Oncogenes are an activated version of cellular proto-oncogenes that control cell growth and differentiation. It is believed that these genes participate in signaling pathways. We have identified a dominant, negative ras mutant capable of inhibiting c-ras proto-oncogene function (Nll6Y mutation of v-H-ras). This mutation is present in one of the consensus sequences conserved in the GTP-binding proteins and is critical in the interaction of ras proteins with the GTP or GDP nucleotides. The N116Y mutant blocks transformation by retroviruses carrying protein tyrosine kinase oncogenes, including v-abl, v-fes/flp and v-fms, but did not affect v-H-ras, v-K-ras and v-bas (for details see Project No. ZOICPO5594-03). We have established a series of NIH3T3 cell lines to investigate the biochemical target and signaling pathways mediated by ras. In this study we have found that phosphorylation of a protein kinase C (PKC) substrate (80 kD protein) was lower in cells transfected with the N116Y ras mutant than in normal NIH3T3 cells. Expression of the N116Y ras mutant completely blocked DNA synthesis induced by EGF, but not by other growth factors or serum. Sequestration or inhibition of ras targets could be an explanation for these differences.