Figure 2.

HIF-1α knockdown results in G₁ accumulation and p21 induction in HeLa cells. (A) Confirmation of HIF-1α knockdown using HeLa whole cell lysates (WCL) for western blot analysis. Samples were harvested following siRNA protocol and 2 h of DFX treatment. (B) HeLa cells were transfected with nontargeted or HIF-1α siRNA oligonucleotides prior to harvesting for cell cycle analysis using the propidium iodide staining protocol. (C) HeLa cells were transfected using nontargeted or HIF-1α siRNA prior to harvest of WCL. Samples were analyzed by western blot using the specific antibodies indicated. (D) HeLa cells were transfected as in (B) but mRNA was extracted. HIF-1α, p21, p27 and GLUT3 mRNA were analyzed using quantitative PCR. Graph depicts mRNA levels normalized to actin. Student's t-tests (two tailed) were performed and p-values calculated. *p ≤ 0.05 and **p ≤ 0.01. (E) HeLa cells were transfected with control and HIF-2α siRNA oligonucleotides and mRNA extracted. HIF-1α, HIF-2α, p21, p27 and GLUT3 mRNA were analyzed using quantitative PCR. Graph depicts mRNA levels normalized to actin. Student's t-tests (two tailed) were performed and p-values calculated. *p ≤ 0.05 and **p ≤ 0.01. Data refers to a minimum of three independent experiments.