No abstract

Tyrosinase catalyzes the rate‐limiting step in melanin synthesis. Melanin is synthesized from l‐tyrosin in the melanosomes, where tyrosinase and other melanogenic factors are recruited via the vesicle transport system. Genetic and biochemical approaches have revealed a correlation between impairments in the vesicle transport system and albinism. However, the specificity of the individual transport systems for the corresponding melanogenic factors has not been well elucidated yet. Here, we report that the thioxothiazolidin derivative, 4‐OST (4‐[(5E)‐5‐[(4‐fluorophenyl)methylidene]‐4‐oxo‐2‐sulfanylidene‐1,3‐thiazolidin‐3‐yl]‐4‐azatricyclo [5.2.1.02,6]dec‐8‐ene‐3,5‐dione: CAS RN. 477766‐87‐3) strongly inhibited melanogenesis in mouse melanoma B16F10 cells. 4‐OST reduces tyrosinase protein levels without affecting its messenger RNA levels or enzymatic activity. Although a reduction in tyrosinase protein level was observed in the presence of a protein synthesis inhibitor, the reduction may be coupled with protein synthesis. Similarly, GIF‐2202 (a derivative of 4‐OST) lowers tyrosinase protein levels without affecting the levels of another melanogenic enzyme, tyrosinase‐related protein 1 (TYRP1) level. The reduction in tyrosinase protein level is associated with an increase in the levels of the lysosomal proteinase cathepsin S. Chloroquine, a lysosome inhibitor, restored the tyrosinase protein level downregulated by GIF‐2202, although no effects of other inhibitors (against proteasome, autophagy, or exocytosis) were observed. In addition, GIF‐2202 segregated the immunofluorescence signals of tyrosinase from those of TYRP1. Chloroquine treatment resulted in co‐localization of tyrosinase and cathepsin S signals near the perinuclear region, suggesting that 4‐OST and GIF‐2202 may alter the destination of the tyrosinase vesicle from the melanosome to the lysosome. 4‐OST and GIF‐2202 can be new tools for studying the tyrosinase‐specific vesicle transport system.

No abstract

Melanogenesis and melanosome secretion are regulated by several mechanisms. In this study, we found that the oxindole derivative GIF-2209 accelerated melanogenesis associated with the discrimination in the expression and intracellular distributions of two melanogenic enzymes, tyrosinase (TYR) and tyrosinase-related protein-1 (TYRP-1). GIF-2209 upregulated the expression of TYR via a microphthalmia transcription factor (MITF)-independent mechanism, leading to high expression of protein. In contrast, GIF-2209 did not alter the mRNA levels of TYRP-1 and suppressed its protein levels. GIF-2209 induced the dissociation of TYR from TYRP-1 but did not alter the association between TYR and CD63, a melanosome and lysosome marker. The protein levels of CD63 were also upregulated by GIF-2209. GIF-2209 induced lysosome expansion and redistribution in all areas of the cytosol, accompanied by autophagy acceleration (upregulation of LC3BII protein levels and downregulation of p62 protein levels). In addition, GIF-2209 stimulated the secretion of melanosomes containing high levels of TYR, TYRP-1, and CD63 proteins. The GIF-2209 mediated melanosome secretion was sensitive to the lysosome inhibitor chloroquine. These results suggest that GIF-2209 may activate lysosomal functions with TYR gene expression, while it accelerates melanosome secretion, which finally leads to the depletion of intracellular melanogenic enzyme, especially TYRP-1 protein.