Imperial College London

ProfessorAnneDell

Faculty of Natural SciencesDepartment of Life Sciences

Professor of Carbohydrate Bichemistry
 
 
 
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a.dell

 
 
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101BSir Ernst Chain BuildingSouth Kensington Campus

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Summary

 

Publications

Publication Type
Year
to

330 results found

Kikuchi C, Antonopoulos A, Wang S, Maemura T, Karamanska R, Lee C, Thompson AJ, Dell A, Kawaoka Y, Haslam SM, Paulson JCet al., 2023, Glyco-engineered MDCK cells display preferred receptors of H3N2 influenza absent in eggs used for vaccines, Nature Communications, Vol: 14, ISSN: 2041-1723

Evolution of human H3N2 influenza viruses driven by immune selection has narrowed the receptor specificity of the hemagglutinin (HA) to a restricted subset of human-type (Neu5Acα2-6 Gal) glycan receptors that have extended poly-LacNAc (Galβ1-4GlcNAc) repeats. This altered specificity has presented challenges for hemagglutination assays, growth in laboratory hosts, and vaccine production in eggs. To assess the impact of extended glycan receptors on virus binding, infection, and growth, we have engineered N-glycan extended (NExt) cell lines by overexpressing β3-Ν-acetylglucosaminyltransferase 2 in MDCK, SIAT, and hCK cell lines. Of these, SIAT-NExt cells exhibit markedly increased binding of H3 HAs and susceptibility to infection by recent H3N2 virus strains, but without impacting final virus titers. Glycome analysis of these cell lines and allantoic and amniotic egg membranes provide insights into the importance of extended glycan receptors for growth of recent H3N2 viruses and relevance to their production for cell- and egg-based vaccines.

Journal article

Gimeno-Molina B, Bayar E, Mountain K, Love R, Lee Y, Muller I, Dell A, Haslam S, Grassi P, Wu G, MacIntyre D, Bennett P, Kropf P, Sykes Let al., 2023, The role of cervical neutrophils in cervicovaginal inflammation in women at high-risk of delivering preterm, 12th International Workshop Reunion Island Reproductive Immunology, Immunological tolerance and Immunology of preeclampsia, Publisher: ELSEVIER IRELAND LTD, Pages: 31-31, ISSN: 0165-0378

Conference paper

Mountain K, MacIntyre D, Chan D, Hyde A, Pasint-Magyar J, Lee Y, Brown R, David A, Dell A, Ten F, Haslam S, Liu Y, Lewis H, Norman J, Stock S, Teoh T, Terzidou V, Kundu S, Bennett P, Sykes Let al., 2023, ABO blood group antigens and preterm birth risk, 12th International Workshop Reunion Island Reproductive Immunology, Immunological tolerance and Immunology of preeclampsia, Publisher: ELSEVIER IRELAND LTD, Pages: 37-37, ISSN: 0165-0378

Conference paper

Abouelhadid S, Atkins ER, Kay EJ, Passmore IJ, North SJ, Lehri B, Hitchen P, Bakke E, Rahman M, Bossé JT, Li Y, Terra VS, Langford PR, Dell A, Wren BW, Cuccui Jet al., 2023, Development of a novel glycoengineering platform for the rapid production of conjugate vaccines, Microbial Cell Factories, Vol: 22, Pages: 1-13, ISSN: 1475-2859

Conjugate vaccines produced either by chemical or biologically conjugation have been demonstrated to be safe and efficacious in protection against several deadly bacterial diseases. However, conjugate vaccine assembly and production have several shortcomings which hinders their wider availability. Here, we developed a tool, Mobile-element Assisted Glycoconjugation by Insertion on Chromosome, MAGIC, a novel biotechnological platform that overcomes the limitations of the current conjugate vaccine design method(s). As a model, we focused our design on a leading bioconjugation method using N-oligosaccharyltransferase (OTase), PglB. The installation of MAGIC led to at least twofold increase in glycoconjugate yield via MAGIC when compared to conventional N-OTase based bioconjugation method(s). Then, we improved MAGIC to (a) allow rapid installation of glycoengineering component(s), (b) omit the usage of antibiotics, (c) reduce the dependence on protein induction agents. Furthermore, we show the modularity of the MAGIC platform in performing glycoengineering in bacterial species that are less genetically tractable than the commonly used Escherichia coli. The MAGIC system promises a rapid, robust and versatile method to develop vaccines against serious bacterial pathogens. We anticipate the utility of the MAGIC platform could enhance vaccines production due to its compatibility with virtually any bioconjugation method, thus expanding vaccine biopreparedness toolbox.

