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Document Type

Campus-Only Access for Five (5) Years

Degree Name

Doctor of Philosophy (PhD)

Degree Program

Molecular and Cellular Biology

Year Degree Awarded

2018

Month Degree Awarded

May

First Advisor

Cynthia L. Baldwin

Second Advisor

Janice C. Telfer

Third Advisor

Lisa M. Minter

Fourth Advisor

Peter Chien

Subject Categories

Immunology of Infectious Disease

Abstract

WC1 molecules are hybrid PRR/co-receptors exclusively expressed on gd T cells of ruminants while humans and mice have related receptors. Bovine WC1 molecules bind pathogens directly and specifically and augment TCR-mediated signals. Using NGS, we found all 13 WC1 genes transcribed by blood gd T cells with no significant differences at birth although by adulthood the relative proportions of transcripts was somewhat altered perhaps attributable to immunological experience. WC1+ gd T cells form two main subpopulations in blood, WC1.1 and WC1.2, distinguished by reactivity with various anti-WC1 mAbs. Their WC1 molecules also differ in their ability to bind particular microbes suggesting the WC1 molecules expressed by a gd T cell may determine its antigen specificity. Although it was clear that the WC1+ subpopulations differed in WC1 gene expression we did not know the expression of WC1 genes by individual gd T cells. Thus 78 gd T cell clones from flow-cytometrically purified WC1+ subpopulations were generated and evaluated by qRT-PCR. The majority of gd T cell clones transcribed 1 to 4 WC1 genes and, while overlap of some transcripts occurred between the two subpopulations, 80% of clones that were generated in response to Leptospira stimulation had transcripts for a WC1 molecule known to bind Leptopsira which was significantly higher than for non-responsive clones We found WC1 molecules are not detected on thymocytes prior to the gd TCR and that the two WC1+ subpopulations are at a similar ratio in thymus as in blood. Using ChIP, we showed that Sox13, an important transcription factor for gd T cell development, differentially occupied WC1 gene loci in the WC1.1+ versus WC1.2+ cells. Using qRT-PCR and immunofluorescence, we evaluated transcription factors critical to development and effector functions of T lymphocytes as well as cytokines and the gd T cell subset marker CD27 in the two subpopulations. WC1.1+ cells showed significantly higher levels of transcription for CD27, as well as higher transcription and protein for Tbx21 and IFNg than WC1.2+ cells; the latter had higher levels of transcripts for Rorc and Il17a. This suggests a tendency for a Tgd1-like phenotype versus Tgd17-like phenotype, respectively.

Available for download on Saturday, May 11, 2019

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