly suggest that cone snails employ compounds that cause disruption of cardiovascular function as part of their complex envenomation strategy, leading to the enhancement of neurotropic peptide toxin activity. To our knowledge, this is the first study to show the presence of ACE and ECE in the venom of cone snails. Identification of these vasoactive peptide-releasing proteases in the injected venoms of two fish-hunting cone snails highlights their role in envenomation and enhances our understanding of the complex hunting strategies utilized by these marine predators. Our findings on the expression of these enzymes in other cone snail species suggests an important biological role of ACE and ECE in these animals and points towards recruitment into venom from general physiological processes.
Multi-Omic Analysis of Two Common P53 Mutations: Proteins Regulated by Mutated P53 as Potential Targets for Immunotherapy
Publisher: MDPI AG
Date: 17-08-2022
Abstract: The p53 protein is mutated in more than 50% of human cancers. Mutated p53 proteins not only lose their normal function but often acquire novel oncogenic functions, a phenomenon termed mutant p53 gain-of-function. Mutant p53 has been shown to affect the transcription of a range of genes, as well as protein–protein interactions with transcription factors and other effectors however, no one has intensively investigated and identified these proteins, or their MHC presented epitopes, from the viewpoint of their ability to act as targets for immunotherapeutic interventions. We investigated the molecular changes that occurred after the TP53 null osteosarcoma cells, SaOS-2, were transfected with one of two conformational p53-mutants, either R175H or R273H. We then examined the phenotypic and functional changes using macroscopic observations, proliferation, gene expression and proteomics alongside immunopeptidome profiling of peptide antigen presentation in the context of major histocompatibility complex (MHC) class I molecules. We identified several candidate proteins in both TP53 mutant cell lines with differential expression when compared to the TP53 null vector control, SaOS-V. Quantitative SWATH proteomics combined with immune-peptidome analysis of the class-I eluted peptides identified several epitopes presented on pMHC and in silico analysis shortlisted which antigens were expressed in a range of cancerous but not adjacent healthy tissues. Out of all the candidates, KLC1 and TOP2A showed high levels of expression in every tumor type examined. From these proteins, three A2 and four pan HLA-A epitopes were identified in both R175H and R273H from TOP2A. We have now provided a short list of future immunotherapy targets for the treatment of cancers harboring mutated TP53.
The use of CRISPR/Cas9 gene editing to confirm congenic contaminations in host-pathogen interaction studies
Publisher: Frontiers Media SA
Date: 19-03-2018
Embryonic toxin expression in the cone snail Conus victoriae
Publisher: Elsevier BV
Date: 06-2011
PCprophet: a framework for protein complex prediction and differential analysis using proteomic data
Publisher: Springer Science and Business Media LLC
Date: 15-04-2021
DOI: 10.1038/S41592-021-01107-5
Abstract: Despite the availability of methods for analyzing protein complexes, systematic analysis of complexes under multiple conditions remains challenging. Approaches based on biochemical fractionation of intact, native complexes and correlation of protein profiles have shown promise. However, most approaches for interpreting cofractionation datasets to yield complex composition and rearrangements between s les depend considerably on protein-protein interaction inference. We introduce PCprophet, a toolkit built on size exclusion chromatography-sequential window acquisition of all theoretical mass spectrometry (SEC-SWATH-MS) data to predict protein complexes and characterize their changes across experimental conditions. We demonstrate improved performance of PCprophet over state-of-the-art approaches and introduce a Bayesian approach to analyze altered protein-protein interactions across conditions. We provide both command-line and graphical interfaces to support the application of PCprophet to any cofractionation MS dataset, independent of separation or quantitative liquid chromatography-MS workflow, for the detection and quantitative tracking of protein complexes and their physiological dynamics.
HLA Peptide Length Preferences Control CD8+ T Cell Responses
Publisher: The American Association of Immunologists
Date: 15-07-2013
Abstract: Class I HLAs generally present peptides of 8–10 aa in length, although it is unclear whether peptide length preferences are affected by HLA polymorphism. In this study, we investigated the CD8+ T cell response to the BZLF1 Ag of EBV, which includes overlapping sequences of different size that nevertheless conform to the binding motif of the large and abundant HLA-B*44 supertype. Whereas HLA-B*18:01+ in iduals responded strongly and exclusively to the octamer peptide 173SELEIKRY180, HLA-B*44:03+ in iduals responded to the atypically large dodecamer peptide 169EECDSELEIKRY180, which encompasses the octamer peptide. Moreover, the octamer peptide bound more stably to HLA-B*18:01 than did the dodecamer peptide, whereas, conversely, HLA-B*44:03 bound only the longer peptide. Furthermore, crystal structures of these viral peptide–HLA complexes showed that the Ag-binding cleft of HLA-B*18:01 was more ideally suited to bind shorter peptides, whereas HLA-B*44:03 exhibited characteristics that favored the presentation of longer peptides. Mass spectrometric identification of & 1000 naturally presented ligands revealed that HLA-B*18:01 was more biased toward presenting shorter peptides than was HLA-B*44:03. Collectively, these data highlight a mechanism through which polymorphism within an HLA class I supertype can ersify determinant selection and immune responses by varying peptide length preferences.
Immunopeptidogenomics: Harnessing RNA-Seq to Illuminate the Dark Immunopeptidome
Publisher: Elsevier BV
Date: 2021
Biased T cell receptor usage directed against human leukocyte antigen DQ8-restricted gliadin peptides is associated with celiac disease.
Publisher: Elsevier BV
Date: 10-2012
Dissecting the role of peptides in the immune response: Theory, practice and the application to vaccine design
Publisher: Wiley
Date: 2003
DOI: 10.1002/PSC.456
Abstract: Analytical biochemistry and synthetic peptide based chemistry have helped to reveal the pivotal role that peptides play in determining the specificity, magnitude and quality of both humoral (antibody) and cellular (cytotoxic and helper T cell) immune responses. In addition, peptide based technologies are now at the forefront of vaccine design and medical diagnostics. The chemical technologies used to assemble peptides into immunogenic structures have made great strides over the past decade and assembly of highly pure peptides which can be incorporated into high molecular weight species, multimeric and even branched structures together with non-peptidic material is now routine. These structures have a wide range of applications in designer vaccines and diagnostic reagents. Thus the tools of the peptide chemist are exquisitely placed to answer questions about immune recognition and along the way to provide us with new and improved vaccines and diagnostics.
