Cu,Zn superoxide dismutase and copper deprivation and toxicity in Saccharomyces cerevisiae

A wild-type strain of the yeast Saccharomyces cerevisiae grown at a medium [Cu] of ≤ 50 nM contained less Cu,Zn superoxide dismutase (SOD) mRNA (60%), protein (50%), and activity (50%) in comparison with control cultures grown in normal synthetic dextrose medium ([Cu ~ 150 nM). A compensating increase in the activity of MnSOD was observed, as well as a smaller increase in MnSOD mRNA. These medium [Cu]-dependent differences were observed in cultures under N₂ as well. Addition of Cu²⁺ (100 μM) to Cu-depleted cultures resulted in a rapid (30 min) increase in Cu,ZnSOD mRNA (2.5-fold), protein (3.5-fold), and activity (4-fold). Ethidium bromide (200 μg/ml of culture) inhibited by 50% the increase in Cu,ZnSOD mRNA, while cycloheximide (100 μg/ml of culture) inhibited completely the increase in protein and activity. Addition of Cu²⁺ to ≥ 100 μM caused no further increase in these parameters but did result in a loss of total cellular RNA and translatable RNA, a decline in a population of specific mRNAs, a decrease in total soluble protein and the activity of specific enzymes, and an inhibition of incorporation of [³H]uracil and [³H]leucine into trichloroacetic acid-insoluble material. Cu,ZnSOD mRNA, protein, and activity appeared relatively more resistant to these effects of Cu toxicity than did the other cellular constituents examined. When evaluated in cultures under N₂, the cellular response to [Cu] of ≥ 100 μM was limited to the inhibition of radiolabel incorporation into trichloroacetic acid-insoluble material. All other effects were absent in the absence of O₂. The data indicated that medium (cellular) Cu alters the steady-state level of Cu,ZnSOD. This regulation may be at the level of transcription. In addition, Cu,ZnSOD exhibits the characteristics of a Cu-stress protein in that it and its mRNA are enhanced relative to other cellular species under conditions of Cu excess. This observation and the O₂-dependence of some of the manifestations of Cu excess suggest that one mechanism of Cu toxicity involves the superoxide radical anion, O₂^-.