Journal article

Xie Y, Zhao F, Freitag N, Borowski S, Wang Y, Harms C, Pang P-C, Desforges J, Wen T, Schwedhelm E, Singh M, Dechend R, Dell A, Haslam SM, Dveksler G, Garcia MG, Blois SMet al., 2023, Maternal-derived galectin-1 shapes the placenta niche through Sda terminal glycosylation: implication for preeclampsia, PNAS Nexus, Vol: 2, ISSN: 2752-6542

Placental abnormalities cause impaired fetal growth and poor pregnancy outcome (e.g. preeclampsia [PE]) with long-lasting consequences for the mother and offspring. The molecular dialogue between the maternal niche and the developing placenta is critical for the function of this organ. Galectin-1 (gal-1), a highly expressed glycan-binding protein at the maternal–fetal interface, orchestrates the maternal adaptation to pregnancy and placenta development. Down-regulation or deficiency of gal-1 during pregnancy is associated with the development of PE; however, the maternal- and placental-derived gal-1 contributions to the disease onset are largely unknown. We demonstrate that lack of gal-1 imposes a risk for PE development in a niche-specific manner, and this is accompanied by a placental dysfunction highly influenced by the absence of maternal-derived gal-1. Notably, differential placental glycosylation through the Sda-capped N-glycans dominates the invasive trophoblast capacity triggered by maternal-derived gal-1. Our findings show that gal-1 derived from the maternal niche is essential for healthy placenta development and indicate that impairment of the gal-1 signaling pathway within the maternal niche could be a molecular cause for maternal cardiovascular maladaptation during pregnancy.

Journal article

Huang Z, Lai PF, Cocker ATH, Haslam SM, Dell A, Brady HJM, Johnson MRet al., 2023, Roles of N-linked glycosylation and glycan-binding proteins in placentation: trophoblast infiltration, immunomodulation, angiogenesis, and pathophysiology, Biochemical Society Transactions, Vol: 51, Pages: 639-653, ISSN: 0300-5127

Protein N-linked glycosylation is a structurally diverse post-translational modification that stores biological information in a larger order of magnitude than other post-translational modifications such as phosphorylation, ubiquitination and acetylation. This gives N-glycosylated proteins a diverse range of properties and allows glyco-codes (glycan-related information) to be deciphered by glycan-binding proteins (GBPs). The intervillous space of the placenta is richly populated with membrane-bound and secreted glycoproteins. Evidence exists to suggest that altering the structural nature of their N-glycans can impact several trophoblast functions, which include those related to interactions with decidual cells. This review summarizes trophoblast-related activities influenced by N-glycan-GBP recognition, exploring how different subtypes of trophoblasts actively adapt to characteristics of the decidualized endometrium through cell-specific expression of N-glycosylated proteins, and how these cells receive decidua-derived signals via N-glycan-GBP interactions. We highlight work on how changes in N-glycosylation relates to the success of trophoblast infiltration, interactions of immunomodulators, and uterine angiogenesis. We also discuss studies that suggest aberrant N-glycosylation of trophoblasts may contribute to the pathogenesis of pregnancy complications (e.g. pre-eclampsia, early spontaneous miscarriages and hydatidiform mole). We propose that a more in-depth understanding of how N-glycosylation shapes trophoblast phenotype during early pregnancy has the potential to improve our approach to predicting, diagnosing and alleviating poor maternal/fetal outcomes associated with placental dysfunction.

Journal article

Passmore IJ, Faulds-Pain A, Abouelhadid S, Harrison MA, Hall CL, Hitchen P, Dell A, Heap JT, Wren BWet al., 2023, A combinatorial DNA assembly approach to biosynthesis of N-linked glycans in E. coli, GLYCOBIOLOGY, Vol: 33, Pages: 138-149, ISSN: 0959-6658

Journal article

Chakraborty A, Perez M, Carroll JD, Antonopoulos A, Dell A, Ortega L, Mohammed NBB, Wells M, Staudinger C, Griswold A, Chandler KB, Marrero C, Jimenez R, Tani Y, Wilmott JS, Thompson JF, Wang W, Sackstein R, Scolyer RA, Murphy GF, Haslam SM, Dimitroff CJet al., 2023, Hypoxia Controls the Glycome Signature and Galectin-8-Ligand Axis to Promote Protumorigenic Properties of Metastatic Melanoma., J Invest Dermatol, Vol: 143, Pages: 456-469.e8