A Molecular Basis for the Interplay between T Cells, Viral Mutants, and Human Leukocyte Antigen Micropolymorphism
Publisher: Elsevier BV
Date: 06-2014
A Long, Naturally Presented Immunodominant Epitope from NY-ESO-1 Tumor Antigen: Implications for Cancer Vaccine Design
Publisher: American Association for Cancer Research (AACR)
Date: 29-01-2009
DOI: 10.1158/0008-5472.CAN-08-2926
Abstract: The tumor antigen NY-ESO-1 is a promising cancer vaccine target. We describe here a novel HLA-B7–restricted NY-ESO-1 epitope, encompassing amino acids 60-72 (APRGPHGGAASGL), which is naturally presented by melanoma cells. The tumor epitope bound to HLA-B7 by bulging outward from the peptide-binding cleft. This bulged epitope was not an impediment to T-cell recognition, however, because four of six HLA-B7+ melanoma patients vaccinated with NY-ESO-1 ISCOMATRIX vaccine generated a potent T-cell response to this determinant. Moreover, the response to this epitope was immunodominant in three of these patients and, unlike the T-cell responses to bulged HLA class I viral epitopes, the responding T cells possessed a remarkably broad TCR repertoire. Interestingly, HLA-B7+ melanoma patients who did not receive the NY-ESO-1 ISCOMATRIX vaccine rarely generated a spontaneous T-cell response to this cryptic epitope, suggesting a lack of priming of such T cells in the natural anti–NY-ESO-1 response, which may be corrected by vaccination. Together, our results reveal several surprising aspects of antitumor immunity and have implications for cancer vaccine design. [Cancer Res 2009 (3):1046–54]
Posttranslational Modifications of Proteins in Type 1 Diabetes: The Next Step in Finding the Cure?
Publisher: American Diabetes Association
Date: 17-07-2012
DOI: 10.2337/DB11-1675
Abstract: The overall role of modification of β-cell antigens in type 1 diabetes has not been elucidated and was the focus of a recent workshop on posttranslational modification of proteins in type 1 diabetes. The prevailing opinion of the workshop attendees was that novel insights into the mechanism of loss of immune tolerance might be gained and that novel diagnostic and therapeutic approaches could be developed for type 1 diabetes if protein modifications were shown to play a critical role in the disease.
Endogenous and exogenous factors contributing to the surface expression of HLA B27 on mutant APC
Publisher: Elsevier BV
Date: 02-2000
DOI: 10.1016/S0198-8859(99)00139-1
Abstract: We have examined the expression of HLA B*2705 in the mutant cell line 721.220, which lacks endogenous HLA A and B alleles and expresses a defective tapasin molecule. Several peptide sensitive mAbs distinguish between HLA B*2705 expressed on the surface of 721.220 cells (B27.220) and 721.220 cells co-transfected with human tapasin (B27.220.hTsn). This differential staining defines subtle differences in the conformation of HLA B27, which most likely reflect changes in the repertoire of antigenic peptides bound to B27 in the presence and absence of wild type tapasin. HLA B27 molecules expressed on the surface of 721.220 display increased levels of "free" B27 heavy chain (HC-10 staining), an epitope that is dependent on TAP-translocated peptides. The conformation and stability of B27 molecules was examined by investigating the integrity of mAb epitopes and the half-lives of these complexes on cells cultured with and without serum. The decay of surface B27 epitopes occurred more rapidly in B27.220 and this effect was exaggerated in serum free media. Importantly, the decay of surface B27 molecules in B27.220.hTsn cells was characterized by an early increase in HC-10 staining when the cells were grown in serum free media. This decay of B27 molecules via HC-10 reactive intermediates was not observed in B27.220 cells, implying molecules on these cells may already have passed through this stage prior to surface expression. Taken together these observations indicate that tapasin has a significant contribution to the composition and stability of the B27-bound peptide repertoire.
Hierarchical self-tolerance to T cell determinants within the ubiquitous nuclear self-antigen La (SS-B) permits induction of systemic autoimmunity in normal mice.
Publisher: Rockefeller University Press
Date: 11-1996
Abstract: Systemic autoimmune diseases are frequently associated with clustering of high titer autoantibody responses towards nuclear self-antigens. Little is known, however, about the extent of immune tolerance to the target nuclear antigens or the events leading to the complex autoantibody responses that are characteristic of systemic autoimmunity. To address these issues, we have examined the mouse immune response to La autoantigen (mLa) and the homologous human La antigen (hLa), which are components of the La(SS-B)/Ro(SS-A) ribonucleoprotein (RNP) complex targeted in systemic lupus erythematosus and primary Sjögren's syndrome. The findings reveal the presence of hierarchical T cell tolerance involving multiple autodeterminants within the La autoantigen expressed by normal H-2k and H-2a mice. At one end of this spectrum, there was no detectable T or B cell autoimmunity observed in mice that were immunized with the immunodominant mLa287-301 determinant, which differed by a single residue in its core sequence from the homologous but highly immunogenic human La288-302 determinant. Interestingly, the mLa287-301 peptide acted as an altered peptide ligand that specifically antagonized the activation of an hLa288-302-specific T cell hybridoma. In contrast to the tolerogenic mLa287-301 determinant, a range of autoimmune potential was identified among poorly tolerizing, subdominant self-peptides present within mouse La autoantigen. Notably, immunization of normal mice with the autologous subdominant La25-44 and La106-129 determinants resulted in limited or no detectable autoantibody response. In contrast, immunization with the subdominant mouse La13-30 determinant induced a proliferative T cell response associated with the appearance of specific autoantibodies recognizing multiple intrastructural (La) and intermolecular components (Ro) of the murine La/Ro RNP. The findings suggest how ersified autoimmunity might follow initiation of immunity to simple peptide mimics of poorly tolerogenic determinants that are present within ubiquitous self-antigens.
