Hypoxia-inducible factor 1alpha (HIF-1α), which transactivates a variety of hypoxia-induced genes, is rapidly degraded under nomoxia through the hydroxylation-ubiquitination-proteasome pathway. In this study, we addressed how HIF-1α is stabilized by proteasome inhibitors. The ubiquitin pool was rapidly reduced after proteasome inhibition, followed by the accumulation of non-ubiquitinated HIF-1α. The poly-ubiquitination of HIF-1α was resumed by restoration of free ubiquitin, which suggests that the HIF-1α stabilization under proteasome inhibition is attributed to depletion of the free ubiquitin pool. Ni(2+) and Zn(2+) also stabilized HIF-1α with depletion of the free ubiquitin pool and these effects of metal ions were attenuated by restoration of free ubiquitin. Ni(2+) and Zn(2+) may disturb the recycling of free ubiquitin, as MG132 does. Based on these results, the state of the ubiquitin pool seems to be another critical factor determining the cellular level of HIF-1α.
Keywords: Hypoxia-inducible Factor 1, Alpha Subunit; Poly-ubiquitination; Proteasome Inhibitors; Transition Metal Ions; Ubiquitin Depletion.