We are investigating the mechanism of cell transformation by Rous sarcoma virus (RSV) and how the events in these cells relate to those induced by other avian sarcome viruses. The relationship between protein phosphorylation and viral transformation will also be compared to theprotein phosphorylation events associated with the response of cells to epidermal growth factor (EGF). The work proposed includes 1) biochemical characterization of the Rous sarcoma virus transforming gene product, pp60(src), and its normal cell homologue, pp60(c-src), as protein kinases as well as a search for other biochemical functions; 2) characterication of these proteins as phosphoproteins and how they intereact with other cellular protein kinases; 3) comparative functional analysis of pp60(src) and pp60(c-src); 4) identification and characterization of pp60(src)/pp60(c-src) substrates; and 5) biochemical characterization of the protein kinase activity associated with other viral trnasforming gene products and with the response of cells to EGF. These experiments bear directly on the health-related problem of oncongenesis. We shall utilize a) ion-exchange and immunoaffinity techniques for protein purification, b) molecular cloning and DNA transfection techniques, c) two-dimensional and conventional slab gel polyacrylamide gel electrophoresis of proteins, d) immunoprecipitation, and e) conditional and non-conditional mutants of RSV.
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