HIF-1alpha binding to VHL is regulated by stimulus-sensitive proline hydroxylation.
Hypoxia-inducible factor-1alpha (HIF-1alpha) is a global transcriptional regulator of the hypoxic response. Under normoxic conditions, HIF-1alpha is recognized by the von Hippel-Lindau tumor-suppressor protein (VHL), a component of an E3 ubiquitin ligase complex. This interaction thereby promotes the rapid degradation of HIF-1alpha. Under hypoxic conditions, HIF-1alpha is stabilized. We have previously shown that VHL binds in a hypoxia-sensitive manner to a 27-aa segment of HIF-1alpha, and that this regulation depends on a posttranslational modification of HIF-1alpha. Through a combination of in vivo coimmunoprecipitation assays using VHL and a panel of point mutants of HIF-1alpha in this region, as well as MS and in vitro binding assays, we now provide evidence that this modification, which occurs under normoxic conditions, is hydroxylation of Pro-564 of HIF-1alpha. The data furthermore show that this proline hydroxylation is the primary regulator of VHL binding.
Yu
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Department of Pathology and Laboratory Medicine, University of Pennsylvania Cancer Center, University of Pennsylvania School of Medicine, Philadelphia 19104, USA.
White
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Zhao
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Lee
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D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D015687:Cell Hypoxia; D002522:Chlorocebus aethiops; D003035:Cobalt / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D006900:Hydroxylation; D006909:Hydroxyproline / Q000737:chemistry / Q000502:physiology*; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D008025:Ligases; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism; D017354:Point Mutation; D011392:Proline / Q000737:chemistry; D011485:Protein Binding / Q000187:drug effects; D011499:Protein Processing, Post-Translational; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D014157:Transcription Factors; D014162:Transfection; D025521:Tumor Suppressor Proteins; D044767:Ubiquitin-Protein Ligases; D051794:Von Hippel-Lindau Tumor Suppressor Protein
D004268:DNA-Binding Proteins; C497442:HIF1A protein, human; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D046911:Macromolecular Substances; D009687:Nuclear Proteins; D010446:Peptide Fragments; D011506:Proteins; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; D003035:Cobalt; D011392:Proline; D044767:Ubiquitin-Protein Ligases; D051794:Von Hippel-Lindau Tumor Suppressor Protein; D008025:Ligases; C497446:VHL protein, human; D006909:Hydroxyproline