MODULATION OF 5-S-CYSTEINYLDOPA FORMATION BY TYROSINASE ACTIVITY AND INTRACELLULAR THIOLS IN HUMAN-MELANOMA CELLS

The catechol 5-S-cysteinyldopa (5-S-CD) is produced in large amounts i n metastatic malignant melanoma, To further understand the mechanism o f formation of 5-S-CD, we investigated the effects of thiol modulating agents and melanin precursors on human melanoma cells, Under standard culture conditions (0.1 mM cystine), the cell levels of 5-S-CD were h ighly correlated with the degree of melanization and the dopa oxidase activity of the four cell lines investigated (Me8, JUSO, GLL19, Swift) , Inhibition of glutathione (GSH) biosynthesis with buthionine sulphox imine did not affect 5-S-CD levels in the low melanotic GL19 cells, In contrast, the highly pigmented Swift cells showed a strong increase i n the cell levels of cystine (CysH) and 5-S-CD, When the cystine conce ntration of the growth medium was increased to 0.2 mM, a similar situa tion of 5-S-CD synthesis caused by an increase in intracellular CysH l evels was observed in the Swift cells. The GLL19 cells showed enhanced 5-S-CD formation in the presence of 0.1 mM L-dopa. This effect was as sociated with a fourfold increase in dopa oxidase activity, Our data c learly indicate that 5-S-CD is formed in human melanoma cells by a tyr osinase-dependent mechanism involving the addition of CysH to dopaquin one, Based on the enhancing effect of buthionine sulphoximine on 5-S-C D formation, it is proposed that GSH is not directly implicated in 5-S -CD formation, but regulates CysH levels via the enzyme gamma-glutamyl cysteine synthetase.