Cell-specific regulation of hypoxia-inducible factor (HIF)-1alpha and HIF-2alpha stabilization and transactivation in a graded oxygen environment

J Biol Chem. 2006 Aug 11;281(32):22575-85. doi: 10.1074/jbc.M600288200. Epub 2006 Jun 7.

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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Basic Helix-Loop-Helix Transcription Factors
  • COS Cells
  • Caco-2 Cells
  • Chlorocebus aethiops
  • Gene Expression Regulation*
  • HeLa Cells
  • Humans
  • Hypoxia
  • Hypoxia-Inducible Factor 1, alpha Subunit / metabolism*
  • Hypoxia-Inducible Factor 1, alpha Subunit / physiology*
  • Molecular Sequence Data
  • Oxygen / metabolism*
  • PC12 Cells
  • Rats
  • Sequence Homology, Amino Acid
  • Transcription Factors / metabolism*
  • Transcription Factors / physiology*
  • Transcriptional Activation

Substances

  • Basic Helix-Loop-Helix Transcription Factors
  • HIF1A protein, human
  • Hypoxia-Inducible Factor 1, alpha Subunit
  • Transcription Factors
  • endothelial PAS domain-containing protein 1
  • Oxygen