FIG. 10.

Functional HIF-1 binding to the NOR-1 promoter in hypoxic cells. (A) The relative in vivo association of HIF-1 with the human NOR-1 promoter was analyzed by ChIP in HUVEC incubated under normoxia (Norm), hypoxia (1% O₂), or CoCl₂ (1 mM) for 4 h in the presence or in the absence (−) of the PI3K/Akt inhibitor LY294002 (LY). Sheared chromatin was immunoprecipitated with an anti-HIF-1α antibody or a nonspecific goat IgG. The enrichment of HIF-1α was quantified by real-time PCR using NOR-1 promoter-specific primers. Data were normalized to the total input DNA and are represented as means ± SEM of two independent experiments performed in duplicate. *, P < 0.05 versus cells exposed to normoxia; #, P < 0.05 versus cells exposed to hypoxia (or treated with CoCl₂) in the absence of inhibitors. (B) Agarose gel electrophoresis of PCR products. Equal input DNA and control IgG (CT IgG) immunoprecipitations are shown.