We have explored the role of protein kinase C (PKC) in pigmentation induced by alpha-melanocyte stimulating hormone (alpha-MSH). Using the well-studied S91 Cloudman mouse melanoma model system in which 10(-7) M alpha-MSH is known to produce a time-dependent increase in pigmentation, we found an increase in the activity of tyrosinase, the key enzyme in pigmentation, between Days 2 and 6 accompanied by an increase in mRNA and protein levels of tyrosinase, as well as an increase in the level of specifically the beta isoform of PKC. When S91 cells were treated with phorbol dibutyrate, 95% of PKC activity was lost within 48 h and the alpha-MSH-induced melanogenesis was completely blocked, as was the induction of tyrosinase mRNA and protein. Serially passaged S91 cells no longer capable of responding to alpha-MSH had an undetectable level of PKC-beta, although the tyrosinase protein level was identical to that of alpha-MSH-responsive cells. Furthermore, in these S91 cells alpha-MSH also did not increase the level of tyrosinase mRNA. Thus, induction of murine melanogenesis by alpha-MSH involves up-regulation of tyrosinase mRNA and protein mediated in part by the PKC-dependent pathway, associated with an up-regulation of the beta isoform previously demonstrated to specifically activate tyrosinase in human melanocytes.