Changes in cobalamin metabolism are associated with the altered methionine auxotrophy of highly growth autonomous human melanoma cells.

Our aim was to identify the biochemical defect responsible for the inability of highly growth autonomous human tumor cells to proliferate in culture medium devoid of methionine, but containing homocysteine and 5-methyletrahydrofolic acid. We have adopted the terms "homocysteine-responsive" and "homocysteine-nonresponsive" to describe cells which can or cannot proliferate in methionine-free homocysteine-supplemented medium. Using a panel of genetically related homocysteine-responsive and -nonresponsive human melanoma cell lines, the results from a number of experiments indicate that acquisition of the "homocysteine-nonresponsive phenotype" is associated with the reduced intracellular accumulation of methyl-cobalamin, a critical cofactor of the methionine synthase enzyme. When in vitro methionine synthase assays were performed in the presence of exogenously added methyl-cobalamin, specific methionine synthase activity in extracts obtained from homocysteine-responsive cells was only twofold greater than that observed with extracts prepared from homocysteine-nonresponsive cells. However, when exogenous methyl-cobalamin was omitted from the enzyme assays, methionine synthase activity in extracts derived from homocysteine-nonresponsive cells was dramatically reduced, compared with the small decrease observed with homocysteine-responsive cell extracts. Compared with their homocysteine-responsive counterparts, homocysteine-nonresponsive cells exhibited increased levels of cobalamin efflux and decreased intracellular accumulation of methyl-cobalamin. There was a clear relationship between the abilities of these related melanoma cell lines to proliferate in methionine-free homocysteine-supplemented medium, and the extent of cobalamin loss and capacity of exogenously added methyl-cobalamin to stimulate in vitro methionine synthase activity. These results indicate a link between alterations in the intracellular trafficking and/or metabolism of cobalamin and the increased growth autonomy of human melanoma cells.