ORCID Profile
0000-0003-2072-5968
Current Organisations
Centrum Wiskunde en Informatica
,
Technische Universiteit Delft
,
La Trobe 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 | Biological Physics | Cell Development, Proliferation and Death | Proteomics and Intermolecular Interactions (excl. Medical Proteomics) | Characterisation of Biological Macromolecules | Medicinal and Biomolecular Chemistry | Biomechanical Engineering | Crop and Pasture Protection (Pests, Diseases and Weeds) | Biomedical Engineering | Receptors and Membrane Biology | Biochemistry and Cell Biology not elsewhere classified | Structural Biology (incl. Macromolecular Modelling) |
ANZSRC Socio-Economic Objective (SEO)
Expanding Knowledge in the Biological Sciences | Environmentally Sustainable Plant Production not elsewhere classified | Expanding Knowledge in the Chemical Sciences | Expanding Knowledge in the Physical Sciences | Expanding Knowledge in Engineering | Infectious Diseases
Related Links
Publications
Bystander B cells rapidly acquire antigen receptors from activated B cells by membrane transfer
Publisher: Proceedings of the National Academy of Sciences
Date: 18-03-2008
Abstract: The B cell antigen receptor (BCR) efficiently facilitates the capture and processing of a specific antigen for presentation on MHC class II molecules to antigen-specific CD4 + T cells ( 1 ). Despite this, the majority of B cells are thought to play only a limited role in CD4 + T cell activation because BCRs are clonotypically expressed. Here, we show, however, that activated B cells can, both in vitro and in vivo , rapidly donate their BCR to bystander B cells, a process that is mediated by direct membrane transfer between adjacent B cells and is lified by the interaction of the BCR with a specific antigen. This results in a dramatic expansion in the number of antigen-binding B cells in vivo , with the transferred BCR endowing recipient B cells with the ability to present a specific antigen to antigen-specific CD4 + T cells.
Fine structure analysis of interaction of FcεRI with IgE
Publisher: Elsevier BV
Date: 05-1999
Divalent metal binding by histidine-rich glycoprotein differentially regulates higher order oligomerisation and proteolytic processing.
Publisher: Wiley
Date: 21-12-2017
Abstract: The serum protein histidine-rich glycoprotein (HRG) has been implicated in tissue injury and tumour growth. Several HRG functions are regulated by the alent metal Zn
Structure of the defensin NsD7 in complex with PIP 2 reveals that defensin : lipid oligomer topologies are dependent on lipid type
Publisher: Wiley
Date: 08-2017
Abstract: Defensins are innate immune molecules that upon recognition of specific phospholipids can disrupt microbial membranes by forming oligomeric assemblies. Structures of two related plant defensins, NaD1 and NsD7, bound to phosphatidylinositol 4,5-bisphosphate (PIP
Human β-Defensin 2 (HBD-2) Displays Oncolytic Activity but Does Not Affect Tumour Cell Migration
Publisher: MDPI AG
Date: 06-02-2022
DOI: 10.3390/BIOM12020264
Abstract: Defensins form an integral part of the cationic host defence peptide (HDP) family, a key component of innate immunity. Apart from their antimicrobial and immunomodulatory activities, many HDPs exert multifaceted effects on tumour cells, notably direct oncolysis and/or inhibition of tumour cell migration. Therefore, HDPs have been explored as promising anticancer therapeutics. Human β-defensin 2 (HBD-2) represents a prominent member of human HDPs, being well-characterised for its potent pathogen-killing, wound-healing, cytokine-inducing and leukocyte-chemoattracting functions. However, its anticancer effects remain largely unknown. Recently, we demonstrated that HBD-2 binds strongly to phosphatidylinositol-4,5-bisphosphate (PI(4,5)P2), a key mediator of defensin-induced cell death and an instructional messenger during cell migration. Hence, in this study, we sought to investigate the lytic and anti-migratory effects of HBD-2 on tumour cells. Using various cell biological assays and confocal microscopy, we showed that HBD-2 killed tumour cells via acute lytic cell death rather than apoptosis. In addition, our data suggested that, despite the reported PI(4,5)P2 interaction, HBD-2 does not affect cytoskeletal-dependent tumour cell migration. Together, our findings provide further insights into defensin biology and informs future defensin-based drug development.
Binding of phosphatidic acid by NsD7 mediates the formation of helical defensin-lipid oligomeric assemblies and membrane permeabilization
Publisher: Proceedings of the National Academy of Sciences
Date: 19-09-2016
Abstract: Direct attack of target cell membranes by protein oligomerization is a powerful innate defense mechanism used widely throughout nature. Defensins are ubiquitous innate immunity mediators that are able to recognize certain phospholipids, and subsequently oligomerize to attack target cell membranes. We now show that the plant defensin NsD7 is able to bind the cellular phospholipid, phosphatidic acid (PA), which triggers defensin oligomerization in a unique manner. Our crystal structure of the NsD7–PA oligomer revealed a striking double-helical defensin–lipid oligomer that features a novel phospholipid-binding site mediating PA binding and membrane permeabilization. This demonstrates that defensins use their conserved small fold in a remarkably flexible way to specifically recognize a range of phospholipids during innate defense using different binding sites.
Histidine-rich glycoprotein binds heparanase and regulates its enzymatic activity and cell surface interactions
Publisher: Elsevier BV
Date: 09-2010
DOI: 10.1016/J.BIOCEL.2010.05.008
Abstract: Heparanase, an endo-beta-D-glucuronidase, is involved in numerous normal physiological and pathological processes, such as inflammation, wound healing and tumour metastasis/angiogenesis, through its ability to mediate the degradation of heparan sulfate, a key structural component of the extracellular matrix and on the surface of cells. Identifying endogenous molecules that can regulate heparanase activity will aid the understanding of its molecular function in health and disease and provide the potential for development of novel anti-cancer and anti-inflammatory therapeutics. The ability of the extracellular heparanase to tether onto cell surface heparan sulfate proteoglycans and other receptor(s), such as the cation-independent mannose-6-phosphate receptor, is key to its activation, function and uptake into intracellular compartments. Here we describe experiments demonstrating that a relatively abundant plasma glycoprotein, histidine-rich glycoprotein, directly interacts with platelet-derived heparanase and enhances its enzymatic activity. The findings in this study also show that histidine-rich glycoprotein interferes with heparanase binding to cell surface receptors, particularly heparan sulfate proteoglycans. Thus, the interaction between histidine-rich glycoprotein and heparanase can potentially regulate the role of heparanase in a variety of physiological and pathological conditions.
Novel Formulation of Undecylenic Acid induces Tumor Cell Apoptosis.
Publisher: MDPI AG
Date: 16-11-2022
Abstract: Undecylenic acid, a monounsaturated fatty acid, is currently in clinical use as a topical antifungal agent, however the potential for therapeutic application in other disease settings has not been investigated. In this study, we describe a novel platform for the solubilization of fatty acids using amino acids and utilize this approach to define a tumoricidal activity and underlying mechanism for undecylenic acid. We examined a novel formulation of undecylenic acid compounded with L-Arginine, called GS-1, that induced concentration-dependent tumor cell death, with undecylenic acid being the cytotoxic component. Further investigation revealed that GS-1-mediated cell death was caspase-dependent with a reduction in mitochondrial membrane potential, suggesting a pro-apoptotic mechanism of action. Additionally, GS-1 was found to localize intracellularly to lipid droplets. In contrast to previous studies where lipid droplets have been shown to be protective against fatty acid-induced cell death, we showed that lipid droplets could not protect against GS-1-induced cytotoxicity. We also found a role for Fatty Acid Transport Protein 2 (FATP2) in the uptake of this compound. Collectively, this study demonstrates that GS-1 has effective pro-apoptotic antitumor activity in vitro and, together with the novel platform of fatty acid solubilization, contributes to the re-emerging field of fatty acids as potential anti-cancer therapeutics.
Towards novel herbicide modes of action by inhibiting lysine biosynthesis in plants
Publisher: eLife Sciences Publications, Ltd
Date: 27-07-2021
DOI: 10.7554/ELIFE.69444
Abstract: Weeds are becoming increasingly resistant to our current herbicides, posing a significant threat to agricultural production. Therefore, new herbicides with novel modes of action are urgently needed. In this study, we exploited a novel herbicide target, dihydrodipicolinate synthase (DHDPS), which catalyses the first and rate-limiting step in lysine biosynthesis. The first class of plant DHDPS inhibitors with micromolar potency against Arabidopsis thaliana DHDPS was identified using a high-throughput chemical screen. We determined that this class of inhibitors binds to a novel and unexplored pocket within DHDPS, which is highly conserved across plant species. The inhibitors also attenuated the germination and growth of A. thaliana seedlings and confirmed their pre-emergence herbicidal activity in soil-grown plants. These results provide proof-of-concept that lysine biosynthesis represents a promising target for the development of herbicides with a novel mode of action to tackle the global rise of herbicide-resistant weeds.
The phospholipid code: a key component of dying cell recognition, tumor progression and host–microbe interactions
Publisher: Springer Science and Business Media LLC
Date: 09-10-2015
DOI: 10.1038/CDD.2015.122
The plant defensin NaD1 introduces membrane disorder through a specific interaction with the lipid, phosphatidylinositol 4,5 bisphosphate
Publisher: Elsevier BV
Date: 06-2016
DOI: 10.1016/J.BBAMEM.2016.02.016
Abstract: Plant defensins interact with phospholipids in bilayers as part of their cytotoxic activity. Solanaceous class II defensins with the loop 5 sequence pattern "S-[KR]-[ILVQ]-[ILVQ]-[KR]-[KR]" interact with PI(4,5)P2. Here, the prototypical defensin of this class, NaD1, is used to characterise the biophysical interactions between these defensins and phospholipid bilayers. Binding of NaD1 to bilayers containing PI(4,5)P2 occurs rapidly and the interaction is very strong. Dual polarisation interferometry revealed that NaD1 does not dissociate from bilayers containing PI(4,5)P2. Binding of NaD1 to bilayers with or without PI(4,5)P2 induced disorder in the bilayer. However, permeabilisation assays revealed that NaD1 only permeabilised liposomes with PI(4,5)P2 in the bilayer, suggesting a role for this protein-lipid interaction in the plasma membrane permeabilising activity of this defensin. No defensins in the available databases have the PI(4,5)P2 binding sequence outside the solanaceous class II defensins, leading to the hypothesis that PI(4,5)P2 binding co-evolved with the C-terminal propeptide to protect the host cell against the effects of the tight binding of these defensins to their cognate lipid as they travel along the secretory pathway. This data has allowed us to develop a new model to explain how this class of defensins permeabilises plasma membranes to kill target cells.
ROCK1 but not LIMK1 or PAK2 is a key regulator of apoptotic membrane blebbing and cell disassembly
Publisher: Springer Science and Business Media LLC
Date: 05-2019
ACM international conference on interactive experiences for television and online video (ACM TVX 2014)
Publisher: Institute of Electrical and Electronics Engineers (IEEE)
Date: 10-2014
DOI: 10.1109/MMUL.2014.57
Yin Yang-1 inhibits tumor cell growth and inhibits p21 WAF1/Cip1complex formation with cdk4 and cyclin D1
Publisher: Spandidos Publications
Date: 09-02-2012
Abstract: The GLI-Krüppel zinc finger factor yin yang-1 (YY1) is a complex protein that regulates a variety of processes including transcription, proliferation, development and differentiation. YY1 inhibits cell growth in a cell type-specific manner. The role played by YY1 in its control of tumor cell growth is unclear and controversial. We show here that YY1 can suppress the growth of different tumor cell types in vitro, including human breast carcinoma cells and glioblastoma cells. YY1 also blocked the growth of 13762 MAT mammary adenocarcinoma isografts in rats. YY1 inhibited 13762 MAT tumor growth by approximately 80% compared with the GFP alone group 21 days after injection. YY1 inhibited proliferating cell nuclear antigen (PCNA) expression and pRbSer249/Thr252 phosphorylation without influencing tumor microvascular density. Moreover, YY1 inhibited p21WAF1/Cip1 complex formation with cdk4 and cyclin D1. These findings demonstrate that YY1 can negatively regulate the growth of multiple malignant cell types.
