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PMID: 12912907 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

HIF prolyl-hydroxylase 2 is the key oxygen sensor setting low steady-state levels of HIF-1alpha in normoxia.

The EMBO journal ·Vol. 22 ·No. 16 ·2003-08-15 ·Pages 4082-90

Berra E, Benizri E, Ginouvès A, Volmat V, Roux D, Pouysségur J

Abstract

Hypoxia-inducible factor (HIF), a transcriptional complex conserved from Caenorhabditis elegans to vertebrates, plays a pivotal role in cellular adaptation to low oxygen availability. In normoxia, the HIF-alpha subunits are targeted for destruction by prolyl hydroxylation, a specific modification that provides recognition for the E3 ubiquitin ligase complex containing the von Hippel-Lindau tumour suppressor protein (pVHL). Three HIF prolyl-hydroxylases (PHD1, 2 and 3) were identified recently in mammals and shown to hydroxylate HIF-alpha subunits. Here we show that specific 'silencing' of PHD2 with short interfering RNAs is sufficient to stabilize and activate HIF-1alpha in normoxia in all the human cells investigated. 'Silencing' of PHD1 and PHD3 has no effect on the stability of HIF-1alpha either in normoxia or upon re-oxygenation of cells briefly exposed to hypoxia. We therefore conclude that, in vivo, PHDs have distinct assigned functions, PHD2 being the critical oxygen sensor setting the low steady-state levels of HIF-1alpha in normoxia. Interestingly, PHD2 is upregulated by hypoxia, providing an HIF-1-dependent auto-regulatory mechanism driven by the oxygen tension.

