Evidence for two pathways of protein kinase C induction of 2ar expression: correlation with mitogenesis

2ar is a cDNA clone of an mRNA that is inducible by 12-0-tetradecanoylphorbol-13-acetate (TPA) in mouse JB6 epidermal cells. This mRNA is stably induced in confluent cells but only transiently induced in subconfluent cells; the elevated level of 2ar mRNA in TPA-treated cells appears to be the result of enhanced transcription (Smith and Denhardt: Journal of Cellular Biochemistry 34:13-22, 1987). Phorbol dibutyrate, teleocidin, and aplysiatoxin, which activate protein kinase C and are also tumor promoters, are shown here to induce 2ar to the same extent as TPA at both cell densities. The increased expression by TPA was prevented by cycloheximide and by H7, a specific inhibitor of protein kinase C. Epidermal growth factor, platelet-derived growth factor, and the nonphorbol promoter diethylhexylphthalate were more effective inducers in confluent than in subconfluent cultures. All-trans retinoic acid, dexamethasone and fluocinolone acetonide, inhibitors of tumor promotion, diminished 2ar induction in both acetonide, inhibitors of tumor promotion, diminished 2ar induction in both confluent and subconfluent cells. In TPA-treated subconfluent cultures indomethacin produced a slight inhibition, whereas difluoromethyl ornithine potentiated the induction. In TPA-treated confluent cultures, in contrast, indomethacin enhanced 2ar mRNA levels and difluoromethyl ornithine was inhibitory. We conclude that the protein kinase C-mediated induction of 2ar expression is controlled by different pathways in subconfluent cultures and confluent cultures, indicative of apparent changes in the regulation of gene expression as proliferating cells become quiescent.