Human leukocyte antigen (HLA)-A, B and C class I molecules are expressed on the surface of almost all nucleated cells and present endogenously processed peptide antigen to cytotoxic T lymphocytes. This process forms the basis by which the immune system eliminates virally infected or malignantly transformed cells. HLA-A and B proteins have well characterized structure and function, but HLA-C proteins are poorly characterized. Protein products of the HLA-C class I genes are expressed at low levels and this has been shown to correlate to low HLA-C mRNA abundance caused by rapid mRNA turnover. The protein products of several gene families, including cytokines and oncogenes, are deleterious to the host when over-expressed. These genes maintain low protein expression by destabilizing mRNA. HLA-C MRNA has a half-life similar to cytokine and oncogene genes. These data suggest that over-expression of HLA-C protein products may be discerned by understanding the mRNA regulatory mechanisms.
The specific aims described in this proposal are to determine how cis and trans acting elements regulate HLA-C mRNA stability, and identify locus specific nucleic acid residues that form a regulatory element. Cis acting elements will be detected by generating chimeric reporter plasmid constructs, HLA-B/C genes, site directed mutants and RNase H analysis. Trans acting elements will be assessed using gel mobility shift assays and reporter constructs to test the role for translation. A database of HLA-B and C nucleic acid sequences will be generated to determine locus specific nucleic acid residues; appropriate sequences will be compared to other genes. The immediate goal of these studies is to determine mechanisms associated with HLA mRNA regulation. The long range goals of these studies will be to determine the biological significance of low HLA-C cell surface expression and examine the effects of alteration of HLA-C surface expression levels. These studies will increase our understanding of immune regulation.
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