The prognosis for patients with metastatic melanoma (MM) involving distant organs is grim, and treatment resistance is potentiated by tumor-initiating cells (TICs) that thrive under hypoxia. MM cells, including TICs, express a unique glycome featuring i-linear poly-N-acetyllactosamines through the loss of I-branching enzyme, β1,6 N-acetylglucosaminyltransferase 2. Whether hypoxia instructs MM TIC development by modulating the glycome signature remains unknown. In this study, we explored hypoxia-dependent alterations in MM glycome‒associated genes and found that β1,6 N-acetylglucosaminyltransferase 2 was downregulated and a galectin (Gal)-8-ligand axis, involving both extracellular and cell-intrinsic Gal-8, was induced. Low β1,6 N-acetylglucosaminyltransferase 2 levels correlated with poor patient outcomes, and patient serum samples were elevated for Gal-8. Depressed β1,6 N-acetylglucosaminyltransferase 2 in MM cells upregulated TIC marker, NGFR/CD271, whereas loss of MM cell‒intrinsic Gal-8 markedly lowered NGFR and reduced TIC activity in vivo. Extracellular Gal-8 bound preferentially to i-linear poly-N-acetyllactosamines on N-glycans of the TIC marker and prometastatic molecule CD44, among other receptors, and activated prosurvival factor protein kinase B. This study reveals the importance of hypoxia governing the MM glycome by enforcing i-linear poly-N-acetyllactosamine and Gal-8 expression. This mechanistic investigation also uncovers glycome-dependent regulation of pro-MM factor, NGFR, implicating i-linear poly-N-acetyllactosamine and Gal-8 as biomarkers and therapeutic targets of MM.

Journal article

Yung HW, Zhao X, Glover L, Burrin C, Pang P-C, Jones CJP, Gill C, Duhig K, Olovsson M, Chappell LC, Haslam SM, Dell A, Burton GJ, Charnock-Jones DSet al., 2023, Perturbation of placental protein glycosylation by endoplasmic reticulum stress promotes maladaptation of maternal hepatic glucose metabolism, iScience, Vol: 26, ISSN: 2589-0042

Placental hormones orchestrate maternal metabolic adaptations to support pregnancy. We hypothesized that placental ER stress, which characterizes early-onset pre-eclampsia (ePE), compromises glycosylation, reducing hormone bioactivity and these maladaptations predispose the mother to metabolic disease in later life. We demonstrate ER stress reduces the complexity and sialylation of trophoblast protein N-glycosylation, while aberrant glycosylation of vascular endothelial growth factor reduced its bioactivity. ER stress alters the expression of 66 of the 146 genes annotated with “protein glycosylation” and reduces the expression of sialyltransferases. Using mouse placental explants, we show ER stress promotes the secretion of mis-glycosylated glycoproteins. Pregnant mice carrying placentas with junctional zone-specific ER stress have reduced blood glucose, anomalous hepatic glucose metabolism, increased cellular stress and elevated DNA methyltransferase 3A. Using pregnancy-specific glycoproteins as a readout, we also demonstrate aberrant glycosylation of placental proteins in women with ePE, thus providing a mechanistic link between ePE and subsequent maternal metabolic disorders.

Journal article

Wu G, Grassi P, MacIntyre D, Molina BG, Sykes L, Kundu S, Hsiao C-T, Khoo K-H, Bennett P, Dell A, Haslam Set al., 2022, N-Glycosylation of cervicovaginal fluid reflects microbial community, immune activity, and pregnancy status, Scientific Reports, Vol: 12, Pages: 1-14, ISSN: 2045-2322

Human cervicovaginal fluid (CVF) is a complex, functionally important and glycan rich biological fluid, fundamental in mediating physiological events associated with reproductive health. Using a comprehensive glycomic strategy we reveal an extremely rich and complex N-glycome in CVF of pregnant and non-pregnant women, abundant in paucimannose and highmannose glycans, complex glycans with 2-4 N-Acetyllactosamine (LacNAc) antennae, and Poly-LacNAc glycans decorated with fucosylation and sialylation. N-glycosylation variations were observed to differ in relation to pregnancy status, microbial composition, immune activation, and pregnancy outcome. Compared to CVF from women experiencing term birth, CVF from women who subsequently experienced preterm birth showed lower sialylation, which correlated to the presence of a diverse microbiome, and higher fucosylation, which correlated positively to pro-inflammatory cytokine concentration. This study is the first step towards better understanding the role of cervicovaginal glycans in reproductive health, their contribution to the mechanism of microbial driven preterm birth, and their potential forpreventative therapy.

