Retinoic acid (RA) has been shown to inhibit melanogenesis in B16 mouse melanoma cells (B16 cells). On the other hand, it has been reported that RA increases protein kinase C (PKC) activity in these cells. Further investigation was carried out to identify the PKC subspecies expressed in B16 cells and to examine the changes in the level of each PKC subspecies by RA treatment. Hydroxyapatite column chromatography, immunoblot analysis, and kinetic analysis have shown that B16 cells express the alpha subspecies of PKC. Northern blot analysis has indicated that these cells normally express mRNA for the alpha, delta, epsilon, and zeta subspecies. Upon treatment of B16 cells with 1 microM RA for 48 h, both the activity of the alpha-subspecies and the level of mRNA for the alpha subspecies were increased, resulting in the decrease of melanin polymer formation and tyrosinase activity. Neither the enzyme activities nor mRNA for the beta and gamma subspecies were detected in either the RA-treated or untreated cells. The levels of mRNA for the delta, epsilon, and zeta subspecies were not altered by RA treatment. The demonstration of a selective increase of the alpha subspecies of PKC is a unique finding.