We have investigated the role of protein kinase C (PKC) isoenzymes in the differential growth regulation of human pancreatic carcinoma cell lines by all-trans retinoic acid (RA). RA treatment results in dose-dependent stimulation of anchorage-independent growth in AsPc1 cells and growth inhibition in Capan 2 cells. Both cell lines express an identical pattern of nuclear RA and retinoid X receptors as determined by RT-PCR. Western blotting using monospecific antibodies revealed that both cell lines express PKC isoenzymes alpha and zeta, whereas beta, gamma, delta, and epsilon were not detected. Incubation with RA in the growth-stimulated AsPc1 cell line resulted in induction of PKC alpha expression, whereas PKC alpha expression was decreased by RA in the growth-inhibited Capan 2 cell line. In contrast, PKC zeta expression was not affected by RA in either cell line. Incubation of AsPc1 cells with the phorbol ester 12-O-tetradecanoyl phorbol 13-acetate resulted in a time- and dose-dependent selective down-regulation of PKC alpha but not zeta. The dose-dependent decrease of intracellular PKC alpha concentration correlated well with the anchorage-independent growth rate of AsPc1 cells. Furthermore, selective down-regulation of PKC alpha blocks subsequent growth stimulation by RA in AsPc1 cells. When PKC alpha concentration was decreased by stably transfecting AsPc1 cells with a PKC alpha complementary DNA antisense construct, RA-stimulated growth could also be partially blocked. These data, therefore, suggest that differential regulation of PKC alpha expression plays a central role in determining the bidirectional effects of RA on growth in pancreatic carcinoma cells.