Tumor growth is the orchestration of various oncogenes and tumor suppressors, and the regulation of these genes offers a rational therapeutic approach to cancer treatment. In this study, we found a new regulator of tumor growth, phosphatidylinositol 4-kinase type IIalpha (PI4KIIalpha), the mechanism of which is involved in angiogenesis and hypoxia-inducible factor HIF-1alpha regulation. Results obtained from a human cancer tissue microarray showed that PI4KIIalpha protein expression increases markedly in seven types of cancers compared with normal tissues. Suppression of PI4KIIalpha leads to retarded tumor growth in nude mice. Downregulation of PI4KIIalpha in cancer cells eliminates tumor cell-induced endothelial cell tubulogenesis and migration, and results in impaired angiogenesis. Further investigation showed that PI4KIIalpha can directly regulate HIF-1alpha expression and that the expression of these two proteins is correlated in vivo. At the same time, downregulation of PI4KIIalpha markedly reduces HER-2 autophosphorylation, and PI4KIIalpha specifically triggers HIF-1alpha accumulation through a phosphatidylinositol 3-kinase (PI3K)- and extracellular signal-regulated protein kinase (ERK)-dependent pathway, suggesting that PI4KIIalpha may regulate HIF-1alpha through the HER-2/PI3K, ERK cascade. In summary, we discovered a pivotal role for PI4KIIalpha in the regulation of tumor growth. Our results shed new light on understanding the novel functions of PI4KIIalpha in cancer and suggest that PI4KIIalpha may be a promising specific target for tumor therapy.