CAMP RESPONSE ELEMENT OF MURINE CYTOMEGALOVIRUS IMMEDIATE-EARLY GENE ENHANCER IS TRANSACTIVATED BY RAS ONCOGENE PRODUCTS

Products of ras oncogenes strongly stimulate the activity of the repor ter gene, chloramphenicol acetyltransferase (CAT), driven by a 1 . 2kb fragment of the murine cytomegalovirus (MCMV) immediate early (IE) ge ne enhancer (pCMVCAT). To define the role of proteins binding to the u nique cAMP response element (CRE) present in the IE enhancer, NIH 3T3 cells were cotransfected with prasZip6 plasmid, a mammalian expression vector containing a v-Ha-ras cDNA, together with P Delta ACMVCAT (pCM VCAT without the CRE sequence). Lower stimulation of CAT activity was indeed observed upon deletion of the CRE sequence. Decreased levels of p Delta ACMVCAT were also observed in cell lines carrying stably tran sfected ras oncogenes. Further support for the role of the CRE sequenc e in MCMV enhancer activation comes from the finding that v-Ha-ras exp ression increases the activity of a reporter gene, beta-galactosidase, driven by three tandem copies of CRE sequence about six-fold. Moreove r, this transactivation was prevented by cotransfection of the dominan t inhibitor mutant Ha-ras (Leu-61; Ser-186) and was not suppressed by cotransfection of Ha-vas (Asn-17), suggesting that the effect is due t o activated ras protein, rather than normal p21(ras). Finally the tran sactivation observed is accompanied by an increase in nuclear proteins binding to a labelled oligonucleotide homologous to the CRE sequence, as shown in a gel retardation assay. These results suggest that the C RE element contributes to the transactivation of the MCMV IE gene enha ncer by vas oncogenes.