Studies of NAD+ - mediated melanogenesis.

Abstract

The melanogenesis pathway in mammalian models has been extensively characterized. However, no study has yet to show the involvement of NAD+ in melanogenesis. Our present study showed that melanin content and tyrosinase enzyme activity in B16 melanoma cells and human primary melanocytes were significantly increased by extracellular b-NAD+ with no observed cytotoxicity to the cells. The b-NAD+ stimulated increase of melanin content was blocked by H89, an inhibitor of PKA. In B16 melanoma cells treated with extracellular b-NAD+, there was a marked increase in the protein levels of tyrosinase and MITF, which were mediated via the phosphorylation of CREB. In addition, transcripts of expected size for P2 receptor isoforms - P2Y1, P2Y2, P2Y6 and P2Y12 were found to be expressed in B16 melanoma cells. The nonselective P2 receptor antagonist, PPADS displayed significant antagonizing effects on the stimulated enzyme activity and increased level of tyrosinase resulting from the addition of extracellular b-NAD+. These data suggests the presence of a novel melanogenesis pathway in melanocytes, which is activated/stimulated by b-NAD+. In this pathway, the regulation of tyrosinase gene expression and melanin level by b-NAD+ is mediated via the activation of PKA, the phosphorylation of CREB and followed by the upregulation of MITF.