Manganese superoxide dismutase modulates hypoxia-inducible factor-1α induction via superoxide

Hypoxia-inducible factor 1 (HIF-1) is a transcription factor that plays an important role in O-2 homeostasis. Numerous observations suggest that changes in reactive oxygen species affect HIF-1 alpha stabilization and HIF-1 alpha transcriptional activation in many cell types. The antioxidant enzyme manganese superoxide dismutase (MnSOD) modulates the cellular redox environment by converting superoxide (O-2(center dot-)) to hydrogen peroxide and dioxygen. Previous results from our group have shown that overexpression of MnSOD in MCF-7 cells alters stabilization of HIF-1 alpha under hypoxic conditions; however, the underlying mechanism(s) is not known. Here, we tested the hypothesis that MnSOD regulates the expression of HIF-1 alpha by modulating the steady-state level of O-2(center dot-). We found that decreasing MnSOD with small interfering RNA in MCF-7 cells resulted in (a) an associated increase in the hypoxic accumulation of HIF-1 alpha immunoreactive protein, (b) a significant increase in the levels of O-2(center dot-) (P < 0.01), but (c) no significant change in the steady-state level of H2O2. Removal of O-2(center dot-) using spin traps (alpha-4-pyridyl-l-oxide-N-tert-butylnitrone and 5,5-dimethyl-1-pyrroline N-oxide) or the O-2(center dot-) scavenger Tempol or an SOD mimic (AEOL10113) resulted in a decrease in HIF-1 alpha protein, consistent with the hypothesis that O-2(center dot-) is an important molecular effector responsible for hypoxic stabilization of HIF-1 alpha. The evidence from both genetic and pharmaceutical manipulation is consistent with our hypothesis that O-2(center dot-) can contribute to the stabilization of HIF-1 alpha.