Heparanase and the hallmarks of cancer
Publisher: Springer Science and Business Media LLC
Date: 30-11-2020
DOI: 10.1186/S12967-020-02624-1
Abstract: Heparanase is the only mammalian enzyme that cleaves heparan sulphate, an important component of the extracellular matrix. This leads to the remodelling of the extracellular matrix, whilst liberating growth factors and cytokines bound to heparan sulphate. This in turn promotes both physiological and pathological processes such as angiogenesis, immune cell migration, inflammation, wound healing and metastasis. Furthermore, heparanase exhibits non-enzymatic actions in cell signalling and in regulating gene expression. Cancer is underpinned by key characteristic features that promote malignant growth and disease progression, collectively termed the ‘hallmarks of cancer’. Essentially, all cancers examined to date have been reported to overexpress heparanase, leading to enhanced tumour growth and metastasis with concomitant poor patient survival. With its multiple roles within the tumour microenvironment, heparanase has been demonstrated to regulate each of these hallmark features, in turn highlighting the need for heparanase-targeted therapies. However, recent discoveries which demonstrated that heparanase can also regulate vital anti-tumour mechanisms have cast doubt on this approach. This review will explore the myriad ways by which heparanase functions as a key regulator of the hallmarks of cancer and will highlight its role as a major component within the tumour microenvironment. The dual role of heparanase within the tumour microenvironment, however, emphasises the need for further investigation into defining its precise mechanism of action in different cancer settings.
Histidine-rich glycoprotein: The Swiss Army knife of mammalian plasma
Publisher: American Society of Hematology
Date: 17-02-2011
DOI: 10.1182/BLOOD-2010-09-303842
Abstract: Histidine-rich glycoprotein (HRG), also known as histidine-proline-rich glyco-protein, is an abundant and well-characterized protein of vertebrate plasma. HRG has a multidomain structure that allows the molecule to interact with many ligands, including heparin, phospholipids, plasminogen, fibrinogen, immunoglobulin G, C1q, heme, and Zn2+. The ability of HRG to interact with various ligands simultaneously has suggested that HRG can function as an adaptor molecule and regulate numerous important biologic processes, such as immune complex/necrotic cell athogen clearance, cell adhesion, angiogenesis, coagulation, and fibrinolysis. The present review covers the proposed multifunctional roles of HRG with a focus on recent findings that have led to its emergence as a key regulator of immunity and vascular biology. Also included is a discussion of the striking functional similarities between HRG and other important multifunctional proteins found in plasma, such as C-reactive protein, C1q, β2 glycoprotein I, and thrombospondin-1.
Monocyte apoptotic bodies are vehicles for influenza A virus propagation
Publisher: Springer Science and Business Media LLC
Date: 08-05-2020
DOI: 10.1038/S42003-020-0955-8
Abstract: The disassembly of apoptotic cells into small membrane-bound vesicles termed apoptotic bodies (ApoBDs) is a hallmark of apoptosis however, the functional significance of this process is not well defined. We recently discovered a new membrane protrusion (termed beaded apoptopodia) generated by apoptotic monocytes which fragments to release an abundance of ApoBDs. To investigate the function of apoptotic monocyte disassembly, we used influenza A virus (IAV) infection as a proof-of-concept model, as IAV commonly infects monocytes in physiological settings. We show that ApoBDs generated from IAV-infected monocytes contained IAV mRNA, protein and virions and consequently, could facilitate viral propagation in vitro and in vivo, and induce a robust antiviral immune response. We also identified an antipsychotic, Haloperidol, as an unexpected inhibitor of monocyte cell disassembly which could impair ApoBD-mediated viral propagation under in vitro conditions. Together, this study reveals a previously unrecognised function of apoptotic monocyte disassembly in the pathogenesis of IAV infections.
Gasdermin E Does Not Limit Apoptotic Cell Disassembly by Promoting Early Onset of Secondary Necrosis in Jurkat T Cells and THP-1 Monocytes
Publisher: Frontiers Media SA
Date: 04-12-2018
GS-2: A Novel Broad-Spectrum Agent for Environmental Microbial Control
Publisher: MDPI AG
Date: 13-09-2022
DOI: 10.3390/BIOM12091293
Abstract: The environmental control of microbial pathogens currently relies on compounds that do not exert long-lasting activity on surfaces, are impaired by soil, and contribute to the growing problem of antimicrobial resistance. This study presents the scientific development and characterization of GS-2, a novel, water-soluble ammonium carboxylate salt of capric acid and L-arginine that demonstrates activity against a range of bacteria (particularly Gram-negative bacteria), fungi, and viruses. In real-world surface testing, GS-2 was more effective than a benzalkonium chloride disinfectant at reducing the bacterial load on common touch-point surfaces in a high-traffic building (average 1.6 vs. 32.6 CFUs recovered from surfaces 90 min after application, respectively). Toxicology testing in rats confirmed GS-2 ingredients were rapidly cleared and posed no toxicities to humans or animals. To enhance the time-kill against Gram-positive bacteria, GS-2 was compounded at a specific ratio with a naturally occurring monoterpenoid, thymol, to produce a water-based antimicrobial solution. This GS-2 with thymol formulation could generate a bactericidal effect after five minutes of exposure and a viricidal effect after 10 min of exposure. Further testing of the GS-2 and thymol combination on glass slides demonstrated that the compound retained bactericidal activity for up to 60 days. Based on these results, GS-2 and GS-2 with thymol represent a novel antimicrobial solution that may have significant utility in the long-term reduction of environmental microbial pathogens in a variety of settings.
Monoclonal antibodies and synthetic peptides define the active site of Fc(ε)RI and a potential receptor antagonist
Publisher: Wiley
Date: 07-2000
DOI: 10.1034/J.1398-9995.2000.00485.X
Abstract: Defining the structure of the human high-affinity receptor for IgE, Fc,RI, is crucial to understand the receptor:ligand interaction, and to develop drugs to prevent IgE-dependent allergic diseases. To this end, a series of four anti-FcepsilonRI monoclonal antibodies (mAbs), including three new mAbs, 47, 54, and 3B4, were used in conjunction with synthetic FcepsilonRI peptides to define functional regions of the Fc IgE-binding site and identify an antagonist of IgE binding. The spatial orientation of the epitopes detected by these antibodies and their relationship to the IgE-binding region of FcepsilonRI was defined by a homology model based on the closely related FcepsilonRIIa. Using recombinant soluble FcRI-alpha as well as FcepsilonRI-alpha expressed on the cell surface, a series of direct and competitive binding experiments indicated that the mAbs detected nonoverlapping epitopes. One antibody (15-1), previously thought to be located close to the IgE-binding site, was precisely mapped to a single loop within the IgE-binding site by both mutagenesis and overlapping synthetic peptides encompassing the entire extracellular domain. A synthetic peptide epsilonRI-11, containing the amino acids 101-120 and the mAb 15-1 epitope, inhibited IgE binding and may form the basis for the development of a useful receptor-based therapy.
Mutagenesis within human FcεRIα differentially affects human and murine IgE binding
Publisher: The American Association of Immunologists
Date: 15-02-2002
DOI: 10.4049/JIMMUNOL.168.4.1787
Abstract: Soluble fragments of the α-chain of FcεRI, the high-affinity receptor for IgE, compete with membrane-bound receptors for IgE and may thus provide a means to combat allergic responses. Mutagenesis within FcεRIα is used in this study, in conjunction with the crystal structure of the FcεRIα/IgE complex, to define the relative importance of specific residues within human FcεRIα for IgE binding. We have also compared the effects of these mutants on binding to both human and mouse IgE, with a view to evaluating the mouse as an appropriate model for the analysis of future agents designed to mimic the human FcεRIα and attenuate allergic disease. Three residues within the C-C′ region of the FcεRI α2 domain and two residues within the α2 proximal loops of the α1 domain were selected for mutagenesis and tested in binding assays with human and mouse IgE. All three α2 mutations (K117D, W130A, and Y131A) reduced the affinity of human IgE binding to different extents, but K117D had a far more pronounced effect on mouse IgE binding, and although Y131A had little effect, W130A modestly enhanced binding to mouse IgE. The mutations in α1 (R15A and F17A) diminished binding to both human and mouse IgE, with these effects most likely caused by disruption of the α1/α2 interface. Our results demonstrate that the effects of mutations in human FcεRIα on mouse IgE binding, and hence the inhibitory properties of human receptor-based peptides assayed in rodent models of allergy, may not necessarily reflect their activity in a human IgE-based system.
Nicotiana alata Defensin Chimeras Reveal Differences in the Mechanism of Fungal and Tumor Cell Killing and an Enhanced Antifungal Variant.
Publisher: American Society for Microbiology
Date: 10-2016
DOI: 10.1128/AAC.01479-16
Abstract: The plant defensin NaD1 is a potent antifungal molecule that also targets tumor cells with a high efficiency. We examined the features of NaD1 that contribute to these two activities by producing a series of chimeras with NaD2, a defensin that has relatively poor activity against fungi and no activity against tumor cells. All plant defensins have a common tertiary structure known as a cysteine-stabilized α-β motif which consists of an α helix and a triple-stranded β-sheet stabilized by four disulfide bonds. The chimeras were produced by replacing loops 1 to 7, the sequences between each of the conserved cysteine residues on NaD1, with the corresponding loops from NaD2. The loop 5 swap replaced the sequence motif (SKILRR) that mediates tight binding with phosphatidylinositol 4,5-bisphosphate [PI(4,5)P 2 ] and is essential for the potent cytotoxic effect of NaD1 on tumor cells. Consistent with previous reports, there was a strong correlation between PI(4,5)P 2 binding and the tumor cell killing activity of all of the chimeras. However, this correlation did not extend to antifungal activity. Some of the loop swap chimeras were efficient antifungal molecules, even though they bound poorly to PI(4,5)P 2 , suggesting that additional mechanisms operate against fungal cells. Unexpectedly, the loop 1B swap chimera was 10 times more active than NaD1 against filamentous fungi. This led to the conclusion that defensin loops have evolved as modular components that combine to make antifungal molecules with variable mechanisms of action and that artificial combinations of loops can increase antifungal activity compared to that of the natural variants.
Phosphoinositide-mediated oligomerization of a defensin induces cell lysis
Publisher: eLife Sciences Publications, Ltd
Date: 04-2014
DOI: 10.7554/ELIFE.01808
Abstract: Cationic antimicrobial peptides (CAPs) such as defensins are ubiquitously found innate immune molecules that often exhibit broad activity against microbial pathogens and mammalian tumor cells. Many CAPs act at the plasma membrane of cells leading to membrane destabilization and permeabilization. In this study, we describe a novel cell lysis mechanism for fungal and tumor cells by the plant defensin NaD1 that acts via direct binding to the plasma membrane phospholipid phosphatidylinositol 4,5-bisphosphate (PIP2). We determined the crystal structure of a NaD1:PIP2 complex, revealing a striking oligomeric arrangement comprising seven dimers of NaD1 that cooperatively bind the anionic headgroups of 14 PIP2 molecules through a unique ‘cationic grip’ configuration. Site-directed mutagenesis of NaD1 confirms that PIP2-mediated oligomerization is important for fungal and tumor cell permeabilization. These observations identify an innate recognition system by NaD1 for direct binding of PIP2 that permeabilizes cells via a novel membrane disrupting mechanism.
Cytotoxic properties of rhenium(I) tricarbonyl complexes of N-heterocyclic carbene ligands.
Publisher: Royal Society of Chemistry (RSC)
Date: 2022
DOI: 10.1039/D2DT00447J
Abstract: A family of eight rhenium(I) tricarbonyl complexes bearing pyridyl-imidazolylidene or bis-imidazolylidene ligands in combination with a series of
Histidine-rich glycoprotein: A novel adaptor protein in plasma that modulates the immune, vascular and coagulation systems
Publisher: Wiley
Date: 04-2005
DOI: 10.1111/J.1440-1711.2005.01320.X
Abstract: Histidine-rich glycoprotein (HRG) is an abundant plasma glycoprotein that has a multidomain structure, interacts with many ligands, and has been shown to regulate a number of important biological processes. HRG ligands include Zn(2+) and haem, tropomyosin, heparin and heparan sulphate, plasminogen, plasmin, fibrinogen, thrombospondin, IgG, FcgammaR and complement. In many cases, the histidine-rich region of the molecule enhances ligand binding following interaction with Zn(2+) or exposure to low pH, conditions associated with sites of tissue injury or tumour growth. The multidomain nature of HRG indicates that it can act as an extracellular adaptor protein, bringing together disparate ligands, particularly on cell surfaces. HRG binds to most cells primarily via heparan sulphate proteoglycans, binding which is also potentiated by elevated free Zn(2+) levels and low pH. Recent reports have shown that HRG can modulate angiogenesis and additional studies have shown that it may regulate other physiological processes such as cell adhesion and migration, fibrinolysis and coagulation, complement activation, immune complex clearance and phagocytosis of apoptotic cells. This review outlines the molecular, structural, biological and clinical properties of HRG as well as describing the role of HRG in various physiological processes.