Dominant protection from HLA-linked autoimmunity by antigen-specific regulatory T cells.
Publisher: Springer Science and Business Media LLC
Date: 05-2017
DOI: 10.1038/NATURE22329
The Structure of H-2Kb and Kbm8 Complexed to a Herpes Simplex Virus Determinant: Evidence for a Conformational Switch That Governs T Cell Repertoire Selection and Viral Resistance
Publisher: The American Association of Immunologists
Date: 07-2004
DOI: 10.4049/JIMMUNOL.173.1.402
Abstract: Polymorphism within the MHC not only affects peptide specificity but also has a critical influence on the T cell repertoire for ex le, the CD8 T cell response toward an immunodominant HSV glycoprotein B peptide is more erse and of higher avidity in H-2bm8 compared with H-2b mice. We have examined the basis for the selection of these distinct antiviral T cell repertoires by comparing the high-resolution structures of Kb and Kbm8, in complex with cognate peptide Ag. Although Kb and Kbm8 differ by four residues within the Ag-binding cleft, the most striking difference in the two structures was the disparate conformation adopted by the shared residue, Arg62. The altered dynamics of Arg62, coupled with a small rigid-body movement in the α1 helix encompassing this residue, correlated with biased Vα usage in the B6 mice. Moreover, an analysis of all known TCR/MHC complexes reveals that Arg62 invariably interacts with the TCR CDR1α loop. Accordingly, Arg62 appears to function as a conformational switch that may govern T cell selection and protective immunity.
Drug Hypersensitivity and Human Leukocyte Antigens of the Major Histocompatibility Complex
Publisher: Annual Reviews
Date: 10-02-2012
DOI: 10.1146/ANNUREV-PHARMTOX-010611-134701
Abstract: The human leukocyte antigen (HLA) genes are the most polymorphic in the human genome and are critical in regulating specific immunity, hence their historical discovery as “immune response” genes. HLA allotypes are also implicated in unwanted immune reactions, including drug hypersensitivity syndrome, in which small therapeutic drugs interact with antigenic peptides to drive T cell responses restricted by host HLA. Abacavir, allo-purinol, and carbamazepine are three commonly used drugs that cause a T cell–mediated hypersensitivity that is HLA linked, with each drug exhibiting striking specificity for presentation by defined HLA allotypes. Recent findings have begun to unearth the mechanistic basis for these HLA associations, and here we review recent advances in the field of HLA-associated drug hypersensitivities.
Determinant spreading: Lessons from animal models and human disease
Publisher: Wiley
Date: 08-1998
DOI: 10.1111/J.1600-065X.1998.TB01222.X
Abstract: Spreading of the immune response is a common theme in organ-specific and systemic autoimmune diseases. We evaluated whether some of the mixed antinuclear antibody patterns characteristic of systemic autoimmunity might be the result of determinant spreading from a single initiating event. Immunisation of healthy mice with in idual protein components of the La/Ro ribonucleoprotein (RNP) targeted in systemic lupus erythematosus and primary Sjögren's syndrome induced autoantibodies recognising Ro60 (SS-A), Ro52 (SS-A) and La (SS-B) and in some cases the molecular chaperones calreticulin and Grp78. The endogenous antigen(s) driving determinant spreading might be derived from physiological apoptosis which could explain the involvement of some chaperone proteins in the autoimmune response. Diversified anti-La/Ro antibody responses were initiated by challenge with a single subdominant T epitope of La even though some self epitopes of La were efficiently tolerised. The pattern of autoantibody responses in primary Sjögren's syndrome was strongly influenced by HLA class II phenotype which we speculate controls activation of T cells recognising defined peptides from the La/Ro RNP. In this way, HLA class II alleles may be critical in influencing initiation and spreading of systemic autoimmune reactions. Molecular mimicry of such determinants by exogenous agents might readily initiate spreading of an autoimmune response in genetically susceptible hosts.
T cell receptor reversed polarity recognition of a self-antigen major histocompatibility complex
Publisher: Springer Science and Business Media LLC
Date: 05-10-2015
DOI: 10.1038/NI.3271
Abstract: Central to adaptive immunity is the interaction between the αβ T cell receptor (TCR) and peptide presented by the major histocompatibility complex (MHC) molecule. Presumably reflecting TCR-MHC bias and T cell signaling constraints, the TCR universally adopts a canonical polarity atop the MHC. We report the structures of two TCRs, derived from human induced T regulatory (iT(reg)) cells, complexed to an MHC class II molecule presenting a proinsulin-derived peptide. The ternary complexes revealed a 180° polarity reversal compared to all other TCR-peptide-MHC complex structures. Namely, the iT(reg) TCR α-chain and β-chain are overlaid with the α-chain and β-chain of MHC class II, respectively. Nevertheless, this TCR interaction elicited a peptide-reactive, MHC-restricted T cell signal. Thus TCRs are not 'hardwired' to interact with MHC molecules in a stereotypic manner to elicit a T cell signal, a finding that fundamentally challenges our understanding of TCR recognition.
