B Cell Biology

National Institute Of Allergy And Infectious Diseases

Investigators

Linked publications, trials & patents

Abstract

Understanding B cell responses to membrane associated antigens A fundamental feature of B cell responses to antigen in secondary lymphoid organs (SLOs) in vivo is the presentation of the antigen on the surface of FDCs. We compared the responses of human tonsil nave B cells to antigen associated with membranes, in solution or displayed as a virus-like particle in vitro. 1) The role of PIEZO in B cell responses. We discovered that human B cells express the mechano-sensitive membrane calcium channel PIEZO. Using Fluorescence Lifetime Image Microscopy we showed that when B cells interact with antigen on membranes, the B cell membrane is stretched, PIEZO opens and allows the second messenger calcium to flow into the B cell to modulate signaling. Specific inhibitors of PIEZO and knock-down of PIEZO dampen B cell responses to membrane antigens but not to antigens in solution. 2) The molecular basis of the efficacy of the human papillomavirus-virus like particle (HPV-VLP) vaccines to induce long-lasting B cell memory. HPV-VLP vaccines when administered in a single dose demonstrated remarkable prophylactic efficacy. The unexpected potency of one dose of the HVP-VLP vaccine may largely be attributed to structural features of the particles that lead to the efficient generation of long-lived antigen-specific antibody-producing cells. To explore this possibility we established a collaboration with Dr. John Schiller (NCI) a developer of the HPV-VLP vaccines. HPV-VLPs are formed by the self-assembly of the HPV major capsid protein, L1. To determine how HP-VLPs activate B cells we generated human B cell lines that express BCRs containing HP L1-specific mAb IgH and IgL chains and are assessing the responses to the HPV-VLP vaccines versus pentamers of the L1 protein alone. Our preliminary results showed the HPV-VLP induced a more robust B cell response as compared to the pentamer of L1 alone, due in part to the ability of the HPV-VLP to activate the mechanosensitive membrane calcium channel PIEZO in contrast to the L1 pentamer that does not. These results reveal a property of the HPV-VLP that confers its efficacy in inducing LLPCs that could be applied to other subunit vaccines. Understanding the role of antigen-affinity thresholds in B cell memory. At the cellular level antibody memory is dependent on the acquisition of highly antigen-selected, high affinity long-lived plasma cells (LLPCs) specific for the immunizing antigen that take up residency in the bone marrow and secrete large amounts of antibodies that persist for years. Long-lived memory B cells (MBCs) are also acquired that upon antigen challenge predominantly undergo three fates: rapid differentiation to PCs that produce antibodies specific for heterologous variant antigens; differentiation toward germinal center (GC) B cells that potentially replenish the responding MBC population or induction of apoptosis. The human MBC compartment is composed of roughly equivalent numbers of switched IgG+ and unswitched IgM+ MBCs, however how these MBC subpopulations function in response to antigen challenge is incompletely understood. We discovered that intrinsic affinity thresholds for BCR-dependent antigen-activation are set at least 100-fold higher for IgG+ as compared to IgM+ MBCs independently of a number of cell surface markers that define additional heterogeneity in MBC subsets. Although IgG+ MBCs are unable to respond to low affinity antigens, challenge in vitro with high affinity antigens induced IgG+ MBCs to differentiate almost exclusively towards PC fates. In contrast, challenge with low affinity antigens induced IgM+ MBCs to predominantly differentiate towards GC B cell fates, whereas high affinity antigen induced apoptosis. We propose that human IgG+ and IgM+ MBCs populate two functionally distinct compartments in immune individuals that are restricted to responding to antigens in distinct affinity windows that result in PC fates for high affinity antigens and GC B cell fates for low affinity antigens. These findings have important implications for the design of vaccination strategies that can efficiently generate broadly neutralizing antibodies against infectious pathogens. Understanding the impact of the human HPV-VLP based vaccine, Gardasil, on the LLPC and MBC compartment Gardasil is a highly potent vaccine currently licensed for protection against infection with HPV, a highly carcinogenic virus. At present we know little about the outcome of any vaccine in use today in humans on the antigen-specific B cell compartment with the exception of recent studies on COVID vaccination. In collaboration with John Schiller (NCI) we are carrying out an in-depth analysis of peripheral blood HPV-specific B cells from a cohort of individuals who received the Gardasil vaccine. To do so we established a highly sensitive fluorescent HPV-VLP probe to identify and isolate HPV-specific B cells. Using this tool we determined that Gardasil induces a large expansion of IgG+ MBCs but not of IgM+ MBCs. The IgG+ MBC compartment is composed of at least six phenotypically defined subpopulations and unexpectedly the HPV-specific B cells induced by Gardasil contain numbers of ABCs and two additional MBC subpopulations. In addition we determined that the HPV-specific B cell population contained B cells expressed the inherently autoreactive VH4.34 genes suggesting that some degree of autoreactivity may be beneficial in response to Gardasil. We plan to continue our analyses of the HPV-specific B cells in vaccinated individuals using single cell RNA-seq, Ig-seq and cell surface phenotyping. We believe those analyses will provide insight into the cellular bases of the extraordinary efficacy of the Gardasil vaccine.