Human beta-defensin 3 contains an oncolytic motif that binds PI(4,5)P-2 to mediate tumour cell permeabilisation
Publisher: Impact Journals, LLC
Date: 09-12-2016
Histidine-rich glycoprotein functions cooperatively with cell surface heparan sulfate on phagocytes to promote necrotic cell uptake
Publisher: Oxford University Press (OUP)
Date: 23-06-2010
DOI: 10.1189/JLB.0210087
Abstract: HRG enhances the phagocytosis of necrotic cells via a heparan sulfate-dependent pathway that is inhibitable by heparin. Dying cells, such as apoptotic and necrotic cells, are cleared rapidly from the site of cell death to prevent the exposure of intracellular antigenic and immunostimulatory molecules that may cause tissue injury or facilitate the development of autoimmune diseases. For the immune system to recognize and remove dying cells efficiently, professional phagocytes use a variety of mechanisms that distinguish healthy cells from dying cells. HRG, a relatively abundant heparin/HS-binding protein in human plasma, has been shown recently to tether IgG specifically to necrotic cells and aid the phagocytic uptake of necrotic cells via a FcγRI-dependent pathway. In this study, we provide direct evidence that HRG can function cooperatively with cell surface HS on the monocytic cell line THP-1 to promote necrotic cell removal. In addition, we found that the presence of heparin can markedly inhibit HRG-enhanced necrotic cell clearance by THP-1 cells, possibly by blocking the ability of HRG to interact with necrotic cells as well as THP-1 cells. Thus, these data suggest that HRG can aid the phagocytosis of necrotic cells via a HS-dependent pathway, and this process can be regulated by the presence of certain HRG ligands, such as heparin.
Moving beyond size and phosphatidylserine exposure: evidence for a diversity of apoptotic cell‐derived extracellular vesicles in vitro
Publisher: Wiley
Date: 26-04-2019
The plant defensin NaD1 induces tumor cell death via a non-apoptotic, membranolytic process
Publisher: Springer Science and Business Media LLC
Date: 23-01-2017
DOI: 10.1038/CDDISCOVERY.2016.102
Abstract: Cationic anti-microbial peptides (CAPs) have an important role in host innate defense against pathogens such as bacteria and fungi. Many CAPs including defensins also exhibit selective cytotoxic activity towards mammalian cells via both apoptotic and non-apoptotic processes, and are being investigated as potential anticancer agents. The anti-fungal plant defensin from ornamental tobacco, Nicotiana alata Defensin 1 (NaD1), was recently shown to induce necrotic-like cell death in a number of tumor cell types within 30 min of treatment, at a concentration of 10 μ M. NaD1-mediated cell killing within these experimental parameters has been shown to occur via binding to the plasma membrane phosphatidylinositol 4,5-bisphosphate (PIP2) in target cells to facilitate membrane destabilization and subsequent lysis. Whether NaD1 is also capable of inducing apoptosis in tumor cells has not been reported previously. In this study, treatment of MM170 (melanoma) and Jurkat T (leukemia) cells with subacute ( μ M) concentrations of NaD1 over 6–24 h was investigated to determine whether NaD1 could induce cell death via apoptosis. At subacute concentrations, NaD1 did not efficiently induce membrane permeabilization within 30 min, but markedly reduced cell viability over 24 h. In contrast to other CAPs that have been shown to induce apoptosis through caspase activation, dying cells were not sensitive to a pancaspase inhibitor nor did they display caspase activity or DNA fragmentation over the 24 h treatment time. Furthermore, over the 24 h period, cells exhibited necrotic phenotypes and succumbed to membrane permeabilization. These results indicate that the cytotoxic mechanism of NaD1 at subacute concentrations is membranolytic rather than apoptotic and is also likely to be mediated through a PIP2-targeting cell lytic pathway.
Structural and functional characterization of the membrane-permeabilizing activity of Nicotiana occidentalis defensin NoD173 and protein engineering to enhance oncolysis
Publisher: Wiley
Date: 22-02-2019
The Tomato Defensin TPP3 Binds Phosphatidylinositol (4,5)-Bisphosphate via a Conserved Dimeric Cationic Grip Conformation To Mediate Cell Lysis
Publisher: Informa UK Limited
Date: 06-2015
DOI: 10.1128/MCB.00282-15
Regulation of histidine-rich glycoprotein (HRG) function via plasmin-mediated proteolytic cleavage
Publisher: Portland Press Ltd.
Date: 23-10-2009
DOI: 10.1042/BJ20090794
Abstract: The plasminogen lasmin system is involved in a variety of normal physiological and pathological processes, including tissue remodelling, angiogenesis and tumour metastasis. Plasminogen activators and receptors for plasminogen lasminogen activators are essential for the processing of plasminogen to form the active serine protease plasmin. Plasmin can in turn positively or negatively regulate further plasminogen activation via plasminmediated cleavage of receptors and activators. HRG (histidine-rich glycoprotein), a relatively abundant (approx. 100–150 μg/ml) plasma glycoprotein, has a multi-domain structure that can interact with many ligands, including Zn2+, heparin, HS (heparan sulfate) and plasminogen. HRG has been shown to function as an adaptor molecule to tether plasminogen to GAG (glycosaminoglycan)-bearing surfaces and to regulate plasminogen activation via various mechanisms. As HRG itself is sensitive to plasmin cleavage, the present study examines in detail the cleavage of human HRG by plasmin and the effect of this cleavage on various functions of HRG. HRG fragments, generated by plasmin cleavage, are held together by disulfide linkages and are not released from the molecule under non-reducing conditions. Plasmin-mediated cleavage partially inhibited HRG binding to cell surface HS, but enhanced HRG binding to necrotic cells and to plasminogen. However, both intact and plasmin-cleaved HRG enhanced the binding of plasminogen to heparin-coated surfaces to a similar extent. Furthermore, the presence of heparin, Zn2+ or acidic pH was found to protect HRG from plasmin cleavage. Thus proteolytic cleavage of HRG by plasmin may provide a feedback mechanism to regulate the effects of HRG on the plasminogen lasmin system and other functions of HRG.
Cloning of mammalian heparanase, an important enzyme in tumor invasion and metastasis
Publisher: Springer Science and Business Media LLC
Date: 07-1999
DOI: 10.1038/10525
Abstract: The endoglycosidase heparanase is an important in the degradation of the extracellular matrix by invading cells, notably metastatic tumor cells and migrating leukocytes. Here we report the cDNA sequence of the human platelet enzyme, which encodes a unique protein of 543 amino acids, and the identification of highly homologous sequences in activated mouse T cells and in a highly metastatic rat adenocarcinoma. Furthermore, the expression of heparanase mRNA in rat tumor cells correlates with their metastatic potential. Exhaustive studies have shown only one heparanase sequence, consistent with the idea that this enzyme is the dominant endoglucuronidase in mammalian tissues.
Dimerization of Plant Defensin NaD1 Enhances Its Antifungal Activity
Publisher: Elsevier BV
Date: 06-2012
Leukocyte Heparanase: A Double-Edged Sword in Tumor Progression
Publisher: Frontiers Media SA
Date: 29-04-2019
Defining the role of cytoskeletal components in the formation of apoptopodia and apoptotic bodies during apoptosis
Publisher: Springer Science and Business Media LLC
Date: 05-09-2019
DOI: 10.1007/S10495-019-01565-5
Abstract: During apoptosis, dying cells undergo dynamic morphological changes that ultimately lead to their disassembly into fragments called apoptotic bodies (ApoBDs). Reorganisation of the cytoskeletal structures is key in driving various apoptotic morphologies, including the loss of cell adhesion and membrane bleb formation. However, whether cytoskeletal components are also involved in morphological changes that occur later during apoptosis, such as the recently described generation of thin apoptotic membrane protrusions called apoptopodia and subsequent ApoBD formation, is not well defined. Through monitoring the progression of apoptosis by confocal microscopy, specifically focusing on the apoptopodia formation step, we characterised the presence of F-actin and microtubules in a subset of apoptopodia generated by T cells and monocytes. Interestingly, targeting actin polymerisation and microtubule assembly pharmacologically had no major effect on apoptopodia formation. These data demonstrate apoptopodia as a novel type of membrane protrusion that could be formed in the absence of actin polymerisation and microtubule assembly.
Recombinant expression and purification of the tomato defensin TPP3 and its preliminary X-ray crystallographic analysis
Publisher: International Union of Crystallography (IUCr)
Date: 22-02-2012
Multiple regions of human FcγRII (CD32) contribute to the binding of IgG
Publisher: Elsevier BV
Date: 09-1995
Repurposed inhibitor of bacterial dihydrodipicolinate reductase exhibits effective herbicidal activity
Publisher: Springer Science and Business Media LLC
Date: 22-05-2023
DOI: 10.1038/S42003-023-04895-Y
Abstract: Herbicide resistance represents one of the biggest threats to our natural environment and agricultural sector. Thus, new herbicides are urgently needed to tackle the rise in herbicide-resistant weeds. Here, we employed a novel strategy to repurpose a ‘failed’ antibiotic into a new and target-specific herbicidal compound. Specifically, we identified an inhibitor of bacterial dihydrodipicolinate reductase (DHDPR), an enzyme involved in lysine biosynthesis in plants and bacteria, that exhibited no antibacterial activity but severely attenuated germination of the plant Arabidopsis thaliana . We confirmed that the inhibitor targets plant DHDPR orthologues in vitro, and exhibits no toxic effects against human cell lines. A series of analogues were then synthesised with improved efficacy in germination assays and against soil-grown A. thaliana . We also showed that our lead compound is the first lysine biosynthesis inhibitor with activity against both monocotyledonous and dicotyledonous weed species, by demonstrating its effectiveness at reducing the germination and growth of Lolium rigidum (rigid ryegrass) and Raphanus raphanistrum (wild radish). These results provide proof-of-concept that DHDPR inhibition may represent a much-needed new herbicide mode of action. Furthermore, this study exemplifies the untapped potential of repurposing ‘failed’ antibiotic scaffolds to fast-track the development of herbicide candidates targeting the respective plant enzymes.
Endothelial cell apoptosis and the role of endothelial cell-derived extracellular vesicles in the progression of atherosclerosis
Publisher: Springer Science and Business Media LLC
Date: 19-12-2018
DOI: 10.1007/S00018-018-2983-9
Abstract: To maintain physiological homeostasis, cell turnover occurs every day in the body via a form of programmed cell death called apoptosis. During apoptosis, cells undergo distinct morphological changes culminating in the disassembly of the dying cell into smaller fragments known as apoptotic bodies (ApoBDs). Dysregulation of apoptosis is associated with diseases including infection, cancer and atherosclerosis. Although the development of atherosclerosis is largely attributed to the accumulation of lipids and inflammatory debris in vessel walls, it is also associated with apoptosis of macrophages, smooth muscle cells (SMCs) and endothelial cells. During cellular activation and apoptosis, endothelial cells can release several types of membrane-bound extracellular vesicles (EVs) including exosomes, microvesicles (MVs)/microparticles and ApoBDs. Emerging evidence in the field suggests that these endothelial cell-derived EVs (EndoEVs) can contribute to intercellular communication during the development of atherosclerosis via the transfer of cellular contents such as protein and microRNA, which may prevent or promote disease progression depending on the context. This review provides an up-to-date overview of the known causes and consequences of endothelial cell death during atherosclerosis along with highlighting current methodological approaches to studying EndoEVs and the potential roles of EndoEVs in atherosclerosis development.
