We propose to continue our investigation of the role of pp60c-src, the cellular homolog of pp60v-src, the transforming protein of Rous sarcoma virus, in cellular growth control to determine what functional differences between the two proteins are responsible for v-src induced neoplastic transformation. We will compare pp60c-src, pp60v-src, and pp60recombinant-src (pp60r-src) activities in vivo using the c-, v- and r-src expression plasmids and overexpressor cell lines that we have created and will use immunopurified pp60src from these overexpressor cell lines for in vitro studies. We will determine if the low-frequency focus formation induced by extremely efficient c-src overexpression plasmids is due to mutation or represents an activity of authentic pp60c-src. We will see if the defect in pp60c-src transforming activity can be activated or complemented in tissue culture by other transforming proteins, tumor promoters or hormones and will investigate the biological effects of pp60c-src overexpression in non-immortalized cells. We will investigate the dose-response behaviors of cells to pp60c-src, pp60v-src and pp60r-src overexpression to compare the behaviors of these proteins in vivo. We will complete the characterization of the activities of c-src - v-src chimeric genes and will determine what biochemical differences are associated with the different biological phenotypes induced by these genes. We intend to identify specific functional difference(s) between pp60c-src and pp60v-src which are responsible for their different transforming activities by comparing their in vitro and in vivo phosphorylating activities. In particular, we will investigate the significance of pp60c-src phosphorylation of hexokinase.
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