Metallothioneins (MTs) protect cells from the toxic effects of heavy metals. It has been suggested that they play a role in cellular resistance to alkylating agents and ionizing radiation because of the coincidence of cadmium- and drug-resistance and by virtue of the reactivity of MT with free radicals. We report the analysis of mouse B16 melanoma cell lines with high and low constitutive MT expression. In these cells, both cisplatin and cadmium resistance were associated with constitutive MT accumulation in the absence of heavy-metal induction. However, in cells with high constitutive MT expression (where zinc treatment did not induce increased MT expression), cisplatin resistance, but not cadmium resistance, was increased approximately twofold by zinc treatment. Methotrexate resistance also was increased by zinc treatment in some cases. We conclude that MT is associated with cisplatin resistance, but that effects of heavy-metal treatment other than MT induction are also responsible for cisplatin and methotrexate, but not cadmium, resistance.