Comparative proteomics evaluation of plasma exosome isolation techniques and assessment of the stability of exosomes in normal human blood plasma
Publisher: Wiley
Date: 18-10-2013
Abstract: Exosomes are nanovesicles released by a variety of cells and are detected in body fluids including blood. Recent studies have highlighted the critical application of exosomes as personalized targeted drug delivery vehicles and as reservoirs of disease biomarkers. While these research applications have created significant interest and can be translated into practice, the stability of exosomes needs to be assessed and exosome isolation protocols from blood plasma need to be optimized. To optimize methods to isolate exosomes from blood plasma, we performed a comparative evaluation of three exosome isolation techniques (differential centrifugation coupled with ultracentrifugation, epithelial cell adhesion molecule immunoaffinity pull-down, and OptiPrep(TM) density gradient separation) using normal human plasma. Based on MS, Western blotting and microscopy results, we found that the OptiPrep(TM) density gradient method was superior in isolating pure exosomal populations, devoid of highly abundant plasma proteins. In addition, we assessed the stability of exosomes in plasma over 90 days under various storage conditions. Western blotting analysis using the exosomal marker, TSG101, revealed that exosomes are stable for 90 days. Interestingly, in the context of cellular uptake, the isolated exosomes were able to fuse with target cells revealing that they were indeed biologically active.
Human mast cell line-1 (HMC-1) cells transfected with FcεRIα are sensitive to IgE/antigen-mediated Stimulation demonstrating selectivity towards cytokine production
Publisher: Elsevier BV
Date: 08-2011
DOI: 10.1016/J.INTIMP.2011.02.017
Abstract: Mast cells play important roles in allergic and inflammatory diseases. Efforts to better understand human mast cell activation and develop novel inhibitory agents have been h ered by the lack of suitable human mast cell lines. The HMC-1 mast cell line has been extensively used, but lacks native expression of the human high-affinity IgE receptor FcεRI limiting its applications. We have stably transfected HMC-1 cells with the IgE-binding α-subunit of FcεRI to generate HMCα cells that are antigen-responsive. We have used flow cytometry, cell signaling assays, pharmacological pathway inhibitors and cell functional assays to characterize the properties of HMCα cells. IgE/antigen responses were compared with those of the adenosine receptor agonist NECA. Surface expression of FcεRI in HMCα cells was demonstrated and was enhanced by prior sensitization with IgE. Activation of HMCα cells with IgE/antigen did not produce degranulation, but did lead to release of numerous cytokines. Whilst there was no measurable increase of intracellular Ca(2+) or marked general changes in protein tyrosine phosphorylation, IgE/antigen stimulation of HMCα cells enhanced phosphorylation of p38(MAPK) and Erk. Inhibitors of these pathways, as well as the src kinase inhibitor PP2, attenuated IgE/antigen-induced cytokine release. In summary, we have generated and characterized HMCα cells and show that they are a useful and relevant human mast cell model to examine FcεRI stabilization, signaling and mediator release. We envisage that HMCα cells will have utility in understanding the importance of mast cells in human allergic disease and in assessing the activity of novel anti-allergic compounds.
Determining the contents and cell origins of apoptotic bodies by flow cytometry
Publisher: Springer Science and Business Media LLC
Date: 31-10-2017
DOI: 10.1038/S41598-017-14305-Z
Abstract: Over 200 billion cells undergo apoptosis every day in the human body in order to maintain tissue homeostasis. Increased apoptosis can also occur under pathological conditions including infection and autoimmune disease. During apoptosis, cells can fragment into subcellular membrane-bound vesicles known as apoptotic bodies (ApoBDs). We recently developed a flow cytometry-based method to accurately differentiate ApoBDs from other particles (e.g. cells and debris). In the present study, we aim to further characterize subsets of ApoBDs based on intracellular contents and cell type-specific surface markers. Utilizing a flow cytometry-based approach, we demonstrated that intracellular contents including nuclear materials and mitochondria are distributed to some, but not all ApoBDs. Interestingly, the mechanism of ApoBD formation could affect the distribution of intracellular contents into ApoBDs. Furthermore, we also showed that ApoBDs share the same surface markers as their cell of origin, which can be used to distinguish cell type-specific ApoBDs from a mixed culture. These studies demonstrate that ApoBDs are not homogeneous and can be ided into specific subclasses based on intracellular contents and cell surface markers. The described flow cytometry-based method to study ApoBDs could be used in future studies to better understand the function of ApoBDs.
Importance of phosphoinositide binding by human β-defensin 3 for Akt-dependent cytokine induction.
Publisher: Wiley
Date: 15-12-2018
DOI: 10.1111/IMCB.1017
Abstract: Host defense peptides (HDPs) are well-characterized for their antimicrobial activities but also variously display potent immunomodulatory effects. Human β-defensin 3 (HBD-3) belongs to a well-known HDP family known as defensins and is able to induce leukocyte chemotactic recruitment, leukocyte activation/maturation, proinflammatory cytokine release, and co-stimulatory marker expression. HBD-3-stimulated cytokine induction is NF-κB-dependent and was initially suggested to act via G protein-coupled C-C chemokine receptor phospholipase C (PLC) and/or Toll-like receptor signaling. Subsequent pharmacological inhibition, however, revealed that NF-κB activation by HBD-3 is receptor-independent and instead involves the phosphoinositide 3-kinase (PI3K)-protein kinase B (Akt) pathway, the mechanism of which remains undetermined. Recently, we have shown that HBD-3 can enter mammalian cells and bind to inner membrane phosphoinositide 4,5-bisphosphate [PI(4,5)P
New method for purifying histidine‐rich glycoprotein from human plasma redefines its functional properties
Publisher: Wiley
Date: 10-04-2013
DOI: 10.1002/IUB.1168
Abstract: Histidine-rich glycoprotein (HRG) is a relatively abundant plasma protein that has been implicated in multiple biological processes including immunity, tumor progression, and vascular biology. However, current protocols for purifying HRG from plasma result in the copurification of contaminating proteins and raise questions over the validity of biological activities ascribed to HRG. In this study, we describe a two-step protocol for the large-scale purification of HRG from human plasma using a combination of metal affinity and ion exchange chromatography. The protocol employs a rapid and simple strategy to isolate highly purified HRG that minimizes proteolytic cleavage of the protein. The purification of HRG was assessed at each stage by measuring the amount of HRG immunoreactive protein using a specific monoclonal antibody against total protein, and demonstrated ~1,000-fold purification with an overall yield of ~32%. Mass spectrometry analysis demonstrated that plasma-derived HRG was free of contaminating proteins and gel electrophoresis showed it to have minimal proteolytic degradation. Characterization of protein by physical method showed that the protein exists as a single, monodisperse species. In contrast to the previous studies of HRG purified by different methods, HRG purified using the new procedure demonstrated a reduced profile of functions. Although the HRG retained binding to heparin and phosphatidic acid, it did not interact with necrotic cells or other cellular lipids. These data demonstrate that HRG does not exhibit the broad interactive properties that have been reported previously, suggesting that copurification of HRG-binding partners or other impurities are responsible for some of the reported functional properties. The findings in this study demonstrate that the new purification procedure can provide a ready source of pure HRG to assess ligand specificity and biological function of this important plasma protein.
ICAM-1-dependent pathways regulate colonic eosinophilic inflammation
Publisher: Oxford University Press (OUP)
Date: 26-05-2006
DOI: 10.1189/JLB.1105643
Abstract: Eosinophilic inflammation is a common feature of numerous eosinophil-associated gastrointestinal (EGID) diseases. Central to eosinophil migration into the gastrointestinal tract are the integrin-mediated interactions with adhesion molecules. Although the mechanisms regulating eosinophil homing into the small intestine have begun to be elucidated, the adhesion pathways responsible for eosinophil trafficking into the large intestine are unknown. We investigated the role of adhesion pathways in eosinophil recruitment into the large intestine during homeostasis and disease. First, using a hapten-induced colonic injury model, we demonstrate that in contrast to the small intestine, eosinophil recruitment into the colon is regulated by a β7-integrin addressin cell adhesion molecule-1-independent pathway. Characterization of integrin expression on colonic eosinophils by flow cytometry analysis revealed that colonic CC chemokine receptor 3+ eosinophils express the intercellular adhesion molecule-1 (ICAM-1) counter-receptor integrins αL, αM, and β2. Using ICAM-1-deficient mice and anti-ICAM-1 neutralizing antibodies, we show that hapten-induced colonic eosinophilic inflammation is critically dependent on ICAM-1. These studies demonstrate that β2-integrin/ICAM-1-dependent pathways are integral to eosinophil recruitment into the colon during GI inflammation associated with colonic injury.
Isolation, tissue distribution, and chromosomal localization of a novel testis-specific human four-transmembrane gene related to CD20 and FcεRI-β
Publisher: Elsevier BV
Date: 2001
Analysis of extracellular vesicles generated from monocytes under conditions of lytic cell death
Publisher: Springer Science and Business Media LLC
Date: 17-05-2019
DOI: 10.1038/S41598-019-44021-9
Abstract: Extracellular vesicles (EVs) are an important class of membrane-bound structures that have been widely investigated for their roles in intercellular communication in the contexts of tumor progression, vascular function, immunity and regenerative medicine. Much of the current knowledge on the functions of EVs pertains to those derived from viable cells (e.g. exosomes and microvesicles) or apoptotic cells (e.g. apoptotic bodies) whilst the generation of EVs from dying cells under non-apoptotic conditions remains poorly characterized. Herein, the release of EVs from THP-1 monocytes under conditions of primary necrosis, secondary necrosis and pyroptosis, was investigated. A comprehensive analysis of THP-1-derived EVs revealed that cells undergoing lytic forms of cell death generated a high number of EVs compared with viable or apoptotic cells in vitro . Differential centrifugation via 16,000 g and 100,000 g revealed that dying THP-1 cells release both medium and small EVs, respectively, consistent with the known characteristics of microvesicles and/or exosomes. In addition, large EVs isolated via 2000 g centrifugation were also present in all s les. These findings suggest that lytic cell death under both sterile and non-sterile inflammatory conditions induces monocytes to generate EVs, which could potentially act as mediators of cell-to-cell communication.
Characterization of FcR Ig-binding sites and epitope mapping.
Publisher: Elsevier BV
Date: 02-1994
Abstract: The low-affinity receptor for IgG, Fc gamma RII, and the high-affinity receptor for IgE, Fc epsilon RI, are functionally distinct but structurally homologous receptors. These characteristics have been exploited using a chimeric receptor strategy to examine segments of human Fc gamma RII for IgG-binding function. A series of chimeric receptors was generated by exchanging coding regions of the extracellular ligand-binding regions between Fc gamma RII and the Fc epsilon RI alpha chain using splice overlap extension by the polymerase chain reaction. The expression of these chimeric receptors in COS-7 cells and analysis of their IgG/IgE binding capacities have enabled the Ig-binding region of Fc gamma RII to be localized to a subregion of the second extracellular domain. The localization of the Ig-binding region of Fc gamma RII has provided the opportunity of performing site-directed mutagenesis to determine the key amino acids involved in the interaction of the receptor with IgG. These findings demonstrate that the chimeric receptor approach is a powerful technique for the dissection of structure/function relationships of structurally related yet functionally different molecules.
Chimeric Fc receptors identify immunoglobulin‐binding regions in human FcγRII and FcϵRI
Publisher: Wiley
Date: 03-1993
Abstract: Fc gamma RII and Fc epsilon RI are functionally distinct cell surface receptors for immunoglobulin (Ig) Fc gamma RII binds IgG with low affinity, whereas Fc epsilon RI binds IgE with high affinity, yet they are homologous in structure and sequence having extracellular regions containing two Ig-like domains with 38% amino acid identity. Chimeric receptors derived from human Fc gamma RII and Fc epsilon RI were produced by exchanging homologous regions of the two receptors to define binding region(s) for IgG in Fc gamma RII and IgE in Fc epsilon RI. Firstly, a chimeric form of the Fc epsilon RI alpha chain was produced by replacing the transmembrane region and cytoplasmic tail with that of Fc gamma RII. This mutant alpha chain could be expressed on the cell surface independently of associated beta and gamma subunits, and retained high-affinity IgE binding, indicating that the extracellular region of the Fc epsilon RI alpha chain is sufficient for high-affinity IgE binding. Secondly, to identify the role of the in idual domains in Fc binding of both Fc gamma RII and Fc epsilon RI, chimeric receptors were generated by exchanging the first extracellular domains between Fc gamma RII and the alpha chain mutant and used to demonstrate that the second extracellular domain of both receptors contains region(s) directly involved in Ig binding. Additional chimeric receptors were constructed to localize the Ig interactive regions in domain two of Fc gamma RII and Fc epsilon RI these identified a single region of IgG binding in Fc gamma RII located between residues Ser136 to Val169, and at least three independent IgE binding regions in the Fc epsilon RI alpha chain, between residues Trp87 to Lys128, Tyr129 to Asp145, and Ser146 to Val169.
