Hydroxylation at an asparagine residue at the COOH-terminal activation domain of hypoxia-inducible factor (HIF)-1/2 alphas is essential for its inactivation under normoxic condition. To date, the mechanism by which HIF-alpha avoids the inhibitory effect of asparagine hydroxylase in renal cell carcinoma (RCC) in normoxia is undefined. We have shown herein that protein kinase C (PKC) zeta has an important role in HIF-alpha activation in RCC. By using dominant negative mutant and small interference RNA approaches, we have demonstrated that the association between HIF-alpha and p300 is modulated by PKCzeta. Moreover, a novel signaling pathway involving phosphatidylinositol 3'-kinase and PKCzeta has been shown to be responsible for the activation of HIF-alpha by inhibiting the mRNA expression of FIH-1 (factor inhibiting HIF-1) in RCC and thereby promoting the transcription of hypoxia-inducible genes such as vascular permeability factor/vascular endothelial growth factor.