A prominent role of PDIA6 in processing of misfolded proinsulin
Publisher: Elsevier BV
Date: 06-2016
DOI: 10.1016/J.BBAPAP.2016.03.002
Abstract: Despite its critical role in maintaining glucose homeostasis, surprisingly little is known about proinsulin folding in the endoplasmic reticulum. In this study we aimed to understand the chaperones involved in the maturation and degradation of proinsulin. We generated pancreatic beta cell lines expressing FLAG-tagged proinsulin. Several chaperones (including BiP, PDIA6, calnexin, calreticulin, GRP170, Erdj3 and ribophorin II) co-immunoprecipitated with proinsulin suggesting a role for these proteins in folding. To investigate the chaperones responsible for targeting misfolded proinsulin for degradation, we also created a beta cell line expressing FLAG-tagged proinsulin carrying the Akita mutation (Cys96Tyr). All chaperones found to be associated with wild type proinsulin also co-immunoprecipitated with Akita proinsulin. However, one additional protein, namely P58(IPK), specifically precipitated with Akita proinsulin and approximately ten fold more PDIA6, but not other PDI family members, was bound to Akita proinsulin. The latter suggests that PDIA6 may act as a key reductase and target misfolded proinsulin to the ER-degradation pathway. The preferential association of PDIA6 to Akita proinsulin was also confirmed in another beta cell line (βTC-6). Furthermore, for the first time, a physiologically relevant substrate for PDIA6 has been evidenced. Thus, this study has identified several chaperones/foldases that associated with wild type proinsulin and has also provided a comprehensive interactome for Akita misfolded proinsulin.
A shared TCR bias toward an immunogenic EBV epitope dominates in HLA-B*07: 02-Expressing individuals
Publisher: The American Association of Immunologists
Date: 15-09-2020
Abstract: EBV is one of the most common viruses found in humans and is prototypic of a persistent viral infection characterized by periods of latency. Across many HLA class I molecules, the latent-specific CD8+ T cell response is focused on epitopes derived from the EBNA-3 protein family. In the case of HLA-B*07:02 restriction, a highly frequent class I allele, the T cell response is dominated by an epitope spanning residues 379–387 of EBNA-3 (RPPIFIRRL [EBVRPP]). However, little is known about either the TCR repertoire specific for this epitope or the molecular basis for this observed immunodominance. The EBVRPP CD8+ T cell response was common among both EBV-seropositive HLA-B*07:02+ healthy and immunocompromised in iduals. Similar TCRs were identified in EBVRPP–specific CD8+ T cell repertoires across multiple HLA-B7+ in iduals, indicating a shared Ag-driven bias in TCR usage. In particular, TRBV4-1 and TRAV38 usage was observed in five out of six in iduals studied. In this study, we report the crystal structure of a TRBV4-1+ TCR–HLA-B*07:02/EBVRPP complex, which provides a molecular basis for the observed TRBV4-1 bias. These findings enhance our understanding of the CD8+ T cell response toward a common EBV determinant in HLA-B*07:02+ in iduals.
A comprehensive analysis of peptides presented by HLA‐A1
Publisher: Wiley
Date: 16-04-2015
DOI: 10.1111/TAN.12565
Abstract: Human leukocyte antigen (HLA)-A1 is one of the most common Caucasian HLA-A alleles. Here, we describe the comprehensive analysis of the HLA-A*01:01 ligand repertoire with the identification of 4735 naturally processed and presented peptides derived from 2477 source proteins. We found HLA-A*01:01 bound an equivalent number of ligands of 9 or 10 amino acids in length as well as being remarkably tolerant of even longer peptides. Indeed close to half of the HLA-A1 bound peptides identified ranged between 11 and 13 amino acids in length. These longer peptides contained the strong canonical motif of and acidic E/D residue at position 3 (P3) and Y at the C-terminus (CΩ), a motif that was still apparent in peptides of up to 18 amino acids in length. The identification of this large database of natural ligands will facilitate the refinement of predictive algorithms particularly with respect to longer peptide ligands.
Acute experimental infection of bats and ferrets with Hendra virus: Insights into the early host response of the reservoir host and susceptible model species
Publisher: Public Library of Science (PLoS)
Date: 30-03-2020
Thermostability profiling of MHC-bound peptides: a new dimension in immunopeptidomics and aid for immunotherapy design
Publisher: Springer Science and Business Media LLC
Date: 09-12-2020
DOI: 10.1038/S41467-020-20166-4
Abstract: The features of peptide antigens that contribute to their immunogenicity are not well understood. Although the stability of peptide-MHC (pMHC) is known to be important, current assays assess this interaction only for peptides in isolation and not in the context of natural antigen processing and presentation. Here, we present a method that provides a comprehensive and unbiased measure of pMHC stability for thousands of in idual ligands detected simultaneously by mass spectrometry (MS). The method allows rapid assessment of intra-allelic and inter-allelic differences in pMHC stability and reveals profiles of stability that are broader than previously appreciated. The additional dimensionality of the data facilitated the training of a model which improves the prediction of peptide immunogenicity, specifically of cancer neoepitopes. This assay can be applied to any cells bearing MHC or MHC-like molecules, offering insight into not only the endogenous immunopeptidome, but also that of neoepitopes and pathogen-derived sequences.
Epitope discovery and their use in peptide based vaccines
Publisher: Bentham Science Publishers Ltd.
Date: 09-2010
DOI: 10.2174/138161210793292447
Abstract: With recent advances in the design and delivery of peptide-based therapeutics there has been a growing interest on the use of peptides in vaccine design. Moreover, functional dissection and proteomic analysis of the immunogenic epitopes of proteins from pathogenic micro-organisms, cancers and self-tissues targeted by autoimmune responses, have broadened the range of target epitopes and given clues to enhancing peptide immunogenicity. Consistent with these observations peptides can be synthesised with defined chemical modifications to mimic natural epitopes and/or deliberately introduce protease resistant peptide bonds to regulate their processing independent of tissue specific proteolysis and to stabilize these compounds in vivo. We discuss the potential of peptide-based vaccines for the treatment of chronic viral diseases and cancer and review recent developments in the field of epitope discovery and peptide-based vaccines.
Tracking protein aggregation and mislocalization in cells with flow cytometry.
Publisher: Springer Science and Business Media LLC
Date: 18-03-2012
DOI: 10.1038/NMETH.1930
Abstract: We applied pulse-shape analysis (PulSA) to monitor protein localization changes in mammalian cells by flow cytometry. PulSA enabled high-throughput tracking of protein aggregation, translocation from the cytoplasm to the nucleus and trafficking from the plasma membrane to the Golgi as well as stress-granule formation. Combining PulSA with tetracysteine-based oligomer sensors in a cell model of Huntington's disease enabled further separation of cells enriched with monomers, oligomers and inclusion bodies.