Plasminogen Is Tethered with High Affinity to the Cell Surface by the Plasma Protein, Histidine-rich Glycoprotein
Publisher: Elsevier BV
Date: 09-2004
Defining the morphologic features and products of cell disassembly during apoptosis
Publisher: Springer Science and Business Media LLC
Date: 19-01-2017
Murine histidine-rich glycoprotein: Cloning, characterization and cellular origin
Publisher: Wiley
Date: 06-2000
DOI: 10.1046/J.1440-1711.2000.00940.X
Abstract: Histidine-rich glycoprotein (HRG) is a plasma protein of vertebrates that has been implicated in the regulation of several important biological functions, including the immune response and blood clotting. In the present study, we have isolated and determined the sequence of the cDNAs for both mouse and rat HRG. The deduced amino acid sequences of mouse and rat HRG are 525 and 510 amino acids, respectively, and they show the same three-domain structure that has been predicted for human HRG, with which they share high amino acid identity. Northern blot analysis indicates that the mouse HRG mRNA is 1.7 kb and is localized specifically to the liver. It has been suggested, somewhat controversially, that some immune cells, such as monocytes and megakaryocytes, also synthesize HRG. Reverse transcriptase-polymerase chain reaction analysis has failed to show any HRG mRNA in immune tissues of the mouse, including the spleen, thymus, lymph node, bone marrow and peripheral blood leucocytes. These data suggest that HRG expression by immune cells is due to the acquisition of plasma HRG derived from the liver. Finally, genomic Southern blot analysis of the mouse HRG gene suggests that it is a single copy gene.
A 4-cyano-3-methylisoquinoline inhibitor of Plasmodium falciparum growth targets the sodium efflux pump PfATP4
Publisher: Springer Science and Business Media LLC
Date: 16-07-2019
DOI: 10.1038/S41598-019-46500-5
Abstract: We developed a novel series of antimalarial compounds based on a 4-cyano-3-methylisoquinoline. Our lead compound MB14 achieved modest inhibition of the growth in vitro of the human malaria parasite, Plasmodium falciparum . To identify its biological target we selected for parasites resistant to MB14. Genome sequencing revealed that all resistant parasites bore a single point S374R mutation in the sodium (Na + ) efflux transporter PfATP4. There are many compounds known to inhibit PfATP4 and some are under preclinical development. MB14 was shown to inhibit Na + dependent ATPase activity in parasite membranes, consistent with the compound targeting PfATP4 directly. PfATP4 inhibitors cause swelling and lysis of infected erythrocytes, attributed to the accumulation of Na + inside the intracellular parasites and the resultant parasite swelling. We show here that inhibitor-induced lysis of infected erythrocytes is dependent upon the parasite protein RhopH2, a component of the new permeability pathways that are induced by the parasite in the erythrocyte membrane. These pathways mediate the influx of Na + into the infected erythrocyte and their suppression via RhopH2 knockdown limits the accumulation of Na + within the parasite hence protecting the infected erythrocyte from lysis. This study reveals a role for the parasite-induced new permeability pathways in the mechanism of action of PfATP4 inhibitors.
Cell Surface-expressed Cation-independent Mannose 6-Phosphate Receptor (CD222) Binds Enzymatically Active Heparanase Independently of Mannose 6-Phosphate to Promote Extracellular Matrix Degradation
Publisher: Elsevier BV
Date: 02-2008
Phosphoinositides: Multipurpose cellular lipids with emerging roles in cell death
Publisher: Springer Science and Business Media LLC
Date: 11-02-2019
Fcγ receptors: Gene structure and receptor function
Publisher: Springer Science and Business Media LLC
Date: 12-1992
DOI: 10.1007/BF02919128
Cloning and characterization of a novel NK cell-specific serine protease gene and its functional 5′-flanking sequences
Publisher: Springer Science and Business Media LLC
Date: 06-1995
DOI: 10.1007/BF00178584
Crystallization and preliminary X-ray crystallographic analysis of the plant defensin NaD1
Publisher: International Union of Crystallography (IUCr)
Date: 24-12-2011
Expression of the heparan sulfate-degrading enzyme heparanase is induced in infiltrating CD4+ T cells in experimental autoimmune encephalomyelitis and regulated at the level of transcription by early
Publisher: Oxford University Press (OUP)
Date: 26-07-2007
DOI: 10.1189/JLB.0507315
Abstract: The heparan sulfate-cleaving enzyme heparanase (HPSE) plays an important role in remodeling of the basement membrane and extracellular matrix during inflammation. Inducible HPSE enzymatic activity has been reported in leukocytes however, little is known of the molecular mechanisms that regulate HPSE gene expression during inflammatory disease. In this study, HPSE expression and regulation in the T cell-mediated disease model, experimental autoimmune encephalomyelitis (EAE), were investigated. Expression analysis showed that HPSE mRNA is induced in rat CD4+ antigen-specific T lymphocytes upon activation and correlates with the encephalitogenicity of the cells. Examination of the kinetics and cell type-specific expression of HPSE throughout the progression of active EAE in rats, indicated that HPSE was highly expressed in CD4+ T cells infiltrating the central nervous system (CNS) during clinical disease. Little or no HPSE expression was observed in CD8+ T cells, macrophages, or astrocytes during disease progression. To investigate the mechanism of inducible HPSE gene regulation in T cells, studies were extended into human primary T cells. HPSE mRNA, protein, and enzymatic activity were induced upon activation. Functional analysis of the human HPSE promoter identified an EGR1 binding motif that contained high inducible activity and was transactivated by EGR1. Furthermore, the treatment of primary T lymphocytes with an EGR1 siRNA inhibited inducible HPSE mRNA expression. These data provide evidence to suggest that inducible HPSE expression in primary T lymphocytes is regulated at the transcriptional level by EGR1 and is important in facilitating CD4+ T cell infiltration into the CNS to promote EAE.
Neurotoxic Amyloidogenic Peptides in the Proteome of SARS-COV2: Potential Implications for Neurological Symptoms in COVID-19
Publisher: Research Square Platform LLC
Date: 07-01-2022
DOI: 10.21203/RS.3.RS-1201481/V1
Abstract: COVID-19 is primarily known as a respiratory disease caused by the virus SARS-CoV-2. However, neurological symptoms such as memory loss, sensory confusion, cognitive and psychiatric issues, severe headaches, and even stroke are reported in as many as 30% of cases and can persist even after the infection is over (so-called ‘long COVID’). These neurological symptoms are thought to be caused by brain inflammation, caused by the virus infecting the central nervous system of COVID-19 patients, however we still don’t understand the molecular mechanisms that trigger these symptoms. The neurological effects of COVID-19 share many similarities to neurodegenerative diseases such as Alzheimer’s and Parkinson’s in which the presence of cytotoxic protein-based amyloid aggregates is a common etiological feature. Following the hypothesis that some neurological symptoms of COVID-19 may also follow an amyloid etiology we performed a bioinformatic scan of the SARS-CoV-2 proteome, detecting peptide fragments that were predicted to be highly amyloidogenic. We selected two of these peptides from the open reading frame 6 (ORF6) and open reading frame 10 (ORF10) proteins. The amyloidogenic virus-derived proteins studied in this work did not include spike (S) protein or any other proteins that have been modified to function as antigens in any current vaccines. We discovered that these ORF protein fragments rapidly self-assemble into amyloid aggregates. Furthermore, these amyloid assemblies were shown to be highly toxic to a neuronal cell line. We introduce and support the idea that cytotoxic amyloid aggregates of SARS-CoV-2 proteins are causing some of the neurological symptoms commonly found in COVID-19 and contributing to long COVID.
The Heparanase Regulatory Network in Health and Disease
Publisher: MDPI AG
Date: 14-10-2021
Abstract: The extracellular matrix (ECM) is a structural framework that has many important physiological functions which include maintaining tissue structure and integrity, serving as a barrier to invading pathogens, and acting as a reservoir for bioactive molecules. This cellular scaffold is made up of various types of macromolecules including heparan sulfate proteoglycans (HSPGs). HSPGs comprise a protein core linked to the complex glycosaminoglycan heparan sulfate (HS), the remodeling of which is important for many physiological processes such as wound healing as well as pathological processes including cancer metastasis. Turnover of HS is tightly regulated by a single enzyme capable of cleaving HS side chains: heparanase. Heparanase upregulation has been identified in many inflammatory diseases including atherosclerosis, fibrosis, and cancer, where it has been shown to play multiple roles in processes such as epithelial-mesenchymal transition, angiogenesis, and cancer metastasis. Heparanase expression and activity are tightly regulated. Understanding the regulation of heparanase and its downstream targets is attractive for the development of treatments for these diseases. This review provides a comprehensive overview of the regulators of heparanase as well as the enzyme’s downstream gene and protein targets, and implications for the development of new therapeutic strategies.
NK cell heparanase controls tumor invasion and immune surveillance
Publisher: American Society for Clinical Investigation
Date: 05-06-2017
DOI: 10.1172/JCI92958
The MS4A family: counting past 1, 2 and 3
Publisher: Wiley
Date: 19-05-2015
DOI: 10.1038/ICB.2015.48
Abstract: The MS4A (membrane-spanning 4-domain family, subfamily A) family of proteins contains some well-known members including MS4A1 (CD20), MS4A2 (FcɛRIβ) and MS4A3 (HTm4). These three MS4A family members are expressed on the cell surface of specific leukocyte subsets and have been well characterized as having key roles in regulating cell activation, growth and development. However, beyond MS4A1-3 there are a large number of related molecules (18 to date in humans) where our understanding of their biological roles is at a relatively nascent stage. This review examines the larger MS4A family focusing on their structure, expression, regulation and characterized and/or emerging biological roles. Our own work on one family member MS4A8B, and its possible role in epithelial cell regulation, is also highlighted.
The relationship between CCR6 and its binding partners: Does the CCR6-CCL20 axis have to be extended?
Publisher: Elsevier BV
Date: 03-2015
DOI: 10.1016/J.CYTO.2014.11.029
Abstract: Chemokines and their receptors are vital for the trafficking of immune cells. In an orchestrated fashion, up- and down-regulation of chemokines and their receptors contribute to both immune system homeostasis as well as inflammation. The CC chemokine, CCL20 and its cognate receptor, CCR6, are described as one of the few chemokine-receptor pairs that show exclusivity. In our review, we analyze observations which indicate that CCR6 does not have CCL20 as an exclusive ligand as once appreciated. For ex le, attempts to study the pair, utilizing mainly CCR6-deficient mice, are confounded by a family of non-chemokine ligands known as β-defensins that can bind to CCR6 and potentially can activate the cell. Therefore, a review of the activities of other potential binding partners of CCR6 is essential for interpretation of the current literature on this matter and for an understanding of their involvement in basic immunology and pathology.
Access to the Parent Tetrakis(pyridine)gold(III) Trication, Facile Formation of Rare Au(III) Terminal Hydroxides, and Preliminary Studies of Biological Properties
Publisher: American Chemical Society (ACS)
Date: 03-2016
DOI: 10.1021/ACS.INORGCHEM.5B02667
Abstract: In this paper we report on the use of [NO][BF4] to access tricationic tetrakis(pyridine)gold(III) from Au powder, a species inaccessible using the more traditional (tetrahydrothiophene)AuCl route. It is then demonstrated that this family of compounds can be used to access new terminal Au(III) hydroxides, a challenging class of compounds, and the first crystallographically characterized ex les employing bidentate ligands. Finally, preliminary biological studies indicate good activity for derivatives featuring polydentate ligands against the HeLa and PC3 cell lines but also strong inhibition of primary HUVEC cells.
The endoglycosidase heparanase enters the nucleus of T lymphocytes and modulates H3 methylation at actively transcribed genes via the interplay with key chromatin modifying enzymes.
