The spectrum of autoimmune and rheumatic diseases spans a variety of defects in the immune response. However, to appreciate the underlying pathophysiology of these diseases and thus be able to develop therapeutics in a rational fashion, the biochemical basis of the immune response must be understood. The T cell represents one of the principal effectors and regulators of the immune system. Its role in host defense includes killing of virally infected cells (cytotoxicity), immunoregulation of other cells involved in the immune response (e.g., B cell-T cell interactions), and the ability to secrete lymphokines to effect a variety of tissues and cells. Activation of T cells by foreign pathogens resulting in these effector functions is mediated by the T cell antigen receptor (TCR). Over the past decade, the TCR has been defined biochemically and its structural requirements for normal assembly and function studied in great detail. However, the mechanism by which TCR-mediated signals are transmitted into effector functions remain poorly defined. A variety of intracellular biochemical signals have been implicated, including protein phosphorylation via both tyrosine and serine/threonine protein kinases, dephosphorylation by phosphatases, and mobilization of inositol phosphate with resultant increases in cytoplasmic calcium levels. However, the interplay of these biochemical processes are not understood.
The aim of this proposal is to dissect and define the cascade of signals involved in TCR activation. Irving and Weiss have recently demonstrated the importance of the cytoplasmic domain of the zeta-chain in TCR signal transduction. Furthermore, we have also recently described a protein, ZAP- 70, that associates with zeta and undergoes tyrosine phosphorylation after stimulation of the TCR. This proposal utilizes a combination of biochemical, molecular, and immunologic approaches to characterize ZAP-70. Specifically, these include: l. purification of ZAP-70 for protein microsequencing, 2. isolation of cDNA clones encoding for ZAP-70 utilizing degenerate oligonucleotides generated from protein sequence, and 3. generation of antibodies against ZAP-70. Together, the deduced sequence of ZAP-70 may provide insight into its function based on sequence homology. Moreover, these structural studies and availability of reagents against ZAP-70 will permit further biologic and functional investigation. These studies will allow for a greater understanding of the activation of T cells by foreign pathogens.
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