HLA‐associated antiepileptic drug‐induced cutaneous adverse reactions
Publisher: Wiley
Date: 09-04-2019
DOI: 10.1111/TAN.13530
Abstract: Adverse drug reactions (ADRs) are a common cause of hospital admissions (up to 19%), with the majority of cases due to off-target predictable drug effects (type A reactions). However, idiosyncratic drug-induced immune activated (type B) reactions contribute to a range of hypersensitivity reactions, with T-cell-mediated type IV hypersensitivity reactions mainly manifesting as cutaneous ADRs (cADRs). Aromatic antiepileptic drugs (AEDs), used in the treatment of epilepsy as well as bipolar disorder or neuropathic pain, have been implicated as culprit drugs in a spectrum of pathologies ranging from mild maculopapular exanthema (MPE) to severe and life-threatening conditions including drug reaction with eosinophilia and systemic symptoms (DRESS), Stevens-Johnson syndrome (SJS) and toxic epidermal necrolysis (TEN). These AED-induced cADRs are unpredictable based on pharmacological and clinical factors alone, thereby prompting investigations into genomic contributors mediating risk of pathology. The most strongly associated risk genes identified are from the human leukocyte antigen (HLA) class I alleles, which play a critical role in adaptive immunity by flagging either infected or aberrant cells for recognition by surveying T-cells. In the setting of drug hypersensitivity, the immunogenicity of HLA molecules and their peptide cargo can be modulated by interactions with small drug molecules that drive inappropriate T-cell responses. This review discusses the current understanding of HLA class I molecules in modifying risk of AED-induced cADRs.
TCR_Explore: a novel webtool for T cell receptor repertoire analysis
Publisher: Cold Spring Harbor Laboratory
Date: 04-11-2022
DOI: 10.1101/2022.11.03.514642
Abstract: T cells expressing either alpha-beta or gamma-delta T cell receptors (TCR) are critical sentinels of the adaptive immune system, with receptor ersity being essential for protective immunity against a broad array of pathogens and agents. Programs available to profile TCR clonotypic signatures can be limiting for users with no coding expertise. Current analytical pipelines can be inefficient due to manual processing steps, open to data transcription errors and have multiple analytical tools with unique inputs that require coding expertise. Here we present a bespoke webtool designed for users irrespective of coding expertise, coined ‘TCR_Explore’, incorporating automated quality control steps that generates a single output file for creation of flexible and publication ready figures. TCR_Explore will elevate a user’s capacity to undertake in-depth TCR repertoire analysis of both new and pre-existing datasets for identification of T cell clonotypes associated with health and disease. The web application is located at tcr-explore.erc.monash.edu for users to interactively explore TCR repertoire datasets. Bespoke program for non-specialists in computerised methodologies for deep exploration of TCR repertoire analysis Automated QC and analysis pipelines for Sanger based TCR sequencing coupled with immunophenotyping, with the capacity for integration of other sequencing platform outputs Automated summary processes to aid data visualisation and generation of publication-ready graphical displays
Supplementary Table S5 from Spliced Peptides and Cytokine-Driven Changes in the Immunopeptidome of Melanoma
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/2326-6066.22542552.V1
Abstract: Table S5
Twenty years of bioinformatics research for protease-specific substrate and cleavage site prediction: a comprehensive revisit and benchmarking of existing methods
Publisher: Oxford University Press (OUP)
Date: 29-08-2018
DOI: 10.1093/BIB/BBY077
Abstract: The roles of proteolytic cleavage have been intensively investigated and discussed during the past two decades. This irreversible chemical process has been frequently reported to influence a number of crucial biological processes (BPs), such as cell cycle, protein regulation and inflammation. A number of advanced studies have been published aiming at deciphering the mechanisms of proteolytic cleavage. Given its significance and the large number of functionally enriched substrates targeted by specific proteases, many computational approaches have been established for accurate prediction of protease-specific substrates and their cleavage sites. Consequently, there is an urgent need to systematically assess the state-of-the-art computational approaches for protease-specific cleavage site prediction to further advance the existing methodologies and to improve the prediction performance. With this goal in mind, in this article, we carefully evaluated a total of 19 computational methods (including 8 scoring function-based methods and 11 machine learning-based methods) in terms of their underlying algorithm, calculated features, performance evaluation and software usability. Then, extensive independent tests were performed to assess the robustness and scalability of the reviewed methods using our carefully prepared independent test data sets with 3641 cleavage sites (specific to 10 proteases). The comparative experimental results demonstrate that PROSPERous is the most accurate generic method for predicting eight protease-specific cleavage sites, while GPS-CCD and LabCaS outperformed other predictors for calpain-specific cleavage sites. Based on our review, we then outlined some potential ways to improve the prediction performance and ease the computational burden by applying ensemble learning, deep learning, positive unlabeled learning and parallel and distributed computing techniques. We anticipate that our study will serve as a practical and useful guide for interested readers to further advance next-generation bioinformatics tools for protease-specific cleavage site prediction.
A subset of HLA-I peptides are not genomically templated: Evidence for cis- and trans-spliced peptide ligands
Publisher: American Association for the Advancement of Science (AAAS)
Date: 26-10-2018
DOI: 10.1126/SCIIMMUNOL.AAR3947
Abstract: A considerable proportion of HLA-I peptides likely derive from cis- and trans-splicing events.
