Molecular Basis for Immunogenic B cell responses to Virus-Like Antigen Display

About This Grant

PROJECT SUMMARY Self-reactive B cells must distinguish self from foreign antigens (Ag), mounting immunogenic responses to the latter but not the former. However, the mechanisms they employ to do so are not fully understood. Long-term, we aim to exploit new molecular insight gained by filling this knowledge gap to restore B cell tolerance in autoimmunity, enhance anti-tumor B cell responses to self-like Ag, and optimize host defense. In this proposal, we leverage a unique and modular platform of liposomes decorated with model Ag, enabling independent control of epitope affinity and density. Using this system, we recently demonstrated that such particulate Ag are much more potent activators of B cells than identical Ag in soluble, monovalent form. We found that this stems not merely from avidity but from the ability of particulate Ag to evade inhibitory signaling pathways normally engaged by soluble Ag and mediated by the Src family tyrosine kinase Lyn. Moreover, we discovered that transcriptional programs and B cell functional responses to particulate and soluble Ag diverge markedly; despite an identical epitope recognized by the BCR, we found that particulate Ag produce robust NF- kB activation even in the absence of T cell help, while soluble Ag drive an NFAT-associated anergy program. We propose that biophysical characteristics of particulate Ag display serves as a stand-alone danger signal that evades tolerogenic transcriptional programs and elicits immunogenic responses by B cells. In this grant, we propose to: (1) Identify proximal biochemical pathways downstream of the Lyn Src family tyrosine kinase that enable B cells to distinguish tolerogenic and immunogenic Ag display. We will edit mouse and human B cells in vitro and in vivo to test candidate substrates of Lyn. We will complement this with an unbiased proteomics approach to identify novel negative regulators of B cell responses to self-like soluble Ag. (2) Elucidate how early signaling events triggered by particulate Ag are transformed into immunogenic transcriptional programming of B cell fate through robust activation of NF-kB but not NFAT pathways. We will take a genetic approach to manipulate the second messenger diacylglycerol (DAG) which we hypothesize toggles between these immunogenic and tolerogenic B cell transcriptional responses. We will test this in human B cells. We will pursue a genomic strategy to survey the epigenetic landscape of B cells in order to define the transcriptional architecture that translates pattern of Ag display into B cell fate. (3) Systematically map the lower boundary of Ag affinity, density, and particle dose, and test the biophysical properties (particle size, membrane fluidity, linker length) required to activate B cells, including anergic self- reactive cells. We will test the hypothesis that these thresholds are controlled by Lyn-dependent inhibitory signaling pathways.