ORCID Profile
0000-0003-3803-871X
Current Organisations
Australian National University
,
University of Oxford
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Publisher: American Association for the Advancement of Science (AAAS)
Date: 22-07-2022
Abstract: Many pathogens exploit host cell-surface glycans. However, precise analyses of glycan ligands binding with heavily modified pathogen proteins can be confounded by overlapping sugar signals and/or compounded with known experimental constraints. Universal saturation transfer analysis (uSTA) builds on existing nuclear magnetic resonance spectroscopy to provide an automated workflow for quantitating protein-ligand interactions. uSTA reveals that early-pandemic, B-origin-lineage severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike trimer binds sialoside sugars in an “end-on” manner. uSTA-guided modeling and a high-resolution cryo–electron microscopy structure implicate the spike N-terminal domain (NTD) and confirm end-on binding. This finding rationalizes the effect of NTD mutations that abolish sugar binding in SARS-CoV-2 variants of concern. Together with genetic variance analyses in early pandemic patient cohorts, this binding implicates a sialylated polylactosamine motif found on tetraantennary N-linked glycoproteins deep in the human lung as potentially relevant to virulence and/or zoonosis.
Publisher: Elsevier BV
Date: 12-2022
Publisher: Elsevier BV
Date: 10-2022
DOI: 10.1111/JTH.15794
Abstract: Breast cancer results in a three- to four-fold increased risk of venous thromboembolism (VTE), which is associated with reduced patient survival. Despite this, the mechanisms underpinning breast cancer-associated thrombosis remain poorly defined. Tumor cells can trigger endothelial cell (EC) activation resulting in increased von Willebrand factor (VWF) secretion. Importantly, elevated plasma VWF levels constitute an independent biomarker for VTE risk. Moreover, in a model of melanoma, treatment with low molecular weight heparin (LMWH) negatively regulated VWF secretion and attenuated tumor metastasis. To investigate the role of VWF in breast cancer metastasis and examine the effect of LMWH in modulating EC activation and breast tumor transmigration. von Willebrand factor levels were measured by ELISA. Primary ECs were used to assess tumor-induced activation, angiogenesis, tumor adhesion, and transendothelial migration. Patients with metastatic breast cancer have markedly elevated plasma VWF:Ag levels that also correlate with poorer survival. MDA-MB-231 and MCF-7 breast cancer cells induce secretion of VWF, angiopoietin-2, and osteoprotegerin from ECs, which is further enhanced by the presence of platelets. Vascular endothelial growth factor-A (VEGF-A) plays an important role in modulating breast cancer-induced VWF release. Moreover, VEGF-A from breast tumor cells also contributes to a pro-angiogenic effect on ECs. VWF multimers secreted from ECs, in response to tumor-VEGF-A, mediate adhesion of breast tumor cells along the endothelium. LMWH inhibits VWF-breast tumor adhesion and transendothelial migration. Our findings highlight the significant crosstalk between tumor cells and the endothelium including increased VWF secretion which may contribute to tumor metastasis.
Publisher: Cold Spring Harbor Laboratory
Date: 14-04-2021
DOI: 10.1101/2021.04.14.439284
Abstract: Many host pathogen interactions such as human viruses (including non-SARS-coronaviruses) rely on attachment to host cell-surface glycans. There are conflicting reports about whether the Spike protein of SARS-CoV-2 binds to sialic acid commonly found on host cell-surface N-linked glycans. In the absence of a biochemical assay, the ability to analyze the binding of glycans to heavily- modified proteins and resolve this issue is limited. Classical Saturation Transfer Difference (STD) NMR can be confounded by overlapping sugar resonances that compound with known experimental constraints. Here we present ‘universal saturation transfer analysis’ (uSTA), an NMR method that builds on existing approaches to provide a general and automated workflow for studying protein-ligand interactions. uSTA reveals that B-origin-lineage-SARS-CoV-2 spike trimer binds sialoside sugars in an ‘end on’ manner and modelling guided by uSTA localises binding to the spike N-terminal domain (NTD). The sialylated-polylactosamine motif is found on tetraantennary human N-linked-glycoproteins in deeper lung and may have played a role in zoonosis. Provocatively, sialic acid binding is abolished by mutations in some subsequent SARS- CoV-2 variants-of-concern. A very high resolution cryo-EM structure confirms the NTD location and ‘end on’ mode it rationalises the effect of NTD mutations and the structure-activity