Co-regulation of melanin precursors and tyrosinase in human pigment cells:Roles of cysteine and glutathione

Glutathione (GSH) and cysteine (CysH) have both been implicated in the biog enesis of the pheomelanin precursor 5-S-cysteinyldopa (5-S-CD), However, re cent studies have shown that only CysH is transported across the membrane o f isolated melanosomes, and that the positive regulation of CysH in pigment cells leads to an increased production of 5-S-CD, In the present study, th e question was examined as to whether melanin precursors and tyrosinase cou ld be coregulated by cellular thiols, To address this issue, the levels of CysH and GSH were varied in normal melanocytes and melanoma cells using but hionine sulfoximine (BSO), an inhibitor of GSH biosynthesis, Treatment with 50-100 mu M BSO decreased GSH levels to less than 10% of control, and incr eased CysH levels between two- and five-fold in both cell types. Concomitan t with this, an increase in the ratio of 5-S-CD to DOPA and a decrease in t he pigment content of the cells were observed. The decrease in cell pigment ation was associated with strong decreases in tyrosine hydroxylase activity and C-14-melanin production. Only melanoma cells showed a modified tyrosin ase isozyme pattern on Western immunoblots in response to BSO, while the mR NA expression of tyrosinase and TRP-1 were unchanged in both cell types. Th ese results suggest that the balance between CysH and GSH, which is partly determined by the rate of utilization of CysH for GSH biosynthesis, regulat es not only the levels of 5-S-CD and DOPA but also the melanogenic activity of pigment cells. Since DOPA functions as a cofactor in the monophenolase reaction of tyrosinase, it is proposed that the ratio of S-CD to DOPA may b e an important factor in the regulation of tyrosinase activity in situ.