Genome-wide HIF-1 binding sites in HepG2 cells. Homo sapiens

Adaptation to hypoxia is mediated through a coordinated transcriptional response driven largely by Hypoxia-Inducible Factor 1 (HIF-1). The direct transcriptional targets of HIF-1 play important roles in facilitating both short-term and long-term adaptation to hypoxia. Alignment of the sequences encompassing all well-characterized HIF-1 binding sites has revealed a consensus core HRE motif of 5'-RCGTG-3' (R = A or G). Since the consensus HIF-1 binding motif is too promiscuous to accurately predict binding a priori, we used ChIP-chip to define HIF-1 chromatin binding on a genome-wide level. We integrated these results with gene expression profiling to interrogate mechanisms regulating hypoxia-induced gene expression, and to more comprehensively identify direct targets of HIF-1 transactivation. Overall design: HepG2 cells were cultured under normoxic (ambient) or hypoxic (0.5%O2, 4 h) conditions. HIF-1 ChIP was performed with a HIF-1 polyclonal Ab. Triplicate ChIPed DNA and inputs were amplified and hybridized onto the Affymetrix Gene-Chip Human Tiling 2.0R Array Set. The MAT algorithm was used to identify peaks of probe intensity (‘‘hits’’).