Response of amplified metallothionein genes to zinc induction

With a stable cadmium resistant CHO mutant, the authors have examined the effect of exogenous zinc on the induction of two metallothionein isogenes at both the protein and mRNA levels. This mutant is over 200-fold less sensitive than wild type to cadmium toxicity. Restriction mapping and hybridization analysis show a nearly 100-fold amplification in MT gene copies. By dot hybridization, as much as 90-fold increase in MT mRNA could be detected within 24 hours after zinc was added to the culture medium; the elevation is detectable within 20-minutes. Both isospecies, MT 1 and MT 2 are induced coordinately as measured by stringent hybridization with their respective cDNA probes. The response is dose dependent up to 200 ..mu..M of zinc, beyond which cells began to show cytotoxicity. The level of mRNA accumulation increased with time up to 30 hours; thereafter, mRNA levels declined. Zinc supply was not exhausted in this conditioned media which could induce new MT mRNA and protein synthesis in naive cells. It is possible that elevated methallothioneins generated in the induced cells prevent or retard further induction by zinc. Metallothionein may thus regulate the expression of its coding genes by sequestering metals and controlling inducer availability.