Resistance to copper toxicity of cultured hepatoma cells. Characterization of resistant cell lines.

A series of four cell lines resistant to the toxic effect of copper were developed from Morris rat hepatoma cells by gradually increasing the concentration of copper in the growth medium. The EC50, that concentration of copper that kills and/or inhibits the growth of 50% of the cells after 72 h, increased 4-fold over that for wild type cells in the most resistant cell line. These cells were also resistant to zinc, cadmium, and mercury toxicity, but not to nickel or cobalt. The amount of copper in the soluble protein pool of the resistant cells increased proportionally with the concentration of copper in the medium in which they were maintained. Associated with copper accumulation was the production of an 18-kDa cysteine-rich protein which complexes a significant amount of the metal. It is suggested that resistance to copper toxicity is due to sequestration of the metal by this protein. When resistant cells were removed from the copper-enriched environment, cellular copper levels rapidly fell to that observed for wild type cells, but no reduction in either the EC50 or the level of the cysteine-rich protein was noted. This suggests that a permanent change responsible for copper resistance had occurred which is maintained in the absence of the metal.