Immunology seeks to investigate the immune system’s response to pathogens and the diseases that result when the immune system is improperly regulated. B cells are immune cells that function to produce antibodies, proteins that target a specific molecule for destruction by the immune system. However, in autoimmune diseases, antibodies may target the host’s body. To avoid autoimmunity, B cells must have a carefully controlled threshold for activation. This threshold is set in part by the presence of activating and inhibitory receptors on B cells, which regulate signaling pathways for B cell activation. FCRL1 is a receptor found on B cells, and it has been shown to mediate both activating and inhibitory signaling pathways. Therefore, understanding how FCRL1 is involved in B cell signaling will advance what is known about the regulation of B cell activation, a topic with implications in autoimmune disease. We formerly identified that FCRL1 binds to the signaling proteins SHIP-1. Grb2. GRAP, and SOS and inhibits the MAP kinase pathway. In this investigation, we seek to identify the molecular basis of FCRL1’s signaling in B cells in order to determine the mechanism by which it inhibits MAP kinase signaling. We generated FCRL1 mutant proteins, which we analyzed by immunoprecipitation and immunoblotting to determine interaction sites with confirmed binding partners. Next, we employed the CRISPR/Cas9 system for gene editing to knock confirmed binding partners out of B cells. This allowed us to determine that multi-protein interactions are required for the recruitment of SHIP-1 to FCRL1. In the future, we plan to generate a series of fusion protein constructs that will allow us to determine which binding partners are sufficient to regulate the MAP kinase pathway, identify additional binding partners, and probe the role of FCRL1 in regulating levels of lipid messengers involved in B cell signaling.
Author: Maegan Murphy
Faculty Advisors: Timothy Wilson and Jenna DeLuca, Department of Microbiology
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