Upstream stimulatory factor 2 and hypoxia-inducible factor 2α (HIF2α) cooperatively activate HIF2 target genes during hypoxia.

While the functions of hypoxia-inducible factor 1α (HIF1α)/aryl hydrocarbon receptor nuclear translocator (ARNT) and HIF2α/ARNT (HIF2) proteins in activating hypoxia-inducible genes are well established, the role of other transcription factors in the hypoxic transcriptional response is less clear. We report here for the first time that the basic helix-loop-helix-leucine-zip transcription factor upstream stimulatory factor 2 (USF2) is required for the hypoxic transcriptional response, specifically, for hypoxic activation of HIF2 target genes. We show that inhibiting USF2 activity greatly reduces hypoxic induction of HIF2 target genes in cell lines that have USF2 activity, while inducing USF2 activity in cells lacking USF2 activity restores hypoxic induction of HIF2 target genes. Mechanistically, USF2 activates HIF2 target genes by binding to HIF2 target gene promoters, interacting with HIF2α protein, and recruiting coactivators CBP and p300 to form enhanceosome complexes that contain HIF2α, USF2, CBP, p300, and RNA polymerase II on HIF2 target gene promoters. Functionally, the effect of USF2 knockdown on proliferation, motility, and clonogenic survival of HIF2-dependent tumor cells in vitro is phenocopied by HIF2α knockdown, indicating that USF2 works with HIF2 to activate HIF2 target genes and to drive HIF2-depedent tumorigenesis.