Genome-wide Association of Hypoxia-inducible Factor (HIF)-1α and HIF-2α DNA Binding with Expression Profiling of Hypoxia-inducible Transcripts

Hypoxia-inducible factor (HIF) controls an extensive range of adaptive responses to hypoxia. To better understand this transcriptional cascade we performed genome-wide chromatin immuno-precipitation using antibodies to two major HIF-alpha subunits, and correlated the results with genome-wide transcript profiling. Within a tiled promoter array we identified 546 and 143 sequences that bound, respectively, to HIF-1 alpha or HIF-2 alpha at high stringency. Analysis of these sequences confirmed an identical core binding motif for HIF-1 alpha and HIF-2 alpha (RCGTG) but demonstrated that binding to this motif was highly selective, with binding enriched at distinct regions both upstream and downstream of the transcriptional start. Comparison of HIF-promoter binding data with bidirectional HIF-dependent changes in transcript expression indicated that whereas a substantial proportion of positive responses (> 20% across all significantly regulated genes) are direct, HIF-dependent gene suppression is almost entirely indirect. Comparison of HIF-1 alpha- versus HIF-2 alpha-binding sites revealed that whereas some loci bound HIF-1 alpha in isolation, many bound both isoforms with similar affinity. Despite high-affinity binding to multiple promoters, HIF-2 alpha contributed to few, if any, of the transcriptional responses to acute hypoxia at these loci. Given emerging evidence for biologically distinct functions of HIF-1 alpha versus HIF-2 alpha understanding the mechanisms restricting HIF-2 alpha activity will be of interest.