Rotenone-mediated mitochondrial ROS generation inhibits melanogenesis in B16F10 cells by inducing the ERK activation-MITF degradation pathway

Pachyrhizus erosus seeds have been reported to have various biological activities, including antifungal, antisecretory, insecticidal, antibacterial, and antispasmodic properties. In this study, we evaluated the hypopigmentation effects of the ethanol extract of Pachyrhizus erosus seeds (PESE), identified rotenone as a representative active metabolite, and proposed a mechanism for inhibiting α-MSH-mediated melanogenesis in B16F10 cells. PESE treatment effectively inhibited melanin synthesis in B16F10 cells stimulated with α-MSH or forskolin. Among the three major metabolites characterized from PESE, pachyrrhizine, neotenone, and rotenone, only rotenone exhibited a strong inhibitory effect on melanin synthesis at a concentration of 8 nM, with minimal cytotoxicity. Rotenone suppressed transcriptional expression of melanosomal genes, TRP-1 and TYR, in B16F10 cells stimulated by α-MSH, primarily due to a reduction in the protein level of microphthalmia-associated transcription factor (MITF). Rotenone, an inhibitor of mitochondrial electron transport chain complex I, induced mitochondrial reactive oxygen species (ROS) production, and the increased ROS activated ERK. Treatment with N-acetylcystein (NAC), a ROS scavenger, or PD98059, an ERK inhibitor, suppressed the decrease in MITF protein induced by rotenone, thereby eliminating the hypopigmentation effect of rotenone. These findings provide novel insights into the whitening activity mechanism of rotenone and suggest that mitochondrial damage may affect melanogenesis.