Biology subjects

Cobb, B. A.

Publications and source records attributed to Cobb, B. A..

7 recordsLinked to original sources

bioRxiv2025

Not all alloantibodies are created equal: IgG glycosylation and severity of antibody-mediated rejection in kidney transplantation

IntroductionAntibody-mediated rejection (AMR) is a leading cause of kidney transplant (KT) failure, driven by donor-specific anti-HLA antibodies (DSA). However, not all patients with DSA experience accelerated graft loss, suggesting that factors beyond antibody presence influence AMR severity. Post-translational modifications, particularly glycosylation of Immunoglobulin-G (IgG), play a critical role in modulating antibody function. This study investigates the association between IgG glycosylation profiles and the risk and severity of AMR in KT recipients. MethodsWe prospectively analyzed 65 KT patients, including 26 with acute AMR (aAMR), 27 with chronic-active AMR (caAMR), and 12 controls without rejection. IgG glycosylation was quantified using lectin-based ELISA, focusing on mannose, fucose, sialic acid, and bisecting N-acetylglucosamine (GlcNAc) levels. ResultsResults showed that bisecting GlcNAc levels of total IgG were significantly higher in caAMR patients than controls (p=0.019) and aAMR patients (p=0.045). Multivariable analysis revealed that higher bisecting GlcNAc levels of IgG were independently associated with glomerulitis [g-score, OR: 2.7 (95%CI: 1.2-6.7), p=0.019] and chronic glomerulopathy [cg-score, OR: 2.8 (95%CI: 1.3-7.5), p=0.021], independent of DSA presence. ConclusionsThese findings indicate an association between IgG glycosylation, particularly bisecting GlcNAc, and AMR severity. IgG glycosylation profiles could serve as biomarkers for AMR risk and severity, offering new insights into the mechanisms of AMR and potential therapeutic targets.

immunology↗

bioRxiv2025

Pregnant Dairy Heifers Express Influenza A Virus Receptors in the Mammary Gland

Highly pathogenic avian influenza (HPAI) A(H5N1) virus emerged in lactating dairy cattle in March 2024, causing mastitis-related disease and infections in other farm animals and workers. Recent work identified 2,6 and 2,3-linked sialic acids (SA), which serve as influenza virus receptors, in the lactating bovine mammary gland; however, their distribution across stages of mammary growth and development remains unknown. We compared the distribution of tissue sialylation in mammary glands of prepubertal dairy calves, pregnant dairy heifers, and lactating cows. Mammary glands at all physiological stages expressed both 2,6 SA, the preferred receptor linkage for human influenza viruses, and 2,3 SA, the preferred receptor linkage for avian influenza viruses. Importantly, mammary glands of pregnant dairy heifers exhibited the highest overall expression of 2,3 SA, observed in both tissue and alveolar lumens. Our results suggest that pregnant dairy heifers, like lactating dairy cows, are susceptible to H5N1 infection in the mammary gland.

pathology↗

bioRxiv2023

IgG sialylation occurs via the FcRn-mediated recycling pathway in endothelial cells

IgG is a key mediator of immune responses throughout the human body, and the structure of the conserved glycan on the Fc region has been identified as a key inflammatory switch regulating its downstream effects. In particular, the absence of terminal sialic acid has been shown to increase the affinity of IgG for activating Fc receptors, cascading the inflammatory response in a variety of diseases and conditions. Previously, we have shown that IgG sialylation is mediated by B cell-extrinsic processes. Here, we show that the FcRn-mediated recycling pathway within endothelial cells is a critical modulator of IgG sialylation. Building a deeper understanding of how IgG sialylation is regulated will drive the development of novel therapeutics which dynamically tune IgG functionality in vivo. One-Sentence SummaryEndothelial cells remodel IgG glycans within the FcRn-mediated recycling pathway.

immunology↗

bioRxiv2023

Platelet-Localized ST6Gal1 Does Not Impact IgG Sialylation

The IgG antibody class forms an important basis of the humoral immune response, conferring reciprocal protection from both pathogens and autoimmunity. IgG function is determined by the IgG subclass, as defined by the heavy chain, as well as the glycan composition at N297, the conserved site of N-glycosylation within the Fc domain. For example, lack of core fucose promotes increased antibody-dependent cellular cytotoxicity, whereas 2,6-linked sialylation by the enzyme ST6Gal1 helps to drive immune quiescence. Despite the immunological significance of these carbohydrates, little is known about how IgG glycan composition is regulated. We previously reported that mice with ST6Gal1-deficient B cells have unaltered IgG sialylation. Likewise, ST6Gal1 released into the plasma by hepatocytes does not significantly impact overall IgG sialylation. Since IgG and ST6Gal1 have independently been shown to exist in platelet granules, it was possible that platelet granules could serve as a B cell-extrinsic site for IgG sialylation. To address this hypothesis, we used a platelet factor 4 (Pf4)-Cre mouse to delete ST6Gal1 in megakaryocytes and platelets alone or in combination with an albumin-Cre mouse to also remove it from hepatocytes and the plasma. The resulting mouse strains were viable and had no overt pathological phenotype. We also found that despite targeted ablation of ST6Gal1, no change in IgG sialylation was apparent. Together with our prior findings, we can conclude that in mice, neither B cells, the plasma, nor platelets have a substantial role in homeostatic IgG sialylation.