Journal article

Mohammed NBB, Antonopoulos A, Dell A, Haslam SM, Dimitroff CJet al., 2022, The pleiotropic role of galectin-3 in melanoma progression: Unraveling the enigma, Advances in Cancer Research, ISSN: 0065-230X

Melanoma is a highly aggressive skin cancer with poor outcomes associated with distant metastasis. Intrinsic properties of melanoma cells alongside the crosstalk between melanoma cells and surrounding microenvironment determine the tumor behavior. Galectin-3 (Gal-3), a ß-galactoside-binding lectin, has emerged as a major effector in cancer progression, including melanoma behavior. Data from melanoma models and patient studies reveal that Gal-3 expression is dysregulated, both intracellularly and extracellularly, throughout the stages of melanoma progression. This review summarizes the most recent data and hypotheses on Gal-3 and its tumor-modulating functions, highlighting its role in driving melanoma growth, invasion, and metastatic colonization. It also provides insight into potential Gal-3-targeted strategies for melanoma diagnosis and treatment.

Journal article

Cao H, Mathur A, Robertson C, Antonopoulos A, Henderson S, Girard L-P, Wong JH, Davie A, Wright S, Brewin J, Rees DC, Dell A, Haslam SM, Vickers MAet al., 2022, Measurement of erythrocyte membrane mannoses to assess splenic function, British Journal of Haematology, Vol: 198, Pages: 155-164, ISSN: 0007-1048

Red blood cells (RBCs) lose plasma membrane in the spleen as they age, but the cells and molecules involved are yet to be identified. Sickle cell disease and infection by Plasmodium falciparum cause oxidative stress that induces aggregates of cross-linked proteins with N-linked high-mannose glycans (HMGs). These glycans can be recognised by mannose-binding lectins, including the mannose receptor (CD206), expressed on macrophages and specialised phagocytic endothelial cells in the spleen to mediate the extravascular haemolysis characteristic of these diseases. We postulated this system might also mediate removal of molecules and membrane in healthy individuals. Surface expression of HMGs on RBCs from patients who had previously undergone splenectomy was therefore assessed: high levels were indeed observable as large membrane aggregates. Glycomic analysis by mass spectrometry identified a mixture of Man5-9GlcNAc2 structures. HMG levels correlated well with manual pit counts (r = 0.75–0.85). To assess further whether HMGs might act as a splenic reticuloendothelial function test, we measured levels on RBCs from patients with potential functional hyposplenism, some of whom exhibited high levels that may indicate risk of complications.

Journal article

Mountain K, MacIntyre D, Chan D, Hyde A, Lee Y, Brown R, David A, Dell A, Ten F, Haslam S, Liu Y, Lewis H, Norman J, Stock S, Teoh TG, Terzidou V, Kundu S, Bennett P, Sykes Let al., 2022, Blood group antigens influence host-microbe interactions and risk of early preterm birth, Publisher: WILEY, Pages: 55-56, ISSN: 1470-0328

Conference paper

Duncombe L, Howells L, Haughey A, Taylor AV, Kaveh D, Erdenliğ Gϋrbilek S, Dell A, Hitchen PG, Haslam SM, Mandal SS, Ganesh NV, Bundle DR, McGiven Jet al., 2022, The tip of brucella O-Polysaccharide is a potent epitope in response to brucellosis infection and enables short synthetic antigens to be superior diagnostic reagents, Microorganisms, Vol: 10, Pages: 1-19, ISSN: 2076-2607

Brucellosis is a global disease and the world’s most prevalent zoonosis. All cases in livestock and most cases in humans are caused by members of the genus Brucella that possess a surface O-polysaccharide (OPS) comprised of a rare monosaccharide 4-deoxy-4-formamido-D-mannopyranose assembled with α1,2 and α1,3 linkages. The OPS of the bacterium is the basis for serodiagnostic tests for brucellosis. Bacteria that also contain the same rare monosaccharide can induce antibodies that cross-react in serological tests. In previous work we established that synthetic oligosaccharides, representing elements of the Brucella A and M polysaccharide structures, were excellent antigens to explore the antibody response in the context of infection, immunisation and cross reaction. These studies suggested the existence of antibodies that are specific to the tip of the Brucella OPS. Sera from naturally and experimentally Brucella abortus-infected cattle as well as from cattle experimentally infected with the cross-reactive bacterium Yersinia enterocolitica O:9 and field sera that cross react in conventional serological assays were studied here with an expanded panel of synthetic antigens. The addition of chemical features to synthetic antigens that block antibody binding to the tip of the OPS dramatically reduced their polyclonal antibody binding capability providing conclusive evidence that the OPS tip (non-reducing end) is a potent epitope. Selected short oligosaccharides, including those that were exclusively α1,2 linked, also demonstrated superior specificity when evaluated with cross reactive sera compared to native smooth lipopolysaccharide (sLPS) antigen and capped native OPS. This surprising discovery suggests that the OPS tip epitope, even though common to both Brucella and Y. enterocolitica O:9, has more specific diagnostic properties than the linear portion of the native antigens. This finding opens the way to the development of improved serological

