Oncogene activation has been suggested to play some role in determining the hormone independency of tumors. In order to study the role of protein kinase C in mediating the inhibition of the glucocorticoid-dependent transcription from the Mouse Mammary Tumor Virus (MMTV)-Long Terminal Repeat induced by overexpressed activated ras oncogene, we studied the effects of protein kinase C activators [the tumor promoting phorbol ester 12-O-tetradecanoyl-phorbol-13-acetate (TPA)] and inhibitors [1-(5-isoquinolinesulfonyl)-2-methylpiperazine (H-7)] on the dexamethasone (DEX)-mediated activation of a MMTV-Long Terminal Repeat-chloramphenicol acetyltransferase (pMMTV-CAT) chimeric reporter gene transiently transfected into NIH-3T3 cells and in Ha-ras-transformed fibroblasts (T24-NIH-3T3). TPA (30 ng/ml) together with DEX (0.1 microM) treatment of NIH-3T3 cells resulted in a significant decrease of CAT activity from pMMTV-CAT, compared to DEX treatment alone. The addition of H-7 (40 microM) was able to overcome the TPA-induced inhibition of DEX-dependent transcription from pMMTV-CAT. DEX-dependent expression of pMMTV-CAT was significantly reduced in T24-NIH-3T3 with respect to wild-type NIH-3T3 cells. Treatment of T24-NIH-3T3 cells with either H-7 or TPA significantly enhanced or decreased, respectively, the DEX-dependent expression of pMMTV-CAT. TPA and/or H-7 did not affect CAT activity from either pMMTV-CAT in the absence of DEX or from CAT gene under the control of the SV40 promoter. Similar glucocorticoid receptor sites and binding affinities were observed in T24-NIH-3T3 or TPA-treated NIH-3T3 cells compared to wild-type untreated cells. Our data suggest that activation of PKC is involved in the reduced transcriptional regulatory activity of glucocorticoid hormone induced by overexpressed Ha-ras oncogene in NIH-3T3 fibroblasts.