Regulation of melanogenesis in B16 mouse melanoma cells by protein kinase C.
Melanogenesis is regulated by a variety of environmental and hormonal factors. In this study, we showed that protein kinase C (PKC) plays a major role in regulating melanogenesis in B16 mouse melanoma cells. Chronic treatment of B16 cells with phorbol dibutyrate resulted in a concentration-dependent loss of density-dependent induction of tyrosinase activity, which correlated positively with a concentration-dependent loss of PKC enzyme activity. In contrast, B16 clones overexpressing PKC alpha had increased tyrosinase activity. Different phorbol derivatives inhibited tyrosinase activity and depleted cellular PKC alpha in a manner that reflected their reported tumor-promoting activity. Western blotting analysis showed that phorbol dibutyrate decreased the amount of the brown locus gene product (TRP-1) by 50% and lowered the amount of the albino locus gene product (tyrosinase) to undetectable levels. None of the phorbol derivatives affected the level of the slaty locus protein (TRP-2). The decrease in tyrosinase and TRP-1 protein levels was found to be due to a decrease in the mRNA encoded by these genes. In addition to inhibiting the density-dependent increase in tyrosinase activity, phorbol dibutyrate inhibited some, but not all, of the 8-bromocyclic AMP-induced increase in tyrosinase activity. This was accompanied by a decrease in the amount of tyrosinase protein induced by 8-bromocyclic AMP. Although 8-bromocyclic AMP did not change the level of TRP-1, it did reverse the decrease in the amount of this protein induced by phorbol dibutyrate. The amount of TRP-2 was not altered by any of these agents. These data suggest that PKC regulates melanogenesis primarily by controlling the constitutive expression of tyrosinase and, to a lesser extent, TRP-1.
Mahalingam
H
H
Department of Biochemistry and Molecular Biology, Marshall University School of Medicine, Huntington, West Virginia 25755, USA.
Vaughn
J
J
Novotny
J
J
Gruber
J R
JR
Niles
R M
RM
D000818:Animals; D000242:Cyclic AMP / Q000502:physiology; D015870:Gene Expression; D008543:Melanins / Q000096:biosynthesis*; D008546:Melanoma, Experimental / Q000378:metabolism*; D008562:Membrane Glycoproteins; D051379:Mice; D014442:Monophenol Monooxygenase / Q000378:metabolism*; D010088:Oxidoreductases; D015240:Phorbol 12,13-Dibutyrate / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D014162:Transfection; D014407:Tumor Cells, Cultured
D008543:Melanins; D008562:Membrane Glycoproteins; D011506:Proteins; D012333:RNA, Messenger; D015240:Phorbol 12,13-Dibutyrate; D000242:Cyclic AMP; D010088:Oxidoreductases; C079621:Tyrp1 protein, mouse; C064056:tyrosinase-related protein-1; D014442:Monophenol Monooxygenase; D011493:Protein Kinase C