Journal article

Kundu S, Lee Y, Sykes L, Chan D, Lewis H, Brown R, Kindinger L, Dell A, Feizi T, Haslam S, Liu Y, Marchesi J, MacIntyre D, Bennett Pet al., 2022, The Effect of Secretor Status and the Vaginal Microbiome on Birth Outcome, Publisher: SPRINGER HEIDELBERG, Pages: 197-197, ISSN: 1933-7191

Conference paper

Edwards E, Livanos M, Krueger A, Dell A, Haslam SM, Mark Smales C, Bracewell DGet al., 2022, Strategies to control therapeutic antibody glycosylation during bioprocessing: synthesis and separation., Biotechnology and Bioengineering, Vol: 119, ISSN: 0006-3592

Glycosylation can be a critical quality attribute (CQA) in biologic manufacturing. In particular, it has implications on the half-life, immunogenicity and pharmacokinetics of therapeutic monoclonal antibodies (mAbs) and must be closely monitored throughout drug development and manufacturing. To address this, advances have been made primarily in upstream processing, including mammalian cell line engineering to yield more predictably glycosylated mAbs, and the addition of media supplements during fermentation to manipulate the metabolic pathways involved in glycosylation. A more robust approach would be a conjoined upstream-downstream processing strategy. This could include implementing novel downstream technologies, such as the use of Fc gamma-based affinity ligands for the separation of mAb glycovariants. This review highlights the importance of controlling therapeutic antibody glycosylation patterns, the challenges faced in terms of glycosylation during mAb biosimilar development, current efforts both upstream and downstream to control glycosylation and their limitations, and the need for research in the downstream space in order to establish holistic and consistent manufacturing processes for the production of antibody therapies. This article is protected by copyright. All rights reserved.

Journal article

Peswani AR, Narkpuk J, Krueger A, Bracewell DG, Lekcharoensuk P, Haslam SM, Dell A, Jaru-Ampornpan P, Robinson Cet al., 2022, Novel constructs and 1-step chromatography protocols for the production of Porcine Circovirus 2d (PCV2d) and Circovirus 3 (PCV3) subunit vaccine candidates, FOOD AND BIOPRODUCTS PROCESSING, Vol: 131, Pages: 125-135, ISSN: 0960-3085

Journal article

Butta NV, Haslam SM, Dell A, Xuan LK, Ball S, Arias-Salgado EG, Monzon Manzano E, Acuna P, Canales M, Jimenez-Yuste V, Alvarez Roman MTet al., 2021, Glycomic Characterization of Platelets from Patients with Immune Thrombocytopenia, 63rd Annual Meeting and Exposition of the American-Society-of-Hematology (ASH), Publisher: AMER SOC HEMATOLOGY, ISSN: 0006-4971

Conference paper

Baksmeier C, Blundell P, Steckel J, Schultz V, Gu Q, Da Silva Filipe A, Kohl A, Linnington C, Lu D, Dell A, Haslam S, Wang J, Czajkowsky D, Goebels N, Pleass RJet al., 2021, Modified recombinant human IgG1-Fc is superior to natural IVIG at inhibiting immune-mediated demyelination., Immunology, Vol: 464, Pages: 90-105, ISSN: 0019-2805

Intravenous immunoglobulin (IVIG) is an established treatment for numerous autoimmune conditions. Although Fc fragments derived from IVIG have shown efficacy in controlling immune thrombocytopenia (ITP) in children, the mechanisms of action are unclear and controversial. The aim of this study is to dissect IVIG effector mechanisms using further adapted Fc fragments on demyelination in an ex vivo model of the central nervous system (CNS)-immune interface. Using organotypic cerebellar slice cultures (OSC) from transgenic mice we induced extensive immune-mediated demyelination and oligodendrocyte loss with an antibody specific for myelin oligodendrocyte glycoprotein (MOG) and complement. Protective effects of adapted Fc fragments were assessed by live imaging of GFP expression, immunohistochemistry and confocal microscopy. Cysteine and glycan adapted Fc fragments protected OSC from demyelination in a dose-dependent manner where equimolar concentrations of either IVIG or control Fc were ineffective. The protective effects of the adapted Fc fragments are partly attributed to interference with complement-mediated oligodendroglia damage. Transcriptome analysis ruled out signatures associated with inflammatory or innate immune responses. Taken together our findings show that recombinant biomimetics can be made that are at least two hundred-fold more effective than IVIG in controlling demyelination by anti-MOG antibodies.

