title: Cell-specific regulation of hypoxia-inducible factor (HIF)-1alpha and HIF-2alpha stabilization and transactivation in a graded oxygen environment.
abstract: The hypoxia-inducible factor (HIF)-1alpha and HIF-2alpha are closely related, key transcriptional regulators of the hypoxic response, countering a low oxygen situation with the up-regulation of target genes associated with numerous processes, including vascularization and glycolysis. This involves a dual mechanism of control through both stabilization and transactivation, regulated via prolyl and asparaginyl hydroxylation. Despite high similarity with respect to protein sequence and activation pathway, a growing number of physiological and mechanistic differences between HIF-1alpha and HIF-2alpha are being reported. To further characterize this nonredundancy, the stabilization of endogenous proteins and regulation of the transactivation domains were compared in a graded oxygen environment across a series of cell lines. Although generally similar results were found, interesting and specific differences between the HIF-alpha proteins were observed within certain cell lines, such as rat adrenal PC12s, emphasizing the cell-specific nature of HIF-alpha regulation. We characterize a conserved amino acid substitution between HIF-1alpha and HIF-2alpha that contributes to the intrinsically higher FIH-1-mediated asparaginyl hydroxylation of HIF-1alpha and, hence, lower HIF-1alpha activity. In addition, our data demonstrate that the different cell lines can be classified into two distinct groups: those in which stabilization and transactivation proceed in conjunction (HeLa, 293T, and COS-1) and those cells in which HIF-alpha is stabilized prior to transactivation (PC12, HepG2, and CACO2). Interestingly, the initial stabilization of HIF-alpha prior to transactivation up-regulation predicted from in vitro derived hydroxylation data is only true for a subset of cells.
authors.affiliation: School of Molecular and Biomedical Science, University of Adelaide, South Australia.
authors.forename: Cameron P
authors.initials: CP
authors.lastname: Bracken
authors.forename: Anthony O
authors.initials: AO
authors.lastname: Fedele
authors.forename: Sarah
authors.initials: S
authors.lastname: Linke
authors.forename: Wiltiana
authors.initials: W
authors.lastname: Balrak
authors.forename: Karolina
authors.initials: K
authors.lastname: Lisy
authors.forename: Murray L
authors.initials: ML
authors.lastname: Whitelaw
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: Peet
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins; D019556:COS Cells; D018938:Caco-2 Cells; D002522:Chlorocebus aethiops; D005786:Gene Expression Regulation; D006367:HeLa Cells; D006801:Humans; D000860:Hypoxia; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000378:metabolism* / Q000502:physiology*; D008969:Molecular Sequence Data; D010100:Oxygen / Q000378:metabolism*; D016716:PC12 Cells; D051381:Rats; D017386:Sequence Homology, Amino Acid; D014157:Transcription Factors / Q000378:metabolism* / Q000502:physiology*; D015533:Transcriptional Activation; D000099162:Endothelial PAS Domain-Containing Protein 1
chemical_list: D051792:Basic Helix-Loop-Helix Proteins; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D010100:Oxygen; D014157:Transcription Factors; D000099162:Endothelial PAS Domain-Containing Protein 1; C497442:HIF1A protein, human