Oxidants are toxic, but at low doses they can stimulate rather than inhibit the growth of mammalian cells and play a role in the etiology of cancer and fibrosis. The effect of oxidants on cells is modulated by multiple interacting antioxidant defense systems. We have studied the individual roles and the interaction of Cu,Zn-superoxide dismutase (SOD) and catalase (CAT) in transfectants with human cDNAs of mouse epidermal cells JB6 clone 41. Since only moderate increases in these enzymes are physiologically meaningful, we chose the following five clones for in-depth characterization: CAT 4 and CAT 12 with 2.6-fold and 4.2-fold increased catalase activities, respectively, SOD 15 and SOD 3 with 2.3-fold and 3.6-fold increased Cu,Zn-SOD activities, respectively, and SOCAT 3 with a 3-fold higher catalase activity and 1.7-fold higher Cu,Zn-SOD activity than the parent JB6 clone 41. While the increases in enzyme activities were moderate, the human cDNAs were highly expressed in the transfectants. As demonstrated for the clone SOD 15, this discordance between message concentrations and enzyme activities may be due to the low stability of the human Cu,Zn-SOD mRNA in the mouse recipient cells. According to immunoblots the content of Mn-SOD was unaltered in the transfectants. While the activities of glutathione peroxidase were comparable in all strains, the concentrations of reduced glutathione (GSH) were significantly lower in SOD 3 and SOD 15. This decrease in GSH may reflect a chronic prooxidant state in these Cu,Zn-SOD overproducers. The Cu,Zn-SOD overproducers SOD 15 and SOD 3 were hypersensitive to the formation of DNA single-strand breaks, growth retardation, and killing by an extracellular burst of superoxide plus H2O2 while the CAT overproducers were protected relative to the parent clone JB6 clone 41. The double transfectant SOCAT 3 was well protected from oxidant damage because of its increased content in CAT, which counterbalances the increase in Cu,Zn-SOD. The inducibility of the growth-competence-related protooncogene c-fos was decreased in all transfectants, albeit probably for different reasons. We conclude that H2O2 represents the major intracellular oxidant on the pathways to DNA single-strand breakage and cytotoxicity and that the hypersensitivity of Cu,Zn-SOD transfectants is mostly due to the overproduction of H2O2. The balance of SOD and CAT plus glutathione peroxidase is more important for overall sensitivity than the level of Cu,Zn-SOD alone. Growth stimulation may occur when cells are protected from excessive oxidant toxicity but only when a sufficient oxidant signal remains to activate the necessary pathways.