Plasmodium berghei Hsp90 contains a natural immunogenic I-Ab-restricted antigen common to rodent and human Plasmodium species
Publisher: Elsevier BV
Date: 2021
RP-HPLC binding domains of proteins
Publisher: American Chemical Society (ACS)
Date: 29-10-1998
DOI: 10.1021/AC980473C
Abstract: Procedures have been developed to identify the chromatographic binding domains of horse heart cytochrome c (Cyt c) and bovine growth hormone (bGH) during their interaction with reversed-phase sorbent materials. The procedure involves adsorption of the protein solute to the chromatographic sorbent, followed by proteolytic cleavage. Comparison of the proteolytic map obtained for Cyt c and bGH in free solution with the corresponding map obtained when these proteins are adsorbed to the chromatographic sorbent revealed significant differences in the digestion pattern. Following characterization of the peptides generated in both maps, the results indicated that specific regions on the surface of both Cyt c and bGH are inaccessible to tryptic cleavage when adsorbed to the hydrophobic surface of both a C-4 and a C-18 sorbent. Based on the assumption that the region of the protein surface that is in contact with the sorbent remains intact and bound to the sorbent during the digestion step, while the protein surface that is exposed to the solvent is accessible to proteolysis, the regions that were inaccessible to tryptic digestion were found to correspond to hydrophobic domains on the protein surface. These results also suggest that the three-dimensional structures of these proteins remain largely intact upon adsorption to the hydrophobic surface.
A comprehensive review and performance evaluation of bioinformatics tools for HLA class I peptide-binding prediction
Publisher: Oxford University Press (OUP)
Date: 04-05-2020
DOI: 10.1093/BIB/BBZ051
Abstract: Human leukocyte antigen class I (HLA-I) molecules are encoded by major histocompatibility complex (MHC) class I loci in humans. The binding and interaction between HLA-I molecules and intracellular peptides derived from a variety of proteolytic mechanisms play a crucial role in subsequent T-cell recognition of target cells and the specificity of the immune response. In this context, tools that predict the likelihood for a peptide to bind to specific HLA class I allotypes are important for selecting the most promising antigenic targets for immunotherapy. In this article, we comprehensively review a variety of currently available tools for predicting the binding of peptides to a selection of HLA-I allomorphs. Specifically, we compare their calculation methods for the prediction score, employed algorithms, evaluation strategies and software functionalities. In addition, we have evaluated the prediction performance of the reviewed tools based on an independent validation data set, containing 21 101 experimentally verified ligands across 19 HLA-I allotypes. The benchmarking results show that MixMHCpred 2.0.1 achieves the best performance for predicting peptides binding to most of the HLA-I allomorphs studied, while NetMHCpan 4.0 and NetMHCcons 1.1 outperform the other machine learning-based and consensus-based tools, respectively. Importantly, it should be noted that a peptide predicted with a higher binding score for a specific HLA allotype does not necessarily imply it will be immunogenic. That said, peptide-binding predictors are still very useful in that they can help to significantly reduce the large number of epitope candidates that need to be experimentally verified. Several other factors, including susceptibility to proteasome cleavage, peptide transport into the endoplasmic reticulum and T-cell receptor repertoire, also contribute to the immunogenicity of peptide antigens, and some of them can be considered by some predictors. Therefore, integrating features derived from these additional factors together with HLA-binding properties by using machine-learning algorithms may increase the prediction accuracy of immunogenic peptides. As such, we anticipate that this review and benchmarking survey will assist researchers in selecting appropriate prediction tools that best suit their purposes and provide useful guidelines for the development of improved antigen predictors in the future.
Constitutive and inflammatory immunopeptidome of pancreatic beta-cells
Publisher: American Diabetes Association
Date: 16-10-2012
DOI: 10.2337/DB11-1333
Abstract: Type 1 diabetes is characterized by the autoimmune destruction of pancreatic β-cells. Recognition of major histocompatibility complex (MHC)-bound peptides is critical for both the initiation and progression of disease. In this study, MHC peptide complexes were purified from NIT-1 β-cells, interferon-γ (IFN-γ)-treated NIT-1 cells, splenic and thymic tissue of 12-week-old NOD mice, and peptides identified by mass spectrometry. In addition to global liquid chromatography–tandem mass spectrometry analysis, the targeted approach of multiple-reaction monitoring was used to quantitate the immunodominant Kd-restricted T-cell epitope islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP)206–214. We identified & ,000 MHC-bound peptides 1,100 of these presented by β-cells grown under normal conditions or after exposure to IFN-γ. These include sequences from a number of known autoantigens. Quantitation of IGRP206–214 revealed low-level presentation by Kd (∼25 complexes/cell) on NIT-1 cells after IFN-γ treatment compared with the simultaneous presentation of the endogenously processed Kd-restricted peptide Janus kinase-1355–363 (∼15,000 copies/cell). We have successfully sequenced peptides from NIT-1 β-cells under basal and inflammatory conditions. We have shown the feasibility of quantitating disease-associated peptides and provide the first direct demonstration of the disparity between presentation of a known autoantigenic epitope and a common endogenously presented peptide.
T cell autoreactivity directed toward CD1c itself rather than toward carried self lipids.
Publisher: Springer Science and Business Media LLC
Date: 12-03-2018
The 1.5 Å Crystal Structure of a Highly Selected Antiviral T Cell Receptor Provides Evidence for a Structural Basis of Immunodominance
Publisher: Elsevier BV
Date: 11-2002
DOI: 10.1016/S0969-2126(02)00878-X
Abstract: Despite a potential repertoire of >10(15) alphabeta T cell receptors (TcR), the HLA B8-restricted cytolytic T cell response to a latent antigen of Epstein-Barr virus (EBV) is strikingly limited in the TcR sequences that are selected. Even in unrelated in iduals this response is dominated by a single highly restricted TcR clonotype that selects identical combinations of hypervariable Valpha, Vbeta, D, J, and N region genes. We have determined the 1.5 A crystal structure of this "public" TcR, revealing that five of the six hypervariable loops adopt novel conformations providing a unique combining site that contains a deep pocket predicted to overlay the HLA B8-peptide complex. The findings suggest a structural basis for the immunodominance of this clonotype in the immune response to EBV.
