RAS-INDUCED C-FOS EXPRESSION AND PROLIFERATION IN LIVING RAT FIBROBLASTS INVOLVES C-KINASE ACTIVATION AND THE SERUM RESPONSE ELEMENT PATHWAY

We have examined the early events involved in the proliferative activation of quiescent rat embryo fibroblasts by microinjection of oncogenic ras protein. Cells injected with ras show a transient expression of c-fos after 30-60 min visualized by immunofluorescence in the nucleus. This c-fos expression can be specifically suppressed by coinjection of a double-stranded oligonucleotide which corresponds to the serum response element (SRE) present in the c-fos promoter, implying that ras utilizes a pathway which activates the binding of serum response factor(s) (SRF) to SRE to induce c-fos transcription. Inhibition of this pathway also abolished ras-induced DNA synthesis indicating that the proliferative induction by ras requires expression of SRE-regulated genes. Both c-fos induction and DNA synthesis were prevented when ras oncoprotein was injected into quiescent cells together with either antibodies against calcium phospholipid-dependent protein kinase (C-kinase) or a synthetic peptide that specifically inhibits C-kinase. These data demonstrate the involvement of both functional C-kinase and the SRE pathway in the activation of quiescent cells by ras and suggest a potential relationship in their mechanism of action.