Journal article

Bonnardel F, Haslam SM, Dell A, Feizi T, Liu Y, Tajadura-Ortega V, Akune Y, Sykes L, Bennett PR, MacIntyre DA, Lisacek F, Imberty Aet al., 2021, Proteome-wide prediction of bacterial carbohydrate-binding proteins as a tool for understanding commensal and pathogen colonisation of the vaginal microbiome, npj Biofilms and Microbiomes, Vol: 7, Pages: 1-10, ISSN: 2055-5008

Bacteria use carbohydrate-binding proteins (CBPs), such as lectins and carbohydrate-binding modules (CBMs), to anchor to specific sugars on host surfaces. CBPs in the gut microbiome are well studied, but their roles in the vagina microbiome and involvement in sexually transmitted infections, cervical cancer and preterm birth are largely unknown. We established a classification system for lectins and designed Hidden Markov Model (HMM) profiles for data mining of bacterial genomes, resulting in identification of >100,000 predicted bacterial lectins available at unilectin.eu/bacteria. Genome screening of 90 isolates from 21 vaginal bacterial species shows that those associated with infection and inflammation produce a larger CBPs repertoire, thus enabling them to potentially bind a wider array of glycans in the vagina. Both the number of predicted bacterial CBPs and their specificities correlated with pathogenicity. This study provides new insights into potential mechanisms of colonisation by commensals and potential pathogens of the reproductive tract that underpin health and disease states.

Journal article

Ng BG, Sosicka P, Fenaille F, Harroche A, Vuillaumier-Barrot S, Porterfield M, Xia Z-J, Wagner S, Bamshad MJ, Vergnes-Boiteux M-C, Cholet S, Dalton S, Dell A, Dupre T, Fiore M, Haslam SM, Huguenin Y, Kumagai T, Kulik M, McGoogan K, Michot C, Nickerson DA, Pascreau T, Borgel D, Raymond K, Warad D, Flanagan-Steet H, Steet R, Tiemeyer M, Seta N, Bruneel A, Freeze HHet al., 2021, A mutation in <i>SLC37A4</i> causes a dominantly inherited congenital disorder of glycosylation characterized by liver dysfunction, AMERICAN JOURNAL OF HUMAN GENETICS, Vol: 108, Pages: 1040-1052, ISSN: 0002-9297

Journal article

Wu G, Murugesan G, Nagala M, McCraw A, Haslam SM, Dell A, Crocker PRet al., 2021, Activation of regulatory T cells triggers specific changes in glycosylation associated with Siglec-1-dependent inflammatory responses [version 1; peer review: 2 approved], Wellcome Open Research, Vol: 6, ISSN: 2398-502X

Background: Siglec-1 is a macrophage lectin-like receptor that mediates sialic acid-dependent cellular interactions. Its upregulation on macrophages in autoimmune disease was shown previously to promote inflammation through suppressing the expansion of regulatory T cells (Tregs). Here we investigate the molecular basis for Siglec-1 binding to Tregs using in vitro-induced cells as a model system. Methods: Glycosylation changes that affect Siglec‑1 binding were studied by comparing activated and resting Tregs using RNA-Seq, glycomics, proteomics and binding of selected antibodies and lectins. A proximity labelling and proteomics strategy was used to identify Siglec-1 counter-receptors expressed on activated Tregs. Results: Siglec-1 binding was strongly upregulated on activated Tregs, but lost under resting conditions. Glycomics revealed changes in N-glycans and glycolipids following Treg activation and we observed changes in expression of multiple 'glycogenes' that could lead to the observed increase in Siglec-1 binding. Proximity labelling of intact, living cells identified 49 glycoproteins expressed by activated Tregs that may function as Siglec-1 counter-receptors. These represent ~5% of the total membrane protein pool and were mainly related to T cell activation and proliferation. We demonstrate that several of these counter-receptors were upregulated following activation of Tregs and provide initial evidence that their altered glycosylation may also be important for Siglec-1 binding. Conclusions: We provide the first comprehensive analysis of glycan changes that occur in activated Tregs, leading to recognition by the macrophage lectin, Siglec-1 and suppression of Treg expansion. We furthermore provide insights into glycoprotein counter-receptors for Siglec-1 expressed by activated Tregs that are likely to be important for suppressing Treg expansion.

