Figure 5.

HIF-1α/HIF-1β heterodimer is present at the KDM2A promoter and is required for RNA polymerase II recruitment. (A) Schematic diagram of the KDM2A and KDM2B genes and annotation of putative hypoxia responsive elements (HREs) in different species; (B) HeLa cells were exposed to 1% O₂ for 24 h prior to crosslinking and lysis. Chromatin immunoprecipitations (ChIPs) were performed for the levels of HIF-1α and HIF-1β present at the KDM2A and KDM2B promoters. IgG was used as the antibody control; (C) HeLa cells were transfected with the indicated siRNA oligonucleotides for 72 h prior to cross-linking and lysis. In addition, cells were exposed to 1% O₂ for the last 24 h. ChIP was performed for the level of RNA polymerase II present at the KDM2A promoter, with IgG used as the antibody control. Graphs depict mean and standard deviation from a minimum of three independent experiments performed in duplicate. Student t-test was performed and p values calculated. * p < 0.05, ** p < 0.01, *** p < 0.001; (D) HEK293 cells were transfected with 500 ng PGL3 (control) vector or KDM2A HRE–luciferase constructs as indicted for 48 h prior to lysis and luciferase activity measured. Graph depicts mean and standard error of mean of a minimum of three independent experiments. Student t-test was performed and p values calculated. * p < 0.05, ** p < 0.01, *** p < 0.001.