T-cells behaving badly: structural insights into alloreactivity and autoimmunity
Publisher: Elsevier BV
Date: 10-2008
DOI: 10.1016/J.COI.2008.07.006
Abstract: T-cells play a critical role in protective immunity, with their broad receptor repertoire capable of engaging erse foreign pMHC landscapes. While the versatility and specificity of this MHC-restricted response is the hallmark of adaptive immunity, unwanted TCR interactions can profoundly effect the health of the host leading for instance to allograft rejection or autoimmunity. In allogeneic transplantation, such adverse reactions can occur by an indirect pathway when the TCR interacts with self-MHC molecules presenting allogeneic MHC derived peptides. Direct T-cell alloreactivity involves recognition of the allogeneic molecule itself either through molecular mimicry or by novel pMHC binding modes. By contrast, auto-reactive TCRs are considered to interact in a manner distinct from cognate pMHC interactions. Here we review recent advances in the field, focusing on structural data pertaining to alloreactivity and auto-reactivity and discuss implications for T-cell mediated transplant rejection and autoimmune disorders.
T cell receptor recognition of hybrid insulin peptides bound to HLA-DQ8
Publisher: Springer Science and Business Media LLC
Date: 25-08-2021
DOI: 10.1038/S41467-021-25404-X
Abstract: HLA-DQ8, a genetic risk factor in type I diabetes (T1D), presents hybrid insulin peptides (HIPs) to autoreactive CD4+ T cells. The abundance of spliced peptides binding to HLA-DQ8 and how they are subsequently recognised by the autoreactive T cell repertoire is unknown. Here we report, the HIP ( GQV E LGGG NAV E VLK), derived from splicing of insulin and islet amyloid polypeptides, generates a preferred peptide-binding motif for HLA-DQ8. HLA-DQ8-HIP tetramer + T cells from the peripheral blood of a T1D patient are characterised by repeated TRBV5 usage, which matches the TCR bias of CD4+ T cells reactive to the HIP peptide isolated from the pancreatic islets of a patient with T1D. The crystal structure of three TRBV5+ TCR-HLA-DQ8-HIP complexes shows that the TRBV5 -encoded TCR β-chain forms a common landing pad on the HLA-DQ8 molecule. The N- and C-termini of the HIP is recognised predominantly by the TCR α-chain and TCR β-chain, respectively, in all three TCR ternary complexes. Accordingly, TRBV5 + TCR recognition of HIP peptides might occur via a ‘polarised’ mechanism, whereby each chain within the αβTCR heterodimer recognises distinct origins of the spliced peptide presented by HLA-DQ8.
Spliced Peptides and Cytokine-Driven Changes in the Immunopeptidome of Melanoma
Publisher: American Association for Cancer Research (AACR)
Date: 10-2020
DOI: 10.1158/2326-6066.CIR-19-0894
Abstract: Antigen recognition by CD8+ T cells is governed by the pool of peptide antigens presented on the cell surface in the context of HLA class I complexes. Studies have shown not only a high degree of plasticity in the immunopeptidome, but also that a considerable fraction of all presented peptides is generated through proteasome-mediated splicing of noncontiguous regions of proteins to form novel peptide antigens. Here, we used high-resolution mass spectrometry combined with new bioinformatic approaches to characterize the immunopeptidome of melanoma cells in the presence or absence of IFNγ. In total, we identified more than 60,000 peptides from a single patient-derived cell line (LM-MEL-44) and demonstrated that IFNγ induced changes in the peptidome, with an overlap of only approximately 50% between basal and treated cells. Around 6% to 8% of the peptides were identified as cis-spliced peptides, and 2,213 peptides (1,827 linear and 386 cis-spliced peptides) were derived from known melanoma-associated antigens. These peptide antigens were equally distributed between the constitutive- and IFNγ-induced peptidome. We next examined additional HLA-matched patient-derived cell lines to investigate how frequently these peptides were identified and found that a high proportion of both linear and spliced peptides was conserved between in idual patient tumors, drawing on data amassing to more than 100,000 peptide sequences. Several of these peptides showed in vitro immunogenicity across multiple patients with melanoma. These observations highlight the breadth and complexity of the repertoire of immunogenic peptides that can be exploited therapeutically and suggest that spliced peptides are a major class of tumor antigens.
Hydroxychloroquine inhibits the mitochondrial antioxidant system in activated T cells
Publisher: Elsevier BV
Date: 12-2021
Immune responses to abacavir in antigen-presenting cells from hypersensitive patients
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 06-2007
Most viral peptides displayed by class I MHC on infected cells are immunogenic
Publisher: Proceedings of the National Academy of Sciences
Date: 04-02-2019
Abstract: CD8 + T cells are essential effectors in antiviral immunity, recognizing short virus-derived peptides presented by MHC class I (pMHCI) on the surface of infected cells. However, the fraction of viral pMHCI on infected cells that are immunogenic has not been shown for any virus. To approach this fundamental question, we used peptide sequencing by high-resolution mass spectrometry to identify more than 170 vaccinia virus pMHCI presented on infected mouse cells. Next, we screened each peptide for immunogenicity in multiple virus-infected mice, revealing a wide range of immunogenicities. A surprisingly high fraction ( %) of pMHCI were immunogenic in at least one infected mouse, and nearly 40% were immunogenic across more than half of the mice screened. The high number of peptides found to be immunogenic and the distribution of responses across mice give us insight into the specificity of antiviral CD8 + T cell responses.
Direct quantitation of MHC-bound peptide epitopes by selected reaction monitoring
Publisher: Wiley
Date: 20-05-2011
Abstract: We describe a cell-free approach that employs selected reaction monitoring (SRM) in tandem mass spectrometry to identify and quantitate T-cell epitopes. This approach utilises multiple epitope-specific SRM transitions to identify known T-cell epitopes and an absolute quantitation (AQUA) peptide strategy to afford AQUA. The advantage of a mass spectrometry-based approach over more traditional cell-based assays resides in the robustness and transferability of an SRM approach between laboratories and the ability of this strategy to detect multiple peptides simultaneously without the requirement of epitope-specific reagents such as T-cell lines. Thus, the SRM strategy for epitope quantitation will find application in studies of antigen density, the link between epitope abundance and immunogenicity, the dynamic range of epitope presentation and the abundance of T-cell epitopes in disease.
Tadpole-like conformations of huntingtin exon 1 are characterized by conformational heterogeneity that persists regardless of polyglutamine length.
