Hypoxia-inducible factor-1alpha (HIF-1alpha) is a main responder to intracellular hypoxia and is overexpressed in many human cancers, including renal cell carcinoma (RCC). To better understanding of the role of HIF-1alpha in the tumorigenicity of RCC, we used short-hairpin RNA (shRNA) interference to inhibit HIF-1alpha expression in the human renal cancer cell line, Caki-1 and OS-RC-2. Silencing of HIF-1alpha significantly reduced the expression of HIF-1alpha in both of renal cancer cell lines. In vitro downregulation of HIF-1alpha inhibited Caki-1 and OS-RC-2 cell growth, migration and invasion. The results further showed that HIF-1alpha silencing resulted in caspase-dependent apoptosis of Caki-1 and OS-RC-2 through regulation of PI3K/Akt pathway and Bcl-2-related proteins expression. In vivo animal studies showed that tumor growth was significantly inhibited in HIF-1alpha shRNA-transfected RCC. Intratumor gene therapy with polyethylenimine-loaded HIF-1alpha shRNA also resulted in tumor growth suppression. Thus, this study demonstrates that downregulation of HIF-1alpha could suppress tumorigenicity of RCC through induction of apoptosis, and HIF-1alpha shRNA may be a promising strategy for the treatment of RCC.