Methionine dependence, the inability of cells to grow when the amino acid methionine is replaced in culture medium by its metabolic precursor homocysteine, is characteristic of many cancer cell lines and some tumors in situ. Most cell lines proliferate normally under these conditions. The methionine dependent tumorigenic human melanoma cell line MeWo-LC1 was derived from the methionine independent non-tumorigenic line, MeWo. MeWo-LC1 has a cellular phenotype identical to that of cells from patients with the cblC inborn error of cobalamin metabolism, with decreased synthesis of cobalamin coenzymes and decreased activity of the cobalamin-dependent enzymes methionine synthase and methylmalonylCoA mutase. Inability of cblC cells to complement the defect in MeWo-LC1 suggested that it was caused by decreased activity of the MMACHC gene. However, no potentially disease causing mutations were detected in the coding sequence of MMACHC in MeWo-LC1. No MMACHC expression was detected in MeWo-LC1 by quantitative or non-quantitative PCR. There was virtually complete methylation of a CpG island at the 5'-end of the MMACHC gene in MeWo-LC1, consistent with inactivation of the gene by methylation. The CpG island was partially methylated (30-45%) in MeWo and only lightly methylated (2-11%) in control fibroblasts. Infection of MeWo-LC1 with wild type MMACHC resulted in correction of the defect in cobalamin metabolism and restoration of the ability of cells to grow in medium containing homocysteine. We conclude that epigenetic inactivation of the MMACHC gene is responsible for methionine dependence in MeWo-LC1.