Heat induction of the unphosphorylated form of hypoxia-inducible factor-1α is dependent on heat shock protein-90 activity

Hypoxia-inducible factor (HIF)-1alpha is the oxygen-sensitive subunit of HIF-1alpha transcriptional master regulator of oxygen homeostasis. Oxygen-dependent prolyl hydroxylation targets HIF-1alpha for ubiquitinylation and proteasomal degradation. Unexpectedly, we found that exposing mice to elevated temperatures resulted in a strong HIF-1alpha induction in kidney, liver, and spleen. To elucidate the molecular mechanisms responsible for this effect, HepG2 hepatoma cells were exposed to different temperatures (34-42 degreesC) under normoxic (20% O-2) or hypoxic (3% O-2) conditions. Heat was sufficient to stabilize mainly a phosphatase-resistant, low molecular weight form of HIF-1alpha (termed HIF-1alpha.). Heat-induced HIF-1alpha. accumulated in the nucleus but neither bound to DNA nor trans-activated reporter or target gene expression, demonstrating the need for post-translational modifications for these functions. The protein banding pattern of heat-induced HIF-1alpha in immunoblot analyses was clearly distinct from the HIF-1alpha pattern after prolyl hydroxylase inhibition (by hypoxia or iron chelation/replacement) or following proteasome, inhibition, suggesting that heat stabilizes HIF-1alpha by a novel mechanism. Inhibition of the ATP-dependent chaperone activity of HSP90 by novobiocin or geldanamycin prevented heat-induced as well as hypoxia-induced HIF-1alpha accumulation, indicating a common role of the HSP90 chaperone activity in HIF-1alpha stabilization by these two environmental parameters.