MeSH Terms
Biological Transport Cell Hypoxia Cell Line Cell Nucleus/metabolism DNA-Binding Proteins/chemistry,drug effects,metabolism Dose-Response Relationship, Drug Gene Silencing HeLa Cells Humans Hydroxylation Hypoxia/genetics,metabolism Hypoxia-Inducible Factor 1 Hypoxia-Inducible Factor 1, alpha Subunit Hypoxia-Inducible Factor-Proline Dioxygenases Immediate-Early Proteins Isoenzymes/metabolism Nuclear Proteins/chemistry,drug effects,metabolism Oxygen/metabolism Procollagen-Proline Dioxygenase/metabolism RNA, Messenger/genetics,metabolism RNA, Small Interfering/metabolism,pharmacology Recombinant Proteins/metabolism Transcription Factors/metabolism Transfection Tumor Cells, Cultured Up-Regulation
Chemicals
DNA-Binding Proteins HIF1A protein, human Hypoxia-Inducible Factor 1 Hypoxia-Inducible Factor 1, alpha Subunit Immediate-Early Proteins Isoenzymes Nuclear Proteins RNA, Messenger RNA, Small Interfering Recombinant Proteins Transcription Factors EGLN1 protein, human Procollagen-Proline Dioxygenase Hypoxia-Inducible Factor-Proline Dioxygenases Oxygen
Authors & Affiliations
6 authors, click to expand affiliations / ORCID
Berra Edurne
Institute of Signaling, Developmental Biology and Cancer Research, CNRS UMR 6543, Centre Antoine Lacassagne, 33 Avenue Valombrose, 06189 Nice, France. [email protected]
Benizri Emmanuel
Ginouvès Amandine
Volmat Véronique
Roux Danièle
Pouysségur Jacques
References (49)
49 references, click to expand
  1. Establishment and characterisation of a new tumorigenic cell line with a normal karyotype derived from a human breast adenocarcinoma.
    Br J Cancer. 1990 Jul;62(1):8-13 PMID: 2390488
  2. High-efficiency transformation of mammalian cells by plasmid DNA.
    Mol Cell Biol. 1987 Aug;7(8):2745-52 PMID: 3670292
  3. Hypoxia-inducible factor 1 is a basic-helix-loop-helix-PAS heterodimer regulated by cellular O2 tension.
    Proc Natl Acad Sci U S A. 1995 Jun 6;92(12):5510-4 PMID: 7539918
  4. Activation of hypoxia-inducible transcription factor depends primarily upon redox-sensitive stabilization of its alpha subunit.
    J Biol Chem. 1996 Dec 13;271(50):32253-9 PMID: 8943284
  5. The dual specificity mitogen-activated protein kinase phosphatase-1 and -2 are induced by the p42/p44MAPK cascade.
    J Biol Chem. 1997 Jan 10;272(2):1368-76 PMID: 8995446
  6. Endothelial PAS domain protein 1 (EPAS1), a transcription factor selectively expressed in endothelial cells.
    Genes Dev. 1997 Jan 1;11(1):72-82 PMID: 9000051
  7. A novel bHLH-PAS factor with close sequence similarity to hypoxia-inducible factor 1alpha regulates the VEGF expression and is potentially involved in lung and vascular development.
    Proc Natl Acad Sci U S A. 1997 Apr 29;94(9):4273-8 PMID: 9113979
  8. Hypoxia-inducible factor 1alpha (HIF-1alpha) protein is rapidly degraded by the ubiquitin-proteasome system under normoxic conditions. Its stabilization by hypoxia depends on redox-induced changes.
    J Biol Chem. 1997 Sep 5;272(36):22642-7 PMID: 9278421
  9. V-SRC induces expression of hypoxia-inducible factor 1 (HIF-1) and transcription of genes encoding vascular endothelial growth factor and enolase 1: involvement of HIF-1 in tumor progression.
    Cancer Res. 1997 Dec 1;57(23):5328-35 PMID: 9393757
  10. Regulation of hypoxia-inducible factor 1alpha is mediated by an O2-dependent degradation domain via the ubiquitin-proteasome pathway.
    Proc Natl Acad Sci U S A. 1998 Jul 7;95(14):7987-92 PMID: 9653127
  11. Insulin induces transcription of target genes through the hypoxia-inducible factor HIF-1alpha/ARNT.
    EMBO J. 1998 Sep 1;17(17):5085-94 PMID: 9724644
  12. Hypoxia-inducible factor 1: master regulator of O2 homeostasis.
    Curr Opin Genet Dev. 1998 Oct;8(5):588-94 PMID: 9794818
  13. The VHL tumour-suppressor gene paradigm.
    Trends Genet. 1998 Oct;14(10):423-6 PMID: 9820032
  14. Molecular characterization and chromosomal localization of a third alpha-class hypoxia inducible factor subunit, HIF3alpha.
    Gene Expr. 1998;7(3):205-13 PMID: 9840812
  15. Regulation of the hypoxia-inducible transcription factor 1alpha by the ubiquitin-proteasome pathway.
    J Biol Chem. 1999 Mar 5;274(10):6519-25 PMID: 10037745
  16. Structure of the VHL-ElonginC-ElonginB complex: implications for VHL tumor suppressor function.
    Science. 1999 Apr 16;284(5413):455-61 PMID: 10205047
  17. Rbx1, a component of the VHL tumor suppressor complex and SCF ubiquitin ligase.
    Science. 1999 Apr 23;284(5414):657-61 PMID: 10213691
  18. The tumour suppressor protein VHL targets hypoxia-inducible factors for oxygen-dependent proteolysis.
    Nature. 1999 May 20;399(6733):271-5 PMID: 10353251
  19. The von Hippel-Lindau tumor suppressor protein is a component of an E3 ubiquitin-protein ligase activity.
    Genes Dev. 1999 Jul 15;13(14):1822-33 PMID: 10421634
  20. Hypoxia-inducible factor-1 alpha (HIF-1 alpha) escapes O(2)-driven proteasomal degradation irrespective of its subcellular localization: nucleus or cytoplasm.
    EMBO Rep. 2001 Jul;2(7):615-20 PMID: 11454738
  21. Characterization and comparative analysis of the EGLN gene family.
    Gene. 2001 Sep 5;275(1):125-32 PMID: 11574160
