Mechanism of regulation of the hypoxia-inducible factor-1 alpha by the von Hippel-Lindau tumor suppressor protein.
In normoxic cells the hypoxia-inducible factor-1 alpha (HIF-1 alpha) is rapidly degraded by the ubiquitin-proteasome pathway, and activation of HIF-1 alpha to a functional form requires protein stabilization. Here we show that the product of the von Hippel-Lindau (VHL) tumor suppressor gene mediated ubiquitylation and proteasomal degradation of HIF-1 alpha under normoxic conditions via interaction with the core of the oxygen-dependent degradation domain of HIF-1 alpha. The region of VHL mediating interaction with HIF-1 alpha overlapped with a putative macromolecular binding site observed within the crystal structure of VHL. This motif of VHL also represents a mutational hotspot in tumors, and one of these mutations impaired interaction with HIF-1 alpha and subsequent degradation. Interestingly, the VHL binding site within HIF-1 alpha overlapped with one of the minimal transactivation domains. Protection of HIF-1 alpha against degradation by VHL was a multistep mechanism, including hypoxia-induced nuclear translocation of HIF-1 alpha and an intranuclear hypoxia-dependent signal. VHL was not released from HIF-1 alpha during this process. Finally, stabilization of HIF-1 alpha protein levels per se did not totally bypass the need of the hypoxic signal for generating the transactivation response.
Tanimoto
K
K
Department of Cell and Molecular Biology, Medical Nobel Institute, Karolinska Institutet, S-171 77 Stockholm, Sweden.
Makino
Y
Y
Pereira
T
T
Poellinger
L
L
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D019556:COS Cells; D002460:Cell Line; D018360:Crystallography, X-Ray; D003546:Cysteine Endopeptidases / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005656:Fungal Proteins / Q000378:metabolism; D049452:Green Fluorescent Proteins; D006801:Humans; D000860:Hypoxia; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D015151:Immunoblotting; D008025:Ligases; D008164:Luminescent Proteins / Q000378:metabolism; D008954:Models, Biological; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000378:metabolism; D009154:Mutation; D009687:Nuclear Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010100:Oxygen / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D046988:Proteasome Endopeptidase Complex; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D014157:Transcription Factors / Q000378:metabolism; D015533:Transcriptional Activation; D014162:Transfection; D025521:Tumor Suppressor Proteins; D044767:Ubiquitin-Protein Ligases; D014452:Ubiquitins / Q000378:metabolism; D051794:Von Hippel-Lindau Tumor Suppressor Protein
D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C052130:GAL4 protein, S cerevisiae; C497442:HIF1A protein, human; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D008164:Luminescent Proteins; D009097:Multienzyme Complexes; D009687:Nuclear Proteins; D011506:Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; D014452:Ubiquitins; D049452:Green Fluorescent Proteins; D044767:Ubiquitin-Protein Ligases; D051794:Von Hippel-Lindau Tumor Suppressor Protein; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; D008025:Ligases; C497446:VHL protein, human; D010100:Oxygen