relationship of sialic acid analogues. uSTA is demonstrated to be a robust, rapid and quantitative tool for analysis of binding, even in the most demanding systems. The surface proteins found on both pathogens and host cells mediate entry (and exit) and influence disease progression and transmission. Both types can bear host-generated post- translational modifications such as glycosylation that are essential for function but can confound biophysical methods used for dissecting key interactions. Several human viruses (including non- SARS-coronaviruses) attach to host cell-surface N -linked glycans that include forms of sialic acid (sialosides). There remains, however, conflicting evidence as to if or how SARS-associated coronaviruses might use such a mechanism. Here, we demonstrate quantitative extension of ‘saturation transfer’ protein NMR methods to a complete mathematical model of the magnetization transfer caused by interactions between protein and ligand. The method couples objective resonance-identification via a deconvolution algorithm with Bloch-McConnell analysis to enable a structural, kinetic and thermodynamic analysis of ligand binding beyond previously-perceived limits of exchange rates, concentration or system. Using an automated and openly available workflow this ‘universal saturation transfer’ analysis (uSTA) can be readily-applied in a range of even heavily-modified systems in a general manner to now obtain quantitative binding interaction parameters (K D , k Ex ). uSTA proved critical in mapping direct interactions between natural sialoside sugar ligands and relevant virus-surface attachment glycoproteins – SARS-CoV-2-spike and influenza-H1N1-haemagglutinin variants – by quantitating ligand signal in spectral regions otherwise occluded by resonances from mobile protein glycans (that also include sialosides). In B- origin-lineage-SARS-CoV-2 spike trimer ‘end on’-binding to sialoside sugars was revealed contrasting with ‘extended surface’-binding for heparin sugar ligands uSTA-derived constraints used in structural modelling suggested sialoside-glycan binding sites in a beta-sheet-rich region of spike N-terminal domain (NTD). Consistent with this, uSTA-glycan binding was minimally- perturbed by antibodies that neutralize the ACE2-binding domain (RBD) but strongly disrupted in spike from the B1.1.7/alpha and B1.351/beta variants-of-concern, which possess hotspot mutations in the NTD. Sialoside binding in B-origin-lineage-NTD was unequivocally pinpointed by cryo-EM to a site that is created from residues that are notably deleted in variants (e.g. H69,V70,Y145 in alpha). An analysis of beneficial genetic variances in cohorts of patients from early 2020 suggests a model in which this site in the NTD of B-origin-lineage-SARS-CoV-2 (but not in alpha/beta-variants) may have exploited a specific sialylated-polylactosamine motif found on tetraantennary human N -linked-glycoproteins in deeper lung. Together these confirm a novel binding mode mediated by the unusual NTD of SARS-CoV-2 and suggest how it may drive virulence and/or zoonosis via modulation of glycan attachment. Since cell-surface glycans are widely relevant to biology and pathology, uSTA can now provide ready, quantitative, widespread analysis of complex, host-derived and post-translationally modified proteins with putative ligands relevant to disease even in previously confounding complex systems.
Publisher: Elsevier BV
Date: 09-2022
DOI: 10.1016/J.TIM.2022.02.004
Abstract: Glycans are repeating carbohydrate structures added as post-translational modifications (PTMs) to proteins, forming glycoproteins. Self-glycans found on human cells, and viral glycoproteins produced in host cells, are generally weakly immunogenic, which is necessary to avoid autoimmunity. This feature is exploited by many pathogenic viruses, which glycosylate surface proteins to evade or reduce immune recognition. The HIV type-1 (HIV-1) envelope glycoprotein (Env) is heavily glycosylated, which broadly acts to shield neutralisation-relevant protein surfaces with immunorecessive self-glycans to hinder B cell recognition. However, a small subset of HIV-1-infected in iduals develops potent broadly neutralising antibodies (bnAbs), many of which directly engage the glycan shield. This provides hope that such antibodies could be elicited via vaccination and help to provide protective immunity. However, HIV-1 vaccine candidates have thus far failed to fully recapitulate such glycan-specific neutralising responses. In this review we consider the fundamental glycoimmunology and structural biology that underpin glycans in antibody evasion and as antibody targets and discuss potential approaches to harness glycan targeting for HIV-1 vaccine design.
