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Date of Award

9-2013

Document Type

Campus Access

Degree Name

Doctor of Philosophy (PhD)

Degree Program

Molecular and Cellular Biology

First Advisor

Janice Telfer

Second Advisor

Cynthia Baldwin

Third Advisor

Wilmore Webley

Subject Categories

Immunology and Infectious Disease

Abstract

WC1 proteins specifically expressed on the surface of γδ T cells are members of group B Scavenger Receptor Cysteine Rich (SRCR) superfamily, in which receptors contain several SRCR domains in the extracellular region. WC1+ γδ T cells play a critical role in bridging innate and adaptive immunity, organizing granulomas in response to Mycobacterium and producing IFNγ in response to Leptospira . The serologically-defined WC1.1+ γδ T cells exclusively respond to spirochete Leptospira ; the serologically-defined WC1.2+ subpopulation responds to rickettsias Anaplasma ; shRNA silencing three WC1.1+ proteins significantly reduced γδ T cell response toLeptospira antigen. Co-ligation of WC1 with TCR/CD3 potentiates T cell activation, and tyrosine phosphorylation of the WC1 cytoplasmic domain is required for WC1 co-receptor activity. We hypothesized that WC1 receptors encode antigen specificity and contribute to T cell activation in response to leptospirosis. Our data showed SRCR domains from a WC1.1 type receptor, WC1-3, directly interact with vaccines and liquid cultures of leptospires. Vaccine Leptospira interacts with more SRCR domains as compared to liquid cultures, suggesting that vaccine preparation may enhance ligand accessibility to SRCR domains. Importantly, we did not observe any Leptospira binding from WC1-4 SRCR domains, a representative WC1.2 type receptor. The binding assay showed that SRCR a1 domains from WC1.1 type proteins contribute to Leptospira recognition, but none from WC1.2 proteins does. Alkaline phosphatase treatment suggests that a phosphorylation pattern is recognized by SRCR a1 domain, supporting that WC1 functions as a pattern recognition receptor. PMA-induced CD4/WC1-3 endocytosis is mediated by a membrane-proximal dileucine motif, which is essential for the recruitment of AP2 complexes. The disruption of the dileucine motif greatly accumulates the overall levels of CD4/WC1-3 molecules on the cell surface and in the cytoplasm. Co-crosslinking CD4/WC1-3 and TCR/CD3 indicates that the dileucine motif acts as a negative regulator for downstream cytokine production. Moreover, a double serine motif upstream the dileucine motif mediates signaling through WC1-3 for the downstream event. Taken together, the data support that co-ligation of WC1 and the γδ TCR by pathogen-associated molecular patterns (PAMPs) induces specific γδ T cell activation.

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