Journal article

Wang S-S, Solar VD, Yu X, Antonopoulos A, Friedman AE, Agarwal K, Garg M, Ahmed SM, Addhya A, Nasirikenari M, Lau JT, Dell A, Haslam SM, Sampathkumar S-G, Neelamegham Set al., 2021, Efficient inhibition of O-glycan biosynthesis using the hexosamine analog Ac<sub>5</sub>GalNTGc, CELL CHEMICAL BIOLOGY, Vol: 28, Pages: 699-+, ISSN: 2451-9448

Journal article

Wang Y, Khan A, Antonopoulos A, Bouche L, Buckley CD, Filer A, Raza K, Li K-P, Tolusso B, Gremese E, Kurowska-Stolarska M, Alivernini S, Dell A, Haslam SM, Pineda MAet al., 2021, Loss of α2-6 sialylation promotes the transformation of synovial fibroblasts into a pro-inflammatory phenotype in arthritis, Nature Communications, Vol: 12, ISSN: 2041-1723

In healthy joints, synovial fibroblasts (SFs) provide the microenvironment required to mediate homeostasis, but these cells adopt a pathological function in rheumatoid arthritis (RA). Carbohydrates (glycans) on cell surfaces are fundamental regulators of the interactions between stromal and immune cells, but little is known about the role of the SF glycome in joint inflammation. Here we study stromal guided pathophysiology by mapping SFs glycosylation pathways. Combining transcriptomic and glycomic analysis, we show that transformation of fibroblasts into pro-inflammatory cells is associated with glycan remodeling, a process that involves TNF-dependent inhibition of the glycosyltransferase ST6Gal1 and α2-6 sialylation. SF sialylation correlates with distinct functional subsets in murine experimental arthritis and remission stages in human RA. We propose that pro-inflammatory cytokines remodel the SF-glycome, converting the synovium into an under-sialylated and highly pro-inflammatory microenvironment. These results highlight the importance of glycosylation in stromal immunology and joint inflammation.

Journal article

Cao H, Antonopoulos A, Henderson S, Wassall H, Brewin J, Masson A, Shepherd J, Konieczny G, Patel B, Williams M-L, Davie A, Forrester MA, Hall L, Minter B, Tampakis D, Moss M, Lennon C, Pickford W, Erwig L, Robertson B, Dell A, Brown GD, Wilson HM, Rees DC, Haslam SM, Rowe JA, Barker RN, Vickers MAet al., 2021, Red blood cell mannoses as phagocytic ligands mediating both sickle cell anaemia and malaria resistance, Nature Communications, Vol: 12, Pages: 1-13, ISSN: 2041-1723

In both sickle cell disease and malaria, red blood cells (RBCs) are phagocytosed in the spleen, but receptor-ligand pairs mediating uptake have not been identified. Here, we report that patches of high mannose N-glycans (Man5-9GlcNAc2), expressed on diseased or oxidized RBC surfaces, bind the mannose receptor (CD206) on phagocytes to mediate clearance. We find that extravascular hemolysis in sickle cell disease correlates with high mannose glycan levels on RBCs. Furthermore, Plasmodium falciparum-infected RBCs expose surface mannose N-glycans, which occur at significantly higher levels on infected RBCs from sickle cell trait subjects compared to those lacking hemoglobin S. The glycans are associated with high molecular weight complexes and protease-resistant, lower molecular weight fragments containing spectrin. Recognition of surface N-linked high mannose glycans as a response to cellular stress is a molecular mechanism common to both the pathogenesis of sickle cell disease and resistance to severe malaria in sickle cell trait.

Journal article

Antonopoulos A, Broome S, Sharov V, Ziegenfuss C, Easton RL, Panico M, Morris HR, Haslam Set al., 2021, Site-specific characterisation of SARS-CoV-2 spike glycoprotein receptor binding domain, Glycobiology, Vol: 31, Pages: 181-187, ISSN: 0959-6658

The novel coronavirus SARS-CoV-2, the infective agent causing COVID-19, is having a global impact both in terms of human disease as well as socially and economically. Its heavily glycosylated spike glycoprotein is fundamental for the infection process, via its receptor binding domains interaction with the glycoprotein angiotensin converting enzyme 2 on human cell surfaces. We therefore utilized an integrated glycomic and glycoproteomic analytical strategy to characterise both N- and O- glycan site specific glycosylation within the receptor binding domain. We demonstrate the presence of complex type N-glycans with unusual fucosylated LacdiNAc at both sites N331 and N343 and a single site of O-glycosylation on T323.

