Hypoxia-inducible factor-1α (HIF-1α), a member of the transcription family characterized by a basic helix-loop-helix (bHLH) domain and a PAS domain, regulates the transcription of hypoxia-inducible genes involved in erythropoiesis, vascular remodelling and glucose/energy metabolism. It contains bHLH/PAS domains in the N-terminal half, and a nuclear localization signal (NLS) and two transactivation domains (TADs) in the C-terminal half. It also has an oxygen-dependent degradation (ODD) domain, which is required to degrade HIF-1α protein by the ubiquitin—proteasome pathway. In this study, we identified a new alternatively spliced variant of human HIF-1α mRNA, which lacked both exons 11 and 12, producing a frame shift and giving a shorter form of HIF-1α. In the corresponding protein, a part of the ODD domain, both TADs and the C-terminal NLS motif were missing. Expression of endogenous HIF-1α variant protein was identified using immunoprecipitation and immunoblotting methods. The expressed HIF-1α variant exhibited neither the activity of transactivation nor hypoxia-induced nuclear translocation. In contrast with HIF-1α, the variant was strikingly stable in normoxic conditions and not up-regulated to such an extent by hypoxia, cobalt ions or desferrioxamine. It was also demonstrated that the HIF-1α variant competed with endogenous HIF-1α and suppressed HIF-1 activity, resulting in the down-regulation of mRNA expression of hypoxia-inducible genes. The association of the variant and arylhydrocarbon receptor nuclear translocator in the cytoplasm may be related to the inhibition of HIF-1 activity. It is assumed that this isoform preserves the balance between aerobic and anaerobic metabolism by counteracting the overaction of HIF-1α.
Abbreviations used: bHLH, basic helix-loop-helix; HIF-1, hypoxia-inducible factor 1; ARNT, arylhydrocarbon receptor nuclear translocator; AINT, ARNT-interacting protein; TAD, transactivation domain; NLS, nuclear localization signal; ODD, oxygen-dependent degradation; pVHL, von Hippel-Lindau tumour suppressor protein; RT-PCR, reverse transcriptase PCR; EPO, erythropoietin; VEGF, vascular endothelial growth factor; HA, haemagglutinin; EMSA, electrophoretic mobility-shift assay; CBZ-LLL, N-carbobenzoxyl-Leu-Leu-Leu-al; EPAS1, endothelial PAS domain protein 1.