A redox mechanism controls differential DNA binding activities of hypoxia-inducible factor (HIF) 1alpha and the HIF-like factor.
Hypoxia-inducible factor 1alpha (HIF-1alpha) and the HIF-like factor (HLF) are two highly related basic Helix-Loop-Helix/Per-Arnt-Sim (bHLH/PAS) homology transcription factors that undergo dramatically increased function at low oxygen levels. Despite strong similarities in their activation mechanisms (e.g. they both undergo rapid hypoxia-induced protein stabilization, bind identical target DNA sequences, and induce ... synthetic reporter genes to similar degrees), they are both essential for embryo survival via distinct functions during vascularization (HIF-1alpha) or catecholamine production (HLF). It is currently unknown how such specificity of action is achieved. We report here that DNA binding by HLF, but not by HIF-1alpha, is dependent upon reducing redox conditions. In vitro DNA binding and mammalian two-hybrid assays showed that a unique cysteine in the DNA-binding basic region of HLF is a target for the reducing activity of redox factor Ref-1. Although the N-terminal DNA-binding domain of HIF-1alpha can function in the absence of Ref-1, we found that the C-terminal region containing the transactivation domain requires Ref-1 for full activity. Our data reveal that the hypoxia-inducible factors are subject to complex redox control mechanisms that can target discrete regions of the proteins and are the first to establish a discriminating control mechanism for differential regulation of HIF-1alpha and HLF activity.
Mesh Terms:
Amino Acid Sequence, Carbon-Oxygen Lyases, Cysteine, DNA, DNA-(Apurinic or Apyrimidinic Site) Lyase, DNA-Binding Proteins, Gene Expression Regulation, HeLa Cells, Helix-Loop-Helix Motifs, Humans, Hypoxia-Inducible Factor 1, Hypoxia-Inducible Factor 1, alpha Subunit, Molecular Sequence Data, Mutagenesis, Site-Directed, Nuclear Proteins, Oligonucleotides, Antisense, Oxidation-Reduction, Recombinant Proteins, Sequence Homology, Amino Acid, Serine, Transcription Factors
Amino Acid Sequence, Carbon-Oxygen Lyases, Cysteine, DNA, DNA-(Apurinic or Apyrimidinic Site) Lyase, DNA-Binding Proteins, Gene Expression Regulation, HeLa Cells, Helix-Loop-Helix Motifs, Humans, Hypoxia-Inducible Factor 1, Hypoxia-Inducible Factor 1, alpha Subunit, Molecular Sequence Data, Mutagenesis, Site-Directed, Nuclear Proteins, Oligonucleotides, Antisense, Oxidation-Reduction, Recombinant Proteins, Sequence Homology, Amino Acid, Serine, Transcription Factors
J. Biol. Chem.
Date: Feb. 18, 2000
PubMed ID: 10671489
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