Publisher: Informa UK Limited
Date: 05-2012
DOI: 10.4161/TRNS.19998
Neurotoxic amyloidogenic peptides in the proteome of SARS-COV2: potential implications for neurological symptoms in COVID-19
Publisher: Springer Science and Business Media LLC
Date: 13-06-2022
DOI: 10.1038/S41467-022-30932-1
Abstract: COVID-19 is primarily known as a respiratory disease caused by SARS-CoV-2. However, neurological symptoms such as memory loss, sensory confusion, severe headaches, and even stroke are reported in up to 30% of cases and can persist even after the infection is over (long COVID). These neurological symptoms are thought to be produced by the virus infecting the central nervous system, however we don’t understand the molecular mechanisms triggering them. The neurological effects of COVID-19 share similarities to neurodegenerative diseases in which the presence of cytotoxic aggregated amyloid protein or peptides is a common feature. Following the hypothesis that some neurological symptoms of COVID-19 may also follow an amyloid etiology we identified two peptides from the SARS-CoV-2 proteome that self-assemble into amyloid assemblies. Furthermore, these amyloids were shown to be highly toxic to neuronal cells. We suggest that cytotoxic aggregates of SARS-CoV-2 proteins may trigger neurological symptoms in COVID-19.
Immunotherapy of cytotoxic T cell-resistant tumors by T helper 2 cells: An eotaxin and STAT6-dependent process
Publisher: Rockefeller University Press
Date: 27-01-2003
DOI: 10.1084/JEM.20021683
Abstract: Currently most attempts at cancer immunotherapy involve the generation of CD8+ cytotoxic T lymphocytes (CTLs) against tumor-associated antigens. Many tumors, however, have been immunoselected to evade recognition by CTLs and thus alternative approaches to cancer immunotherapy are urgently needed. Here we demonstrate that CD4+ T cells that recognize a secreted tumor-specific antigen and exhibit a cytokine secretion profile characteristic of Th2 cells, are capable of clearing established lung and visceral metastases of a CTL-resistant melanoma. Clearance of lung metastases by the Th2 cells was found to be totally dependent on the eosinophil chemokine, eotaxin, and partially dependent on the transcription activator signal transducer and activator of transcription 6 (STAT6), with degranulating eosinophils within the tumors inducing tumor regression. In contrast, tumor-specific CD4+ Th1 cells, that recruited macrophages into the tumors, had no effect on tumor growth. This work provides the basis for a new approach to adoptive T cell immunotherapy of cancer.
Luminescent iridium(iii ) complexes of N-heterocyclic carbene ligands prepared using the ‘click reaction’
Publisher: Royal Society of Chemistry (RSC)
Date: 2019
DOI: 10.1039/C9DT01362H
Abstract: Luminescent and electrochemiluminescent N-heterocyclic carbene-combined 1,2,3-triazole and 1,2,3-triazolylidene Ir( iii ) complexes have been prepared and their potential as luminescent probes in cell imaging has been evaluated.
Human HPA endoglycosidase heparanase. Map position 4q21.3
Publisher: Springer Science and Business Media LLC
Date: 1999
Histidine-rich glycoprotein is a novel plasma pattern recognition molecule that recruits IgG to facilitate necrotic cell clearance via FcγRI on phagocytes
Publisher: American Society of Hematology
Date: 25-03-2010
DOI: 10.1182/BLOOD-2009-07-234013
Abstract: Under normal physiologic conditions, necrotic cells resulting from tissue injury are rapidly removed from the circulation and tissues by phagocytes, thus preventing the exposure of intracellular antigenic and immunostimulatory molecules that can aid the development of autoimmune disease. Histidine-rich glycoprotein (HRG), a relatively abundant plasma glycoprotein, has a multidomain structure that can interact with many ligands including components of the fibrinolytic and immune systems. Recently, it has been reported that HRG can bind strongly to cytoplasmic ligand(s) exposed in necrotic cells to enhance clearance by phagocytes. Here we describe the molecular mechanisms underpinning this process. A complex consisting of both HRG and immunoglobulin G (IgG) was found as necessary to aid necrotic cell uptake by monocytes, predominantly via an FcγRI-dependent mechanism. The findings in this study also show that HRG can potentially interact with anionic phospholipids exposed in necrotic cells. Furthermore, the enhanced phagocytosis of necrotic cells induced by HRG-IgG complexes triggers phagocytes to release proinflammatory cytokines such as interleukin-8 and tumor necrosis factor. Thus, HRG has the unique property of complexing with IgG and facilitating a proinflammatory innate immune response to promote the clearance of necrotic cells.
Isolation of cell type-specific apoptotic bodies by fluorescence-activated cell sorting
Publisher: Springer Science and Business Media LLC
Date: 06-01-2017
DOI: 10.1038/SREP39846
Abstract: Apoptotic bodies (ApoBDs) are membrane-bound extracellular vesicles that can mediate intercellular communication in physiological and pathological settings. By combining recently developed analytical strategies with fluorescence-activated cell sorting (FACS), we have developed a method that enables the isolation of ApoBDs from cultured cells to 99% purity. In addition, this approach also enables the identification and isolation of cell type-specific ApoBDs from tissue, bodily fluid and blood-derived s les.
Towards Novel Herbicide Modes of Action by Inhibiting Lysine Biosynthesis in Plants
Publisher: Cold Spring Harbor Laboratory
Date: 06-05-2021
DOI: 10.1101/2021.05.06.442928
Abstract: Weeds are becoming increasingly resistant to our current herbicides, posing a significant threat to agricultural production. Therefore, new herbicides are urgently needed. In this study, we exploited a novel herbicide target, dihydrodipicolinate synthase (DHDPS), which catalyses the first and rate-limiting step in lysine biosynthesis. Using a high throughput chemical screen, we identified the first class of plant DHDPS inhibitors that have micromolar potency against Arabidopsis thaliana DHDPS isoforms. Employing X-ray crystallography, we determined that this class of inhibitors binds to a novel and unexplored pocket within DHDPS, which is highly conserved across plant species. We also demonstrated that the inhibitors attenuated the germination and growth of A. thaliana seedlings and confirmed their pre-emergence herbicidal activity in soil-grown plants. These results provide proof-of-concept that lysine biosynthesis represents a promising target for the development of herbicides with a novel mode of action to tackle the global rise of herbicide resistant weeds.
Human beta-defensin 2 kills Candida albicans through phosphatidylinositol 4,5-bisphosphate-mediated membrane permeabilization
Publisher: American Association for the Advancement of Science (AAAS)
Date: 06-07-2018
Abstract: Human β-defensin 2 permeabilizes and kills fungal cells by specifically targeting the important plasma membrane lipid PIP 2 .
Functional Regulation of the Plasma Protein Histidine-Rich Glycoprotein by Zn2+ in Settings of Tissue Injury.
Publisher: MDPI AG
Date: 02-03-2017
DOI: 10.3390/BIOM7010022
Histidine-rich glycoprotein binds to cell-surface heparan sulfate via its N-terminal domain following Zn2+ chelation
Publisher: Elsevier BV
Date: 07-2004
Heparanase: A key enzyme involved in cell invasion
Publisher: Elsevier BV
Date: 03-2001
Redirected cytotoxic effector function. Requirements for expression of chimeric single chain high affinity immunoglobulin E receptors
Publisher: Elsevier BV
Date: 08-1996
Cloning and characterization of a mouse homologue of the human haematopoietic cell‐specific four‐transmembrane gene HTm4
Publisher: Wiley
Date: 08-2001
DOI: 10.1046/J.1440-1711.2001.01017.X
Abstract: Haematopoietic cell-specific transmembrane-4 (HTm4) is a four-transmembrane protein most closely related to CD20 and the beta subunit of the high affinity receptor for IgE (Fc(epsilon)RIbeta). To date, it has only been described in humans, where it is expressed in haematopoietic cells of both myeloid and lymphoid lineages. The function of HTm4 is unknown however, as for CD20 and Fc(epsilon)RI-beta, it is likely to play a role in signal transduction as part of a multi-subunit cell surface receptor complex. In this study, we report the cDNA cloning and expression distribution of mouse HTm4. The deduced mouse HTm4 protein is of 213 amino acids, and contains four putative transmembrane domains. Mouse HTm4 shows 62% overall amino acid identity with human HTm4 the transmembrane regions are highly conserved between both species (75% identity), whereas the N- and C-terminal and inter-transmembrane loop regions are more ergent (52%). Interestingly, the N-terminal domain of mouse HTm4 is predicted to be 23 amino acids shorter, and the C-terminal domain 23 amino acids longer, than that of human HTm4. Northern blot and reverse transcriptase (RT)-PCR analysis suggest that mouse HTm4 mRNA is expressed at low levels only in spleen, bone marrow and peripheral blood leucocytes. This is the first report of the cloning of HTm4 from a species other than human, and provides important sequence information towards the understanding of the function of this poorly characterized four-transmembrane molecule.
Identification of active-site residues of the pro-metastatic endoglycosidase heparanase
Publisher: American Chemical Society (ACS)
Date: 29-11-2000
DOI: 10.1021/BI002080P
Abstract: Heparanase is a beta-D-endoglucuronidase that cleaves heparan sulfate (HS) and has been implicated in many important physiological and pathological processes, including tumor cell metastasis, angiogenesis, and leukocyte migration. We report herein the identification of active-site residues of human heparanase. Using PSI-BLAST and PHI-BLAST searches of sequence databases, similarities were identified between heparanase and members of several of the glycosyl hydrolase families (10, 39, and 51) from glycosyl hydrolase clan A (GH-A), including strong local identities to regions containing the critical active-site catalytic proton donor and nucleophile residues that are conserved in this clan of enzymes. Furthermore, secondary structure predictions suggested that heparanase is likely to contain an (alpha/beta)(8) TIM-barrel fold, which is common to the GH-A families. On the basis of sequence alignments with a number of glycosyl hydrolases from GH-A, Glu(225) and Glu(343) of human heparanase were identified as the likely proton donor and nucleophile residues, respectively. The substitution of these residues with alanine and the subsequent expression of the mutant heparanases in COS-7 cells demonstrated that the HS-degrading capacity of both was abolished. In contrast, the alanine substitution of two other glutamic acid residues (Glu(378) and Glu(396)), both predicted to be outside the active site, did not affect heparanase activity. These data suggest that heparanase is a member of the clan A glycosyl hydrolases and has a common catalytic mechanism that involves two conserved acidic residues, a putative proton donor at Glu(225) and a nucleophile at Glu(343).
Structural and Functional Insight into Canarypox Virus CNP058 Mediated Regulation of Apoptosis
Publisher: MDPI AG
Date: 20-10-2017
DOI: 10.3390/V9100305
Histidine-rich glycoprotein specifically binds to necrotic cells via its amino-terminal domain and facilitates necrotic cell phagocytosis
Publisher: Elsevier BV
Date: 10-2005
Neurotoxic Amyloidogenic Peptides Identified in the Proteome of SARS-COV2: Potential Implications for Neurological Symptoms in COVID-19
Publisher: Cold Spring Harbor Laboratory
Date: 24-11-2021
DOI: 10.1101/2021.11.24.469537
Abstract: COVID-19 is primarily known as a respiratory disease caused by the virus SARS-CoV-2. However, neurological symptoms such as memory loss, sensory confusion, cognitive and psychiatric issues, severe headaches, and even stroke are reported in as many as 30% of cases and can persist even after the infection is over (so-called ‘long COVID’). These neurological symptoms are thought to be caused by brain inflammation, triggered by the virus infecting the central nervous system of COVID-19 patients, however we still don’t fully understand the mechanisms for these symptoms. The neurological effects of COVID-19 share many similarities to neurodegenerative diseases such as Alzheimer’s and Parkinson’s in which the presence of cytotoxic protein-based amyloid aggregates is a common etiological feature. Following the hypothesis that some neurological symptoms of COVID-19 may also follow an amyloid etiology we performed a bioinformatic scan of the SARS-CoV-2 proteome, detecting peptide fragments that were predicted to be highly amyloidogenic. We selected two of these peptides and discovered that they do rapidly self-assemble into amyloid. Furthermore, these amyloid assemblies were shown to be highly toxic to a neuronal cell line. We introduce and support the idea that cytotoxic amyloid aggregates of SARS-CoV-2 proteins are causing some of the neurological symptoms commonly found in COVID-19 and contributing to long COVID, especially those symptoms which are novel to long COVID in contrast to other post-viral syndromes.