Journal article

Mendoza M, Lu D, Ballesteros A, Blois SM, Abernathy K, Feng C, Dimitroff CJ, Zmuda J, Panico M, Dell A, Vasta GR, Haslam SM, Dveksler Get al., 2020, Glycan characterization of pregnancy-specific glycoprotein 1 and its identification as a novel Galectin-1 ligand, Glycobiology, Vol: 30, Pages: 895-909, ISSN: 0959-6658

Pregnancy-specific beta 1 glycoprotein (PSG1) is secreted from trophoblast cells of the human placenta in increasing concentrations as pregnancy progresses, becoming one of the most abundant proteins in maternal serum in the third trimester. PSG1 has seven potential N-linked glycosylation sites across its four domains. We carried out glycomic and glycoproteomic studies to characterize the glycan composition of PSG1 purified from serum of pregnant women and identified the presence of complex N-glycans containing poly LacNAc epitopes with α2,3 sialyation at four sites. Using different techniques, we explored whether PSG1 can bind to galectin-1 (Gal-1) as these two proteins were previously shown to participate in processes required for a successful pregnancy. We confirmed that PSG1 binds to Gal-1 in a carbohydrate-dependent manner with an affinity of the interaction of 0.13 μM. In addition, we determined that out of the three N-glycosylation-carrying domains, only the N and A2 domains of recombinant PSG1 interact with Gal-1. Lastly, we observed that the interaction between PSG1 and Gal-1 protects this lectin from oxidative inactivation and that PSG1 competes the ability of Gal-1 to bind to some but not all of its glycoprotein ligands.

Journal article

Loxley GM, Hooks DO, Antonopoulos A, Dell A, Haslam SM, Linklater WL, Hurst JL, Beynon RJet al., 2020, Vulpeculin: a novel and abundant lipocalin in the urine of the common brushtail possum, Trichosurus vulpecula, Open Biology, Vol: 10, ISSN: 2046-2441

Lipocalins are a family of secreted proteins. They are capable of binding small lipophilic compounds and have been extensively studied for their role in chemosignalling in rodent urine. Urine of the common brushtail possum (Trichosurus vulpecula) contains a prominent glycoprotein of 20 kDa, expressed in both sexes. We have isolated this protein and determined its primary sequence by mass spectrometry, including the use of metabolic labelling to resolve the leucine/isoleucine isobaric ambiguity. The protein sequence was identified as a lipocalin, and phylogenetic analysis grouped the protein with other marsupial lipocalin sequences in a phylogenetic clade distinct from established cross-species lipocalin sub-families. The pattern of expression in possum urine and the similarity in sequence and structure to other lipocalins suggests this protein may have a role in brushtail possum chemosignalling.

Journal article

Li H, Marceau M, Yang T, Liao T, Tang X, Hu R, Xie Y, Tang H, Tay A, Shi Y, Shen Y, Yang T, Pi X, Lamichhane B, Luo Y, Debowski AW, Nilsson H, Haslam SM, Mulloy B, Dell A, Stubbs KA, Marshall BJ, Benghezal Met al., 2020, East-Asian <i>Helicobacter pylori</i> strains synthesize heptan-deficient lipopolysaccharide, Publisher: WILEY, Pages: 75-75, ISSN: 1083-4389

Conference paper

Osimanjiang W, Santos Roballo KC, Houck BD, Ito M, Antonopoulos A, Dell A, Haslam SM, Bushman JSet al., 2020, Analysis of N- and O-linked glycosylation: differential glycosylation after rat spinal cord injury, Journal of Neurotrauma, Vol: 37, Pages: 1954-1962, ISSN: 0897-7151

Glycosylation is a fundamental cellular process that has a dramatic impact on the functionality of glycoconjugates such as proteins or lipids and mediates many different biological interactions including cell migration, cellular signaling, and synaptic interactions in the nervous system. In spinal cord injury (SCI), all of these cellular processes are altered, but the potential contributions of glycosylation changes to these alterations has not been thoroughly investigated.We studied the glycosylation of injured spinal cord tissue from rats that received a contusion SCI. The N- and O-linked glycosylation was assessed at 3 and 14 days post-injury (DPI), and compared with uninjured control and time-matched sham spinal tissue. Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) and tandem MS (MS/MS) were performed to analyze carbohydrate structures. Results revealed diverse and abundant glycosylation in all groups, with some carbohydrate structures differentially produced in SCI animals compared with uninjured controls and shams. One such change occurred in the abundance of the Sda structure, Neu5Ac-α-(2,3)-[GalNAc-β-(1,4)-]Gal-β-(1,4)-GlcNAc, which was increased in SCI samples compared with shams and non-injured controls. Immunohistochemistry (IHC) and western blot were performed on SCI and sham samples using the CT1 antibody, which recognizes the terminal trisaccharide of Sda with high specificity. Both of these metrics confirmed elevated Sda structure in SCI tissue, where IHC further showed that Sda is expressed mainly by microglia. The results of these studies suggest that SCI causes a significant alteration in N- and O-linked glycosylation.

Journal article

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