Publisher: Elsevier BV
Date: 05-2018
Peptidomimetics: modifying peptides in the pursuit of better vaccines
Publisher: Informa UK Limited
Date: 02-2011
DOI: 10.1586/ERV.10.161
Abstract: Peptide vaccines represent a potential strategy for the prevention and treatment of pathogenic diseases, cancers and autoimmune disorders their low cost, ease of synthesis and inherent safety are all attractive features. However, they have remained largely unsuccessful owing to low immunogenicity, predominantly stemming from reduced bioavailability due to rapid proteolytic degradation in vivo. The field of peptidomimetics, through chemical alteration of epitope structure, attempts to modify peptides to engender improved immunogenicity and stability, with the ultimate aim of rationally designing more efficacious vaccines. This article discusses the strategies employed in this process and recent advances within the field.
Crystal structure of the human T cell receptor CD3εγ heterodimer complexed to the therapeutic mAb OKT3
Publisher: Proceedings of the National Academy of Sciences
Date: 10-05-2004
Abstract: The CD3εγ heterodimer is essential for expression and function of the T cell receptor. The crystal structure of the human CD3εγ heterodimer is described to 2.1-Å resolution complexed with OKT3, a therapeutic mAb that not only activates and tolerizes mature T cells but also induces regulatory T cells. The mode of CD3εγ dimerization provides a general structural basis for CD3 assembly and maps candidate T cell antigen receptor docking sites, including a duplicated linear region rich in acidic residues that is unique to human CD3ε. OKT3 binds to an atypically small area of CD3ε and has a low affinity for the isolated CD3εγ heterodimer. The structure of the OKT3/CD3εγ complex has implications for T cell signaling and therapeutic design.
Complete modification of TCR specificity and repertoire selection does not perturb a CD8 + T cell immunodominance hierarchy
Publisher: Proceedings of the National Academy of Sciences
Date: 09-12-2008
Abstract: Understanding T cell immunodominance hierarchies is fundamental to the development of cellular-based vaccines and immunotherapy. A combination of influenza virus infection in C57BL/6J mice and reverse genetics is used here to dissect the role of T cell antigen receptor (TCR) repertoire in the immunodominant D b NP 366 CD8 + T cell response. Infection with an engineered virus (NPM6A) containing a single alanine (A) mutation at the critical p6 NP 366–374 residue induced a noncross-reactive CD8 + T cell response characterized by a novel, narrower TCR repertoire per in idual mouse that was nonetheless equivalent in magnitude to that generated after WT virus challenge. Although of lower overall avidity, the levels of both cytotoxic T lymphocyte activity and cytokine production were comparable with those seen for the native response. Importantly, the overdominance profile characteristic of secondary D b NP 366 -specific clonal expansions was retained for the NPM6A mutant. The primary determinants of immunodominance in this endogenous, non-TCR-transgenic model of viral immunity are thus independent of TCR repertoire composition and ersity. These findings both highlight the importance of effective antigen dose for T cell vaccination and/or immunotherapy and demonstrate the feasibility of priming the memory T cell compartment with engineered viruses to protect against commonly selected mutants viral (or tumor) escape mutants.
HLA-B57 micropolymorphism defines the sequence and conformational breadth of the immunopeptidome
Publisher: Springer Science and Business Media LLC
Date: 08-11-2018
DOI: 10.1038/S41467-018-07109-W
Abstract: Immunophenotypic differences between closely related human leukocyte antigen (HLA) alleles have been associated with ergent clinical outcomes in infection, autoimmunity, transplantation and drug hypersensitivity. Here we explore the impact of micropolymorphism on peptide antigen presentation by three closely related HLA molecules, HLA-B*57:01, HLA-B*57:03 and HLA-B*58:01, that are differentially associated with the HIV elite controller phenotype and adverse drug reactions. For each allotype, we mine HLA ligand data sets derived from the same parental cell proteome to define qualitative differences in peptide presentation using classical peptide binding motifs and an unbiased statistical approach. The peptide repertoires show marked qualitative overlap, with 982 peptides presented by all allomorphs. However, differences in peptide abundance, HLA-peptide stability, and HLA-bound conformation demonstrate that HLA micropolymorphism impacts more than simply the range of peptide ligands. These differences provide grounds for distinct immune reactivity and insights into the capacity of micropolymorphism to ersify immune outcomes.
HLA Class II Specificity Assessed by High-Density Peptide Microarray Interactions
Publisher: The American Association of Immunologists
Date: 07-2020
Abstract: The ability to predict and/or identify MHC binding peptides is an essential component of T cell epitope discovery, something that ultimately should benefit the development of vaccines and immunotherapies. In particular, MHC class I prediction tools have matured to a point where accurate selection of optimal peptide epitopes is possible for virtually all MHC class I allotypes in comparison, current MHC class II (MHC-II) predictors are less mature. Because MHC-II restricted CD4+ T cells control and orchestrated most immune responses, this shortcoming severely h ers the development of effective immunotherapies. The ability to generate large panels of peptides and subsequently large bodies of peptide–MHC-II interaction data are key to the solution of this problem, a solution that also will support the improvement of bioinformatics predictors, which critically relies on the availability of large amounts of accurate, erse, and representative data. In this study, we have used rHLA-DRB1*01:01 and HLA-DRB1*03:01 molecules to interrogate high-density peptide arrays, in casu containing 70,000 random peptides in triplicates. We demonstrate that the binding data acquired contains systematic and interpretable information reflecting the specificity of the HLA-DR molecules investigated, suitable of training predictors able to predict T cell epitopes and peptides eluted from human EBV-transformed B cells. Collectively, with a cost per peptide reduced to a few cents, combined with the flexibility of rHLA technology, this poses an attractive strategy to generate vast bodies of MHC-II binding data at an unprecedented speed and for the benefit of generating peptide–MHC-II binding data as well as improving MHC-II prediction tools.
High Resolution Structures of Highly Bulged Viral Epitopes Bound to Major Histocompatibility Complex Class I
Publisher: Elsevier BV
Date: 06-2005
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