Investigating the Role of Heparanase in Breast Cancer Development Utilising the MMTV-PyMT Murine Model of Mammary Carcinoma
Publisher: MDPI AG
Date: 05-06-2023
Abstract: Breast cancer is the second most common human malignancy and is a major global health burden. Heparanase (HPSE) has been widely implicated in enhancing the development and progression of solid tumours, including breast cancer. In this study, the well-established spontaneous mammary tumour-developing MMTV-PyMT murine model was utilised to examine the role of HPSE in breast cancer establishment, progression, and metastasis. The use of HPSE-deficient MMTV-PyMT (MMTV-PyMTxHPSE−/−) mice addressed the lack of genetic ablation models to investigate the role of HPSE in mammary tumours. It was demonstrated that even though HPSE regulated mammary tumour angiogenesis, mammary tumour progression and metastasis were HPSE-independent. Furthermore, there was no evidence of compensatory action by matrix metalloproteinases (MMPs) in response to the lack of HPSE expression in the mammary tumours. These findings suggest that HPSE may not play a significant role in the mammary tumour development of MMTV-PyMT animals. Collectively, these observations may have implications in the clinical setting of breast cancer and therapy using HPSE inhibitors.
Crystal structure of the human leukocyte Fc receptor, FcγRIIa
Publisher: Springer Science and Business Media LLC
Date: 05-1999
DOI: 10.1038/8241
Abstract: Fc gamma receptors bind IgG to initiate cellular responses against pathogens and soluble antigens. We have determined the three-dimensional structure of the extracellular portion of human Fc gammaRIIa to 2.0 A resolution providing a structural basis for the unique functions of the leukocyte FcR family. The receptor is composed of two immunoglobulin domains and arranged to expose the ligand-binding site at one end of domain 2. Using alanine mutants we find that the binding sites for IgG1 and 2 are similar but the relative importance of specific regions on the receptor varies. In crystals, Fc gammaRIIa molecules associate to resemble V(L)V(H) dimers, suggesting that two Fc gammaRIIa molecules could cooperate to bind IgG in an asymmetric manner.
The second and third extracellular domains of FcγRI (CD64) confer the unique high affinity binding of IgG2a
Publisher: Elsevier BV
Date: 10-1998
DOI: 10.1016/S0161-5890(98)00069-8
Abstract: FcgammaRI (CD64) is functionally unique as it is the only FcgammaR able to bind monomeric IgG with high affinity. FcgammaRI is also structurally distinct, containing an extracellular Ig-interactive region of three Ig-like domains in contrast to the two domains of the low affinity receptors FcgammaRII and FcgammaRIII. Previous studies have demonstrated that the third domain of FcgammaRI plays a crucial role in high affinity IgG binding of the receptor, with the first and second domains together forming a low affinity IgG binding motif. In this study the in idual functional contributions of the first and second domains of FcgammaRI to IgG binding have been investigated. Chimeric FcgammaR were generated by exchanging extracellular domains between mouse FcgammaRI and the structurally related yet distinct low affinity receptor for IgG, mouse FcgammaRII. The replacement of both domains 1 and 2 of FcgammaRI with domains 1 and 2 of FcgammaRII results in a dramatic change in IgG binding characteristics, as this receptor loses the capacity to bind monomeric IgG with high affinity and also demonstrates a broader specificity (binding not only IgG2a but also IgG1 and 2b. IgG3 was not tested). However, the substitution of FcgammaRII domain 2 of this chimeric receptor with domain 2 of FcgammaRI (generating a chimeric receptor with domain 1 of FcgammaRII linked to domains 2 and 3 of FcgammaRI) was found to reconstitute the specific high affinity monomeric IgG2a binding of wild-type FcgammaRI, albeit with a slightly reduced affinity (1.8-fold lower than wild-type FcgammaRI). These findings suggest that it is the specific interaction between domains 2 and 3 of FcgammaRI, with domain 1 playing a supporting role in maintaining the conformational stability of the receptor, that is the major structural requirement to confer the unique Ig binding characteristics of FcgammaRI.
Molecular mechanisms of late apoptotic/necrotic cell clearance
Publisher: Springer Science and Business Media LLC
Date: 18-12-2010
DOI: 10.1038/CDD.2009.195
Abstract: Phagocytosis serves as one of the key processes involved in development, maintenance of tissue homeostasis, as well as in eliminating pathogens from an organism. Under normal physiological conditions, dying cells (e.g., apoptotic and necrotic cells) and pathogens (e.g., bacteria and fungi) are rapidly detected and removed by professional phagocytes such as macrophages and dendritic cells (DCs). In most cases, specific receptors and opsonins are used by phagocytes to recognize and bind their target cells, which can trigger the intracellular signalling events required for phagocytosis. Depending on the type of target cell, phagocytes may also release both immunomodulatory molecules and growth factors to orchestrate a subsequent immune response and wound healing process. In recent years, evidence is growing that opsonins and receptors involved in the removal of pathogens can also aid the disposal of dying cells at all stages of cell death, in particular plasma membrane-damaged cells such as late apoptotic and necrotic cells. This review provides an overview of the molecular mechanisms and the immunological outcomes of late apoptotic/necrotic cell removal and highlights the striking similarities between late apoptotic/necrotic cell and pathogen clearance.
Cell surface expression of a peptide encoded by the unrearranged TCR-Vβ8.2 gene
Publisher: Elsevier BV
Date: 03-2006
DOI: 10.1016/J.MOLIMM.2005.07.040
Abstract: Germline transcription of T-cell receptor (TCR) genes has been described in early lymphoid cells. The most common explanation for this phenomenon is that transcription of unrearranged Vbeta genes directs gene usage during the rearrangement event. Germline transcription of the TCR-Vbeta8.2 gene has been detected in a precursor T-cell line, C1-V13D, which shows no rearrangement at any of the TCR gene loci. This cell line also shows weak binding of specific anti-Vbeta8.2 antibody to the cell surface, consistent with expression of a truncated TCRbeta chain. RT-PCR has been used to confirm expression of spliced germline transcripts of TCR-Vbeta8.2 in C1-V13D initiated from both leader (L)5.1 and L8.2. Transcripts initiated from L8.2 were also detectable in unspliced form. In order to test expression and subcellular localisation of any encoded peptides, lified germline transcripts in both spliced and unspliced form were cloned into the pEGFP-N1 fusion vector for stable transfection and overexpression in C1-V13D. Cell surface expression of a fusion protein between EGFP and a Vbeta peptide has been confirmed in C1-V13D but not in control COS-7 cells. Results presented here raise the possibility of a new pre-TCR structure specific to early lymphoid cells and based on TCR-Vbeta8.2 gene expression.
Combating Human Pathogens and Cancer by Targeting Phosphoinositides and Their Metabolism
Publisher: Elsevier BV
Date: 11-2019
DOI: 10.1016/J.TIPS.2019.09.006
Abstract: Pathogens and tumor cells have adopted various adept strategies to evade immunosurveillance and promote their growth and survival. There has been substantial evidence demonstrating phosphoinositide lipids and their modifying enzymes as essential host targets that are often hijacked by pathogens and tumor cells. The common dependence of pathogen virulence and tumor progression on phosphoinositides presents an exciting disease-combating potential, particularly combinatorial therapeutics. While traditional approaches to pharmacologically inhibit phosphoinositide-metabolizing enzymes has shown some promise, the direct targeting of phosphoinositides has recently emerged as a novel therapeutic strategy. Our review provides a current picture of the role of phosphoinositides during pathogen virulence and tumorigenesis as well as a thorough discussion on promises, challenges, and new perspectives of phosphoinositide-targeting drug development.
TREML4 receptor regulates inflammation and innate immune cell death during polymicrobial sepsis
Publisher: Springer Science and Business Media LLC
Date: 05-10-2020
DOI: 10.1038/S41590-020-0789-Z
Abstract: Sepsis is a biphasic disease characterized by an acute inflammatory response, followed by a prolonged immunosuppressive phase. Therapies aimed at controlling inflammation help to reduce the time patients with sepsis spend in intensive care units, but they do not lead to a reduction in overall mortality. Recently, the focus has been on addressing the immunosuppressive phase, often caused by apoptosis of immune cells. However, molecular triggers of these events are not yet known. Using whole-genome CRISPR screening in mice, we identified a triggering receptor expressed on myeloid cells (TREM) family receptor, TREML4, as a key regulator of inflammation and immune cell death in sepsis. Genetic ablation of Treml4 in mice demonstrated that TREML4 regulates calcium homeostasis, the inflammatory cytokine response, myeloperoxidase activation, the endoplasmic reticulum stress response and apoptotic cell death in innate immune cells, leading to an overall increase in survival rate, both during the acute and chronic phases of polymicrobial sepsis.
Defensin–lipid interactions in membrane targeting: mechanisms of action and opportunities for the development of antimicrobial and anticancer therapeutics
Publisher: Portland Press Ltd.
Date: 11-01-2022
DOI: 10.1042/BST20200884
Abstract: Defensins are a class of host defence peptides (HDPs) that often harbour antimicrobial and anticancer activities, making them attractive candidates as novel therapeutics. In comparison with current antimicrobial and cancer treatments, defensins uniquely target specific membrane lipids via mechanisms distinct from other HDPs. Therefore, defensins could be potentially developed as therapeutics with increased selectivity and reduced susceptibility to the resistance mechanisms of tumour cells and infectious pathogens. In this review, we highlight recent advances in defensin research with a particular focus on membrane lipid-targeting in cancer and infection settings. In doing so, we discuss strategies to harness lipid-binding defensins for anticancer and anti-infective therapies.
Online video and interactive TV experiences
Publisher: Association for Computing Machinery (ACM)
Date: 28-08-2015
DOI: 10.1145/2799629
Heparanase: A Novel Therapeutic Target for the Treatment of Atherosclerosis
Publisher: MDPI AG
Date: 12-10-2022
Abstract: Cardiovascular disease (CVD) is the leading cause of death and disability worldwide, and its management places a huge burden on healthcare systems through hospitalisation and treatment. Atherosclerosis is a chronic inflammatory disease of the arterial wall resulting in the formation of lipid-rich, fibrotic plaques under the subendothelium and is a key contributor to the development of CVD. As such, a detailed understanding of the mechanisms involved in the development of atherosclerosis is urgently required for more effective disease treatment and prevention strategies. Heparanase is the only mammalian enzyme known to cleave heparan sulfate of heparan sulfate proteoglycans, which is a key component of the extracellular matrix and basement membrane. By cleaving heparan sulfate, heparanase contributes to the regulation of numerous physiological and pathological processes such as wound healing, inflammation, tumour angiogenesis, and cell migration. Recent evidence suggests a multifactorial role for heparanase in atherosclerosis by promoting underlying inflammatory processes giving rise to plaque formation, as well as regulating lesion stability. This review provides an up-to-date overview of the role of heparanase in physiological and pathological processes with a focus on the emerging role of the enzyme in atherosclerosis.
Crocodile defensin (CpoBD13) antifungal activity via pH-dependent phospholipid targeting and membrane disruption
Publisher: Springer Science and Business Media LLC
Date: 03-2023
DOI: 10.1038/S41467-023-36280-Y
Abstract: Crocodilians are an order of ancient reptiles that thrive in pathogen-rich environments. The ability to inhabit these harsh environments is indicative of a resilient innate immune system. Defensins, a family of cysteine-rich cationic host defence peptides, are a major component of the innate immune systems of all plant and animal species, however crocodilian defensins are poorly characterised. We now show that the saltwater crocodile defensin CpoBD13 harbors potent antifungal activity that is mediated by a pH-dependent membrane-targeting action. CpoBD13 binds the phospholipid phosphatidic acid (PA) to form a large helical oligomeric complex, with specific histidine residues mediating PA binding. The utilisation of histidine residues for PA engagement allows CpoBD13 to exhibit differential activity at a range of environmental pH values, where CpoBD13 is optimally active in an acidic environment.
Tumor cell membrane-targeting cationic antimicrobial peptides: novel insights into mechanisms of action and therapeutic prospects
Publisher: Springer Science and Business Media LLC
Date: 02-08-2017
DOI: 10.1007/S00018-017-2604-Z
Abstract: There is an ongoing need for effective and targeted cancer treatments that can overcome the detrimental side effects presented by current treatment options. One class of novel anticancer molecules with therapeutic potential currently under investigation are cationic antimicrobial peptides (CAPs). CAPs are small innate immunity peptides found ubiquitously throughout nature that are typically membrane-active against a wide range of pathogenic microbes. A number of CAPs can also target mammalian cells and often display selective activity towards tumor cells, making them attractive candidates as novel anticancer agents warranting further investigation. This current and comprehensive review describes key ex les of naturally occurring membrane-targeting CAPs and their modified derivatives that have demonstrated anticancer activity, across multiple species of origin and structural subfamilies. In addition, we address recent advances made in the field and the ongoing challenges faced in translating experimental findings into clinically relevant treatments.
