Biology subjects

Sustar, V.

Publications and source records attributed to Sustar, V..

4 recordsLinked to original sources

bioRxiv2020

APEX2 proximity biotinylation reveals protein dynamics triggered by B cell receptor activation

Successful B cell activation, critical for high-affinity antibody production, is controlled by the B cell antigen receptor (BCR). However, we still lack a comprehensive protein-level view of the very dynamic multi-branched cellular events triggered by antigen binding. Here, we employed APEX2 proximity biotinylation to study antigen-induced changes, 5-15 min after receptor activation, at the vicinity of the plasma membrane lipid rafts, wherein BCR enriches upon activation. The data reveals dynamics of signaling proteins, as well as various players linked to the subsequent processes, such as actin cytoskeleton remodelling and endocytosis. Interestingly, our differential expression analysis identified dynamic responses in various proteins previously not linked to early B cell activation. We demonstrate active SUMOylation at the sites of BCR activation in various conditions and report its functional role in BCR signaling through Akt and MAPK axes.

immunology↗

bioRxiv2020

AutoCoEv: a high-throughput in silico pipeline for revealing novel protein-protein interactions

Protein-protein communications govern cellular processes via complex regulatory networks, that are still far from being understood. Thus, identifying novel interactions between proteins can significantly facilitate our comprehension of the mechanistic principles of protein functions. Co-evolution between proteins is a sign of functional communication and, as such, provides a powerful approach to search for novel direct or indirect molecular partners. However, evolutionary analysis of large arrays of proteins, in silico, is a highly time-consuming effort, which has limited the usage of this method to protein pairs or small protein groups. Here, we developed AutoCoEv, a user-friendly computational pipeline for the search of co-evolution between a large number of proteins. By driving 15 individual programs, culminating in CAPS2 as the software for detecting co-evolution, AutoCoEv achieves seamless automation and parallelization of the workflow. Importantly, we provide a patch to CAPS2 source code to strengthen its statistical output, allowing for multiple comparisons correction and enhanced analysis of the results. We apply the pipeline to inspect co-evolution among 324 proteins identified to locate at the vicinity of the lipid rafts of B lymphocytes. We successfully detected multiple strong coevolutionary relations between the proteins, predicting many novel partners and previously unidentified clusters of functionally related molecules. We conclude that AutoCoEv, available at https://github.com/mattilalab/autocoev, can be used to predict functional interactions from large datasets in a time and cost-efficient manner.

bioinformatics↗

bioRxiv2019

Missing-In-Metastasis / Metastasis Suppressor 1 modulates B cell receptor signaling, B cell metabolic potential and T cell-independent immune responses.

Efficient generation of antibodies by B cells is one of the prerequisites of protective immunity. B cell activation by cognate antigens via B cell receptors (BCR), or pathogen-associated molecules through pattern-recognition receptors, such as Toll-like receptors (TLRs), leads to transcriptional and metabolic changes that ultimately transform B cells into antibody producing plasma cells or memory cells. BCR signaling and a number of steps downstream of it rely on coordinated action of cellular membranes and the actin cytoskeleton, tightly controlled by concerted action of multiple regulatory proteins, some of them exclusive to B cells. Here, we dissect the role of Missing-In-Metastasis (MIM), or Metastasis suppressor 1 (MTSS1), a cancer-associated membrane and actin cytoskeleton regulating protein, in B cell-mediated immunity by taking advantage of MIM knockout mouse strain. We show undisturbed B cell development and largely normal composition of B cell compartments in the periphery. Interestingly, we found that MIM-/- B cells are defected in BCR signaling in response to surface-bound antigens but, on the other hand, show increased metabolic activity after stimulation with LPS or CpG. In vivo, MIM knockout animals exhibit impaired IgM antibody responses to immunization with T cell-independent antigen. This study provides the first comprehensive characterization of MIM in B cells, demonstrates its regulatory role for B cell-mediated immunity, as well as proposes new functions for MIM in tuning receptor signaling and cellular metabolism, processes which may also contribute to the poorly understood functions of MIM in cancer.

immunology↗

bioRxiv2019

Fast targeting of antigen and surface-derived MHCII into degradative compartments implies endosomal prewiring for antigen presentation in B cells.

In order to mount high-affinity antibody responses, B cells internalise specific antigens and process them into peptides loaded onto MHCII for presentation to TO_SCPLOWHC_SCPLOW cells. While the biochemical principles of antigen processing and MHCII loading have been well dissected, how the endosomal vesicle system is wired to enable these specific functions remains much less studied. Here, we performed a systematic microscopy-based analysis of antigen trafficking in B cells to reveal its route to the MHCII peptide-loading compartment (MIIC). Surprisingly, we detected fast targeting of internalised antigen into peripheral acidic compartments that possessed the hallmarks of MIIC and also showed degradative capacity. In these vesicles, internalised antigen converged rapidly with membrane-derived MHCII and partially overlapped with Cathepsin-S and H2-M, both required for peptide loading. These early compartments appeared heterogenous and atypical as they contained a mixture of both early and late markers, indicating specialized endosomal route. Together, our data suggests that, in addition to previously-reported perinuclear late endosomal MIICs, antigen processing and peptide loading could start already in these specialized early peripheral acidic vesicles (eMIIC) to support fast peptide-MHCII presentation.

immunology↗