Human T cells display a specific receptor (E) to sheep erythrocytes, causing the formation of spontaneous rosettes. Accumulating data suggest that the E-receptor and a p50 cell surface protein associated with it plays a regulatory role of multiple T cell functions that include the release of lymphokine(s) necessary for lymphocyte growth, cytotoxic T cell function and natural killer cell lysis. The long-term objectives of this proposal aim toward increasing our understanding of the biology of the p50 system. In particular we will test the hypothesis in which we propose that the E-receptor and p50 act as a modulator of both cellular and humoral immune reactions. Defective T cell E-rosette formation is observed in association with several diseases including cancer, autoimmune diseases and viral infections. This phenomenon appears to be mediated by serum factors that affect E-rosette formation and T cell function in a similar way than anti-p50 monoclonal antibody. A special emphasis will be given to experiments designed to analyze the mechanism by which E-receptor and p50 regulate T cell activation and the release of lymphokine(s) such as interleukin 2(IL-2). The effect of anti-p50 mAb will be analyzed using normal purified T cells, T cell subsets and various T cell clones. These experiments will include: (a) Kinetics of biosynthesis, maturation and release of p50 molecules of normal and mitogen-induced T cells using pulsechase labeling experiments and SDS gel electrophoresis of immunoprecipitated radio-labeled p50 proteins. (b) Cytofluometric analysis of the cell cycle of mitogen induced T cells, in particular the effect of anti-p50 on the G0-G1 transition will be examined. (c) Special emphasis will be given to examine the role of p50 in regulating IL-2 pathway: expression of IL-2 receptor (Tac antigen), release of IL-2 and more importantly the expression of IL-2 mRNA using cloned IL-2 cDNA probes. (d) In addition the role of p50 in regulating the release of lymphokine necessary for B cell growth, and immunoglobulin secretion will be examined using a B cell proliferation assay and an immunoglobulin ELISA assay respectively. (e) Finally, considerable effort will be made to clone p50 cDNA probes using synthetic oligonucleotide probes prepared on the basis of amino acid sequence analysis of purified p50 polypeptide. p50 cDNA probes will then be used to study the structure, function and expression of p50 gene products and its role in regulating T cell activation and lymphokine(s) productions.
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