Soluble heparan sulfate fragments generated by heparanase trigger the release of pro-inflammatory cytokines through TLR-4.
Publisher: Public Library of Science (PLoS)
Date: 08-10-2014
Mice deficient in heparanase exhibit impaired dendritic cell migration and reduced airway inflammation.
Publisher: Wiley
Date: 14-02-2014
Abstract: Heparanase is a β-d-endoglucuronidase that cleaves heparan sulphate, a key component of the ECM and basement membrane. The remodelling of the ECM by heparanase has been proposed to regulate both normal physiological and pathological processes, including wound healing, inflammation, tumour angiogenesis and cell migration. Heparanase is also known to exhibit non-enzymatic functions by regulating cell adhesion, cell signalling and differentiation. In this study, constitutive heparanase-deficient (Hpse(-/-) ) mice were generated on a C57BL/6 background using the Cre/loxP recombination system, with a complete lack of heparanase mRNA, protein and activity. Although heparanase has been implicated in embryogenesis and development, Hpse(-/-) mice are anatomically normal and fertile. Interestingly, consistent with the suggested function of heparanase in cell migration, the trafficking of dendritic cells from the skin to the draining lymph nodes was markedly reduced in Hpse(-/-) mice. Furthermore, the ability of Hpse(-/-) mice to generate an allergic inflammatory response in the airways, a process that requires dendritic cell migration, was also impaired. These findings establish an important role for heparanase in immunity and identify the enzyme as a potential target for regulation of an immune response.
Author response: Towards novel herbicide modes of action by inhibiting lysine biosynthesis in plants
Publisher: eLife Sciences Publications, Ltd
Date: 20-07-2021
A novel mechanism of generating extracellular vesicles during apoptosis via a beads-on-a-string membrane structure
Publisher: Springer Science and Business Media LLC
Date: 15-06-2015
DOI: 10.1038/NCOMMS8439
Abstract: Disassembly of apoptotic cells into smaller fragments (a form of extracellular vesicle called apoptotic bodies) can facilitate removal of apoptotic debris and intercellular communication. However, the mechanism underpinning this process is unclear. While observing monocytes undergoing apoptosis by time-lapse microscopy, we discovered a new type of membrane protrusion that resembles a ‘beads-on-a-string’ structure. Strikingly, the ‘beads’ are frequently sheared off the ‘string’ to form apoptotic bodies. Generation of apoptotic bodies via this mechanism can facilitate a sorting process and results in the exclusion of nuclear contents from apoptotic bodies. Mechanistically, generation of ‘beads-on-a-string’ protrusion is controlled by the level of actomyosin contraction and apoptopodia formation. Furthermore, in an unbiased drug screen, we identified the ability of sertraline (an antidepressant) to block the formation of ‘beads-on-a-string’ protrusions and apoptotic bodies. These data uncover a new mechanism of apoptotic body formation in monocytes and also compounds that can modulate this process.
Salt-Tolerant antifungal and antibacterial activities of the corn defensin ZmD32
Publisher: Frontiers Media SA
Date: 12-04-2019
X-ray structure of a carpet-like antimicrobial defensin-phospholipid membrane disruption complex
Publisher: Springer Science and Business Media LLC
Date: 17-05-2018
DOI: 10.1038/S41467-018-04434-Y
Abstract: Defensins are cationic antimicrobial peptides expressed throughout the plant and animal kingdoms as a first line of defense against pathogens. Membrane targeting and disruption is a crucial function of many defensins, however the precise mechanism remains unclear. Certain plant defensins form dimers that specifically bind the membrane phospholipids phosphatidic acid (PA) and phosphatidylinositol 4,5-bisphosphate, thereby triggering the assembly of defensin–lipid oligomers that permeabilize cell membranes. To understand this permeabilization mechanism, here we determine the crystal structure of the plant defensin NaD1 bound to PA. The structure reveals a 20-mer that adopts a concave sheet- or carpet-like topology where NaD1 dimers form one face and PA acyl chains form the other face of the sheet. Furthermore, we show that Arg39 is critical for PA binding, oligomerization and fungal cell killing. These findings identify a putative defensin–phospholipid membrane attack configuration that supports a longstanding proposed carpet mode of membrane disruption.
The Defensins Consist of Two Independent, Convergent Protein Superfamilies
Publisher: Oxford University Press (OUP)
Date: 13-06-2016
Abstract: The defensin and defensin-like proteins are an extensive group of small, cationic, disulfide-rich proteins found in animals, plants, and fungi and mostly perform roles in host defense. The term defensin was originally used for small mammalian proteins found in neutrophils and was subsequently applied to insect proteins and plant γ-thionins based on their perceived sequence and structural similarity. Defensins are often described as ancient innate immunity molecules and classified as a single superfamily and both sequence alignments and phylogenies have been constructed. Here, we present evidence that the defensins have not all evolved from a single ancestor. Instead, they consist of two analogous superfamilies, and extensive convergent evolution is the source of their similarities. Evidence of common origin necessarily gets weaker for distantly related genes, as is the case for defensins, which are both ergent and small. We show that similarities that have been used as evidence for common origin are all expected by chance in short, constrained, disulfide-rich proteins. Differences in tertiary structure, secondary structure order, and disulfide bond connectivity indicate convergence as the likely source of the similarity. We refer to the two evolutionarily independent groups as the cis-defensins and trans-defensins based on the orientation of the most conserved pair of disulfides.
Monitoring the progression of cell death and the disassembly of dying cells by flow cytometry
Publisher: Springer Science and Business Media LLC
Date: 03-03-2016
Abstract: The use of annexin A5 (A5) and either propidium iodide or 7-aminoactinomycin D (PI/7-AAD) stains to measure cell death by flow cytometry has been considered the gold standard by most investigators. However, this widely used method often makes the assumption that there are only three types of particles in a s le: viable, apoptotic and necrotic cells. To study the progression of cell death in greater detail, in particular how apoptotic cells undergo fragmentation to generate membrane-bound vesicles known as apoptotic bodies, we established a flow cytometry-based protocol to accurately and rapidly measure the cell death process. This protocol uses a combination of A5 and TO-PRO-3 (a commercially available nucleic acid-binding dye that stains early apoptotic and necrotic cells differentially), and a logical seven-stage analytical approach to distinguish six types of particles in a s le, including apoptotic bodies and cells at three different stages of cell death. The protocol requires 1-5 h for s le preparation (including induction of cell death), 20 min for staining and 5 min for data analysis.
The lure of the lipids: how defensins exploit membrane phospholipids to induce cytolysis in target cells
Publisher: Springer Science and Business Media LLC
Date: 30-03-2017
Domain one of the high affinity IgE receptor, FcεRI, regulates binding to IgE through its interface with domain two
Publisher: Elsevier BV
Date: 03-2000
Early growth response gene 1 (EGR1) regulates heparanase gene transcription in tumor cells.
Publisher: Elsevier BV
Date: 10-2005
Regulation of Inducible Heparanase Gene Transcription in Activated T Cells by Early Growth Response 1
Publisher: Elsevier BV
Date: 12-2003
Regulation of mouse Heparanase gene expression in T lymphocytes and tumor cells
Publisher: Wiley
Date: 09-01-2007
Abstract: Heparanase (HPSE) is an endoglycosidase that cleaves heparan sulfate (HS) and plays an important role in tumor metastasis, angiogenesis and inflammation. The regulation of HPSE expression and function is tightly controlled and the increasing use of the mouse as an animal model to define the role of HPSE in many physiological and pathological settings, makes understanding the regulatory mechanisms of HPSE in this species of fundamental importance. However, the expression distribution of the mouse Hpse gene and the mechanisms that regulate its transcription are poorly defined. In this study, the mouse Hpse gene was determined to encode for two mRNA transcripts of 1.9 and 3.2 kb in length with identical open reading frames that showed similar tissue expression distribution to the human HPSE. The mouse Hpse promoter was cloned and a 478-bp minimal promoter was identified that contained regulatory elements responsible for both basal promoter activity in mouse tumor cells as well as inducible activity in T cells. Mutagenesis and transactivation studies identified a functional site in the minimal promoter region for the transcription factor Early growth response gene 1 (Egr1). Interestingly, Egr1 acted differentially in mouse tumor cells, functioning in an activating or repressive manner in breast carcinoma or melanoma cells, respectively. Furthermore, the proximal region of the promoter, identified as important in the regulation of Hpse transcription, was shown to become accessible in T cells upon cell activation. Significantly, the maximal accessibility of the promoter occurred at 16 h post-stimulation, which correlated with the induction kinetics of Hpse mRNA expression. In summary, this study demonstrates that mouse Hpse is expressed and regulated in a similar manner to human HPSE and also provides some novel insights into mechanisms of Hpse gene regulation that are likely to be relevant to control of the human gene.
Identification of the Immunoglobulin Binding Regions (IBR) of FcγRII and FcɛRI
Publisher: Wiley
Date: 02-1992
DOI: 10.1111/J.1600-065X.1992.TB00623.X
Abstract: In view of the similarity between the reported effects of hyperventilation and recurrent functional symptom presented in primary care, a study was undertaken to establish whether such symptoms are attributable to hyperventilation. Twenty patients with two or more recurrent functional symptoms which their doctors found difficult to diagnose or treat, and 30 controls, were studied using symptom questionnaires and a series of hyperventilation provocation tests during which the partial pressure of carbon dioxide (PCO2) and symptoms were recorded. Sixteen cases (80%) had unexplained breathlessness compared with two of the controls (7%). All of the cases recognized familiar functional symptoms during provoked hyperventilation, and in 16 (80%), these included primary physical symptoms only 23% of the controls recognized any previously experienced symptom. Considerable overlap of PCO2 values between groups meant that absolute values of PCO2 were not useful in differentiating between groups, but cases were more likely than controls to have a PCO2 of less than 4 kPa at rest, three minutes after hyperventilation, or during mental stress (75% of cases fulfilled one or more of these criteria versus 40% of controls). This is the first study in primary care to examine the effect of hyperventilation in a group of patients with multiple somatic symptoms. The findings have implications for the recognition and management of such patients.
Plexin B2 is a regulator of monocyte apoptotic cell disassembly
Publisher: Elsevier BV
Date: 11-2019
DOI: 10.1016/J.CELREP.2019.10.014
Abstract: Billions of cells undergo apoptosis daily and often fragment into small, membrane-bound extracellular vesicles termed apoptotic bodies (ApoBDs). We demonstrate that apoptotic monocytes undergo a highly coordinated disassembly process and form long, beaded protrusions (coined as beaded apoptopodia), which fragment to release ApoBDs. Here, we find that the protein plexin B2 (PlexB2), a transmembrane receptor that regulates axonal guidance in neurons, is enriched in the ApoBDs of THP1 monocytes and is a caspase 3/7 substrate. To determine whether PlexB2 is involved in the disassembly of apoptotic monocytes, we generate PlexB2-deficient THP1 monocytes and demonstrate that lack of PlexB2 impairs the formation of beaded apoptopodia and ApoBDs. Consequently, the loss of PlexB2 in apoptotic THP1 monocytes impairs their uptake by both professional and non-professional phagocytes. Altogether, these data identify PlexB2 as a positive regulator of apoptotic monocyte disassembly and demonstrate the importance of this process in apoptotic cell clearance.
Convergent evolution of defensin sequence, structure and function
Publisher: Springer Science and Business Media LLC
Date: 24-08-2017
DOI: 10.1007/S00018-016-2344-5
Abstract: Defensins are a well-characterised group of small, disulphide-rich, cationic peptides that are produced by essentially all eukaryotes and are highly erse in their sequences and structures. Most display broad range antimicrobial activity at low micromolar concentrations, whereas others have other erse roles, including cell signalling (e.g. immune cell recruitment, self/non-self-recognition), ion channel perturbation, toxic functions, and enzyme inhibition. The defensins consist of two superfamilies, each derived from an independent evolutionary origin, which have subsequently undergone extensive ergent evolution in their sequence, structure and function. Referred to as the cis- and trans-defensin superfamilies, they are classified based on their secondary structure orientation, cysteine motifs and disulphide bond connectivities, tertiary structure similarities and precursor gene sequence. The utility of displaying loops on a stable, compact, disulphide-rich core has been exploited by evolution on multiple occasions. The defensin superfamilies represent a case where the ensuing convergent evolution of sequence, structure and function has been particularly extreme. Here, we discuss the extent, causes and significance of these convergent features, drawing ex les from across the eukaryotes.
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