immunology↗

bioRxiv2022

Plasma ST6Gal1 is Dispensable for IgG Sialylation

The glycosylation of IgG has attracted increased attention due to the impact of N-glycan modifications at N297 on IgG function, acting primarily through modulation of Fc domain conformation and Fc{gamma} receptor binding affinities and signaling. However, the mechanisms regulating IgG glycosylation and especially 2,6-sialylation of its N-glycan remain poorly understood. We observed previously that IgG is normally sialylated in mice with B cells lacking the sialyltransferase ST6Gal1. This supported the hypothesis that IgG may be sialylated outside of B cells, perhaps through the action of hepatocyte-released plasma ST6Gal1. Here we demonstrate that this model is incorrect. Animals lacking hepatocyte expressed ST6Gal1 retain normal IgG 2,6-sialylation, despite the lack of detectable ST6Gal1 in plasma. Moreover, we confirmed that B cells were not a redundant source of IgG sialylation. Thus, while 2,6-sialylation is lacking in IgG from mice with germline ablation of ST6Gal1, IgG 2,6-sialylation is normal in mice lacking ST6Gal1 in either hepatocytes or B cells. These results indicate that IgG 2,6-sialylation arises after release from a B cell, but is not dependent on plasma-localized ST6Gal1 activity.

immunology↗

bioRxiv2022

Divergent Golgi Trafficking Limits B cell-Mediated IgG Sialylation

The degree of 2,6-linked sialylation on IgG glycans is associated with a variety of inflammatory conditions and is thought to drive IgG anti-inflammatory activity. Previous findings revealed that ablation of {beta}-galactoside 2,6-sialyltransferase 1 (ST6Gal1) in B cells failed to alter IgG sialylation in vivo, yet resulted in the loss of B cell surface 2,6 sialylation, suggesting divergent pathways for IgG and cell surface glycoprotein glycosylation and trafficking. Employing both B cell hybridomas and ex vivo murine B cells, we discovered that IgG was poorly sialylated by ST6Gal1 and highly core fucosylated by 1,6-fucosyltransferase 8 (Fut8) in cell culture. In contrast, IgG-producing cells showed the opposite pattern by flow cytometry, with high cell surface 2,6 sialylation and low 1,6 fucosylation. Paired studies further revealed that ex vivo B cell-produced IgG carried significantly less sialylation compared to IgG isolated from the plasma of matched animals, providing evidence that sialylation increases after release in vivo. Finally, confocal analyses demonstrated that IgG poorly localized to subcellular compartments rich in sialylation and ST6Gal1, and strongly to regions rich in fucosylation and Fut8. These findings support a model in which IgG subcellular trafficking diverges from the canonical secretory pathway by promoting Fut8-mediated core fucosylation and limiting exposure to and modification by ST6Gal1, providing a mechanism for why B cell-expressed ST6Gal1 is dispensable for IgG sialylation in vivo.

immunology↗

bioRxiv2020

Integration of IL-2 and IL-4 Signals Coordinates Divergent Regulatory T cell Responses and Drives Therapeutic Efficacy

Cells exist within complex milieus of communicating factors, such as cytokines, that combine to generate context-specific responses, yet nearly all knowledge about the function of each cytokine and the signaling propagated downstream of their recognition is based upon the response to individual cytokines. Here, we found that regulatory T cells (Tregs) integrate concurrent signaling initiated by IL-2 and IL-4 to generate a response divergent from the sum of the two pathways in isolation. IL-4 stimulation of STAT6 phosphorylation was blocked by IL-2, while IL-2 and IL-4 synergized to enhance STAT5 phosphorylation, IL-10 production, and the selective proliferation of IL-10-producing Tregs, leading to increased inhibition of conventional T cell activation and the reversal of asthma and multiple sclerosis in mice. These data define a mechanism of combinatorial cytokine signaling and lay the foundation upon which to better understand the origins of cytokine pleiotropy while informing improved the clinical use of cytokines. Impact StatementSimultaneous cytokine signaling results in unexpected transcription factor changes that fuel a cellular response divergent from the sum of each cytokine alone.

biochemistry↗