Publisher: Cold Spring Harbor Laboratory
Date: 23-08-2023
DOI: 10.1101/2023.08.22.554267
Abstract: Efferocytic clearance of apoptotic cells in general, and T cells in particular, is required for tissue and immune homeostasis. Transmembrane mucins are extended glycoproteins highly expressed in the cell glycocalyx that act as a barrier to phagocytosis. Whether and how mucins may be regulated during cell death to facilitate efferocytic corpse clearance is not understood. Here we show that normal and transformed human T cells express a subset of mucins which are rapidly and selectively removed from the cell surface during apoptosis. This process is mediated by the ADAM10 sheddase, the activity of which is associated with XKR8-catalyzed flipping of phosphatidylserine to the outer leaflet of the plasma membrane. Mucin clearance enhances uptake of apoptotic T cells by macrophages, confirming mucins as an enzymatically-modulatable barrier to efferocytosis. Together these findings reveal a novel glycocalyx regulatory pathway with implications for therapeutic intervention in the clearance of normal and transformed apoptotic T cells.
Publisher: Elsevier BV
Date: 07-2022
Abstract: Immune tolerance to self-glycans is a host mechanism to avoid autoimmunity, which is exploited by HIV-1 coating its envelope glycoproteins with glycans to evade neutralising antibodies (nAbs). Huettner et al. describe cross-reactivity between Schistosoma mansoni glycans and HIV-1 envelope glycoprotein glycans, suggesting a strategy for induction of HIV-1 nAbs by vaccination.
Publisher: Springer Science and Business Media LLC
Date: 18-05-2021
DOI: 10.1038/S41598-021-89860-7
Abstract: IL-4 production is associated with low-avidity, poorly cytotoxic T cell induction that contributes to viral immune evasion and the failure of T cell-based vaccines. Yet, the precise mechanisms that regulate IL-4 signalling in T cells remain elusive. Mounting evidence indicates that cells can dynamically alter their IL-4/IL-13 receptor signature to modulate downstream immune outcomes upon pathogen encounter. Here, we describe how naïve (CD62L + CD44 lo–mid ) CD4 and CD8 T cells distinctly engage both STAT6 and STAT3 in response to IL-4. We further show that IL-4R⍺ expression is both time- and IL-4 concentration-dependent. Remarkably, our findings reveal that STAT3 inhibition can ablate IL-4R⍺ and affect transcriptional expression of other Stat and Jak family members. By extension, the loss of STAT3 lead to aberrant STAT6 phosphorylation, revealing an inter-regulatory relationship between the two transcription factors. Moreover, IL-4 stimulation down-regulated TGF-β1 and IFN-γR1 expression on naïve T cells, possibly signifying the broad regulatory implications of IL-4 in conditioning lineage commitment decisions during early infection. Surprisingly, naïve T cells were unresponsive to IL-13 stimulation, unlike dendritic cells. Collectively, these findings could be exploited to inform more efficacious vaccines, as well as design treatments against IL-4/IL-13-associated disease conditions.
Publisher: Springer Science and Business Media LLC
Date: 23-01-2020
DOI: 10.1038/S41598-020-57815-Z
Abstract: This study demonstrates that 24 h following viral vector-based vaccination IL-13Rα2 functions as a master sensor on conventional dendritic cells (cDCs), abetted by high protein stability coupled with minimal mRNA expression, to rapidly regulate DC mediated IL-13 responses at the lung mucosae, unlike IL-13Rα1. Under low IL-13, IL-13Rα2 performs as a primary signalling receptor, whilst under high IL-13, acts to sequester IL-13 to maintain homeostasis, both in a STAT3-dependent manner. Likewise, we show that viral vector-derived IL-13 levels at the vaccination site can induce differential STAT3/STAT6 paradigms in lung cDC, that can get regulated collaboratively or independently by TGF-β1 and IFN-γ. Specifically, low IL-13 responses associated with recombinant Fowlpox virus (rFPV) is regulated by early IL-13Rα2, correlated with STAT3/TGF-β1 expression. Whilst, high IL-13 responses, associated with recombinant Modified Vaccinia Ankara (rMVA) is regulated in an IL-13Rα1/STAT6 dependent manner associated with IFN-γR expression bias. Different viral vaccine vectors have previously been shown to induce unique adaptive immune outcomes. Taken together current observations suggest that IL-13Rα2-driven STAT3/STAT6 equilibrium at the cDC level may play an important role in governing the efficacy of vector-based vaccines. These new insights have high potential to be exploited to improve recombinant viral vector-based vaccine design, according to the pathogen of interest and/or therapies against IL-13 associated disease conditions.
Publisher: Oxford University Press (OUP)
Date: 02-2019
DOI: 10.1093/GBE/EVZ026
Location: United Kingdom of Great Britain and Northern Ireland
No related grants have been discovered for Lachlan Deimel.