The ubiquitously expressed hypoxia-inducible factor-1 (HIF-1) is involved in expression of a large number of oxygen-regulated genes. HIF-1 is a heterodimer consisting of an α and a β subunit, both belonging to the basic-helix-loop-helix Per-aryl hydrocarbon receptor nuclear translocator-Sim (PAS) family of transcription factors. Whereas HIF-1α is a novel member of this family, HIF-1β is identical to the aryl hydrocarbon receptor nuclear translocator, previously recognized to be involved in xenobiotic metabolism. cDNA cloning revealed that mouse HIF-1α can be expressed as two mRNA isoforms containing alternative 5′ untranslated regions and two different predicted translational start sites. We cloned and characterized 20.5 kb of the mouse HIF-1α gene (Hifla) containing exon II–XV. The two alternative first exons, I.1 and I.2, are separated from exon II by approximately 24 kb and 17 kb, respectively. We also sequenced Hifla exon I.1 and flanking regions, and mapped a single exon I.1 transcription initiation site. Reverse transcription PCR analysis of total RNA derived from normoxic and hypoxic mouse hepatoma and fibroblast cell lines suggested that the two alternative mRNA isoforms are constitutively co-expressed in these cells, and that two different promoters drive transcription of HIF-1α. A minimal exon I.1 promoter was identified which moderately activated heterologous gene expression, indicating that additional cis-elements are required for efficient HIF-1α transcription in vivo.
aryl hydrocarbon receptor
AhR nuclear translocator
- EMS A
electrophoretic mobility shift assay
rapid amplification of cDNA ends
vascular endothelial growth factor