Investigating the differential role of t-cell receptor clonotypes in controlling hiv-1 infection utilizing lentiviral vectors expressing HIV-specific t-cell receptors
Thomas, Tynisha DeAndrea
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HIV elite controllers (EC's) are defined by their undetectable or low viral loads and the ability to maintain healthy CD4+ T cell levels. The association of certain HLA types and HIV disease progression is well documented but the mechanism by which HLA influences disease progression is not well understood. Studies have shown that immune control is seen in T cell receptor (TCR) clones isolated from HLA-B*27-infected individuals but the specific responsiveness and fine-tuning of the TCR interacting with the peptide/MHC complex remains unclear.;To gain insight into the mechanistic basis for differential cytotoxic T lymphocyte (CTL) anti-HIV function demonstrated in TCR clones, we analyzed T cell receptor function of multiple distinct HLA-B*2705-restricted CTL clones from HLA-B*27 elite controllers with divergent capacities to kill HIV-infected cells. We hypothesized that the differences in immune control from different HLA-B*27-KK10 specific clones is related to the TCR peptide/MHC complex.;We evaluated multiple HLA-B*2705 TCR clones from different elite controllers and demonstrated some elite controller TCR clones were reactive to wild type KK10 or KK10 variant peptide L6M while others demonstrated broad recognition. Differences in TCR-responsiveness correlated with cytotoxic differences seen in chromium release experiments and flow cytometric analysis of the mobilization of phosphorylated MAPK. We analyzed TCR functions by cloning the TCR alpha and beta chain genes from multiple KK10-specific CTL clones from elite controllers and cloned the sequences into a single lentiviral vector. Sequence analysis of the clones revealed multiple KK10 TCRs with varying alpha and beta combinations and completely different CDR3 regions. We analyzed TCR function by examining TCR signal transduction by transducing Jurkat/MA cells and demonstrated modest differences in TCR reactivity to wild-type KK10 and viral variants of this peptide.;Successful transduction and tetramer staining of the transduced CTLs demonstrated that naive cells can be molecularly engineered to be HIV-specific CTLs. Transduced CTL differences in viral inhibition were not observed when co cultured with infected cells indicating other mechanistic factors work in concert with the TCR to harness antiviral immunity. These studies should increase our understanding of the role of HLA and the mechanisms that confer immune protection.