  22. C. elegans EGL-9 and mammalian homologs define a family of dioxygenases that regulate HIF by prolyl hydroxylation.
    Cell. 2001 Oct 5;107(1):43-54 PMID: 11595184
  23. p42/p44 mitogen-activated protein kinases phosphorylate hypoxia-inducible factor 1alpha (HIF-1alpha) and enhance the transcriptional activity of HIF-1.
    J Biol Chem. 1999 Nov 12;274(46):32631-7 PMID: 10551817
  24. Loss of PTEN facilitates HIF-1-mediated gene expression.
    Genes Dev. 2000 Feb 15;14(4):391-6 PMID: 10691731
  25. Identification of alternative spliced variants of human hypoxia-inducible factor-1alpha.
    J Biol Chem. 2000 Mar 10;275(10):6922-7 PMID: 10702253
  26. Ubiquitination of hypoxia-inducible factor requires direct binding to the beta-domain of the von Hippel-Lindau protein.
    Nat Cell Biol. 2000 Jul;2(7):423-7 PMID: 10878807
  27. Hypoxia inducible factor-alpha binding and ubiquitylation by the von Hippel-Lindau tumor suppressor protein.
    J Biol Chem. 2000 Aug 18;275(33):25733-41 PMID: 10823831
  28. Nonhypoxic pathway mediates the induction of hypoxia-inducible factor 1alpha in vascular smooth muscle cells.
    J Biol Chem. 2000 Sep 1;275(35):26765-71 PMID: 10837481
  29. Activation of HIF1alpha ubiquitination by a reconstituted von Hippel-Lindau (VHL) tumor suppressor complex.
    Proc Natl Acad Sci U S A. 2000 Sep 12;97(19):10430-5 PMID: 10973499
  30. HIF-1-dependent transcriptional activity is required for oxygen-mediated HIF-1alpha degradation.
    FEBS Lett. 2001 Feb 23;491(1-2):85-90 PMID: 11226425
  31. HIFalpha targeted for VHL-mediated destruction by proline hydroxylation: implications for O2 sensing.
    Science. 2001 Apr 20;292(5516):464-8 PMID: 11292862
  32. Targeting of HIF-alpha to the von Hippel-Lindau ubiquitylation complex by O2-regulated prolyl hydroxylation.
    Science. 2001 Apr 20;292(5516):468-72 PMID: 11292861
  33. SM-20 is a novel mitochondrial protein that causes caspase-dependent cell death in nerve growth factor-dependent neurons.
    J Biol Chem. 2001 Feb 16;276(7):5085-92 PMID: 11060309
  34. Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells.
    Nature. 2001 May 24;411(6836):494-8 PMID: 11373684
  35. A conserved family of prolyl-4-hydroxylases that modify HIF.
    Science. 2001 Nov 9;294(5545):1337-40 PMID: 11598268
  36. Asparagine hydroxylation of the HIF transactivation domain a hypoxic switch.
    Science. 2002 Feb 1;295(5556):858-61 PMID: 11823643
  37. Hypoxia-inducible factor-1alpha mRNA contains an internal ribosome entry site that allows efficient translation during normoxia and hypoxia.
    Mol Biol Cell. 2002 May;13(5):1792-801 PMID: 12006670
  38. FIH-1 is an asparaginyl hydroxylase enzyme that regulates the transcriptional activity of hypoxia-inducible factor.
    Genes Dev. 2002 Jun 15;16(12):1466-71 PMID: 12080085
  39. Hypoxia-inducible factor (HIF) asparagine hydroxylase is identical to factor inhibiting HIF (FIH) and is related to the cupin structural family.
    J Biol Chem. 2002 Jul 19;277(29):26351-5 PMID: 12042299
  40. Overexpression of PH-4, a novel putative proline 4-hydroxylase, modulates activity of hypoxia-inducible transcription factors.
    Biochem Biophys Res Commun. 2002 Aug 16;296(2):343-9 PMID: 12163023
  41. Mammalian EGLN genes have distinct patterns of mRNA expression and regulation.
    Biochem Cell Biol. 2002;80(4):421-6 PMID: 12234095
  42. Sequence determinants in hypoxia-inducible factor-1alpha for hydroxylation by the prolyl hydroxylases PHD1, PHD2, and PHD3.
    J Biol Chem. 2002 Oct 18;277(42):39792-800 PMID: 12181324
  43. Role of prolyl hydroxylation in oncogenically stabilized hypoxia-inducible factor-1alpha.
    J Biol Chem. 2002 Oct 18;277(42):40112-7 PMID: 12186875
  44. Biochemical purification and pharmacological inhibition of a mammalian prolyl hydroxylase acting on hypoxia-inducible factor.
    Proc Natl Acad Sci U S A. 2002 Oct 15;99(21):13459-64 PMID: 12351678
  45. Hypoxia-inducible factor asparaginyl hydroxylase (FIH-1) catalyses hydroxylation at the beta-carbon of asparagine-803.
    Biochem J. 2002 Nov 1;367(Pt 3):571-5 PMID: 12215170
  46. Regulation and destabilization of HIF-1alpha by ARD1-mediated acetylation.
    Cell. 2002 Nov 27;111(5):709-20 PMID: 12464182
  47. von Hippel-Lindau protein binds hyperphosphorylated large subunit of RNA polymerase II through a proline hydroxylation motif and targets it for ubiquitination.
    Proc Natl Acad Sci U S A. 2003 Mar 4;100(5):2706-11 PMID: 12604794
  48. Differential regulation of HIF-1 alpha prolyl-4-hydroxylase genes by hypoxia in human cardiovascular cells.
    Biochem Biophys Res Commun. 2003 Apr 11;303(3):947-53 PMID: 12670503
  49. Cloning of a factor required for activity of the Ah (dioxin) receptor.
    Science. 1991 May 17;252(5008):954-8 PMID: 1852076
Article Info
Journal
The EMBO journal
Abbr.
EMBO J
ISSN
0261-4189
DOI
10.1093/emboj/cdg392
Email
[email protected]
Published
2003-08-15
Pages
4082-90
Language
English
Region
England
NLM ID
8208664
PMCID
PMC175782
Subset
IM
External Links
PubMed source DOI: 10.1093/emboj/cdg392 PMC Full Text View journal details
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