title: Synthetic transactivation screening reveals ETV4 as broad coactivator of hypoxia-inducible factor signaling.
abstract: The human prolyl-4-hydroxylase domain (PHD) proteins 1-3 are known as cellular oxygen sensors, acting via the degradation of hypoxia-inducible factor (HIF) α-subunits. PHD2 and PHD3 genes are inducible by HIFs themselves, suggesting a negative feedback loop that involves PHD abundance. To identify novel regulators of the PHD2 gene, an expression array of 704 transcription factors was screened by a method that allows distinguishing between HIF-dependent and HIF-independent promoter regulation. Among others, the E-twenty six transcription factor ETS translocation variant 4 (ETV4) was found to contribute to PHD2 gene expression particularly under hypoxic conditions. Mechanistically, complex formation between ETV4 and HIF-1/2α was observed by mammalian two-hybrid and fluorescence resonance energy transfer analysis. HIF-1α domain mapping, CITED2 overexpression and factor inhibiting HIF depletion experiments provided evidence for cooperation between HIF-1α and p300/CBP in ETV4 binding. Chromatin immunoprecipitation confirmed ETV4 and HIF-1α corecruitment to the PHD2 promoter. Of 608 hypoxically induced transcripts found by genome-wide expression profiling, 7.7% required ETV4 for efficient hypoxic induction, suggesting a broad role of ETV4 in hypoxic gene regulation. Endogenous ETV4 highly correlated with PHD2, HIF-1/2α and several established markers of tissue hypoxia in 282 human breast cancer tissue samples, corroborating a functional interplay between the ETV4 and HIF pathways.
authors.affiliation: Institute of Physiology and Zürich Center for Integrative Human Physiology (ZIHP), University of Zürich, 8057 Zürich, Switzerland.
authors.forename: Kristin
authors.initials: K
authors.lastname: Wollenick
authors.forename: Jun
authors.initials: J
authors.lastname: Hu
authors.forename: Glen
authors.initials: G
authors.lastname: Kristiansen
authors.forename: Peter
authors.initials: P
authors.lastname: Schraml
authors.forename: Hubert
authors.initials: H
authors.lastname: Rehrauer
authors.forename: Utta
authors.initials: U
authors.lastname: Berchner-Pfannschmidt
authors.forename: Joachim
authors.initials: J
authors.lastname: Fandrey
authors.forename: Roland H
authors.initials: RH
authors.lastname: Wenger
authors.forename: Daniel P
authors.initials: DP
authors.lastname: Stiehl
mesh_terms: D017500:Adenovirus E1A Proteins / Q000378:metabolism*; D001665:Binding Sites; D001943:Breast Neoplasms / Q000378:metabolism; D045744:Cell Line, Tumor; D005260:Female; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000737:chemistry / Q000378:metabolism*; D064799:Hypoxia-Inducible Factor-Proline Dioxygenases; D011393:Procollagen-Proline Dioxygenase / Q000235:genetics*; D011401:Promoter Regions, Genetic; D017434:Protein Structure, Tertiary; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D050783:Proto-Oncogene Proteins c-ets / Q000378:metabolism; D016760:Proto-Oncogene Proteins c-fos / Q000378:metabolism; D015398:Signal Transduction; D015533:Transcriptional Activation; D014168:Transferrin / Q000235:genetics; D050880:p300-CBP Transcription Factors / Q000378:metabolism
chemical_list: D017500:Adenovirus E1A Proteins; C079853:ETV4 protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D011518:Proto-Oncogene Proteins; D050783:Proto-Oncogene Proteins c-ets; D016760:Proto-Oncogene Proteins c-fos; D014168:Transferrin; C490190:EGLN1 protein, human; D011393:Procollagen-Proline Dioxygenase; D064799:Hypoxia-Inducible Factor-Proline Dioxygenases; D050880:p300-CBP Transcription Factors
references.citation: Nucleic Acids Res. 2010 Apr;38(7):2332-45
references.pmid: 20061373
references.citation: Pflugers Arch. 2005 Sep;450(6):363-71
references.pmid: 16007431
references.citation: Oncogene. 2012 May 3;31(18):2283-97
references.pmid: 21927022
references.citation: J Biol Chem. 2006 Aug 18;281(33):23482-91
references.pmid: 16790428
references.citation: J Biol Chem. 1997 Aug 8;272(32):20055-62
references.pmid: 9242677
references.citation: EMBO J. 2003 Aug 15;22(16):4082-90
references.pmid: 12912907
references.citation: Nat Struct Biol. 2003 Jul;10(7):504-12
references.pmid: 12778114
references.citation: Nat Genet. 2009 Dec;41(12):1295-302
references.pmid: 19898483
references.citation: Nat Genet. 2009 May;41(5):509-10
references.pmid: 19399032
references.citation: Biochem J. 2005 May 1;387(Pt 3):711-7
references.pmid: 15563275
references.citation: Circulation. 2007 Aug 14;116(7):774-81
references.pmid: 17646578
references.citation: Nat Med. 2010 Jun;16(6):687-93
references.pmid: 20495569
references.citation: Oncogene. 2010 Feb 4;29(5):625-34
references.pmid: 19946328
references.citation: Blood. 2007 Nov 15;110(10):3610-7
references.pmid: 17684156
references.citation: Nature. 2001 Nov 29;414(6863):550-4
references.pmid: 11734856
references.citation: Curr Biol. 2001 Nov 13;11(22):1739-48
references.pmid: 11719215
references.citation: J Biol Chem. 2004 Apr 30;279(18):18694-700
references.pmid: 14976201
references.citation: Cancer Cell. 2009 Jun 2;15(6):527-38
references.pmid: 19477431
references.citation: J Biol Chem. 2009 Aug 21;284(34):23046-58
references.pmid: 19546213
references.citation: J Biol Chem. 2010 Jan 22;285(4):2601-9
references.pmid: 19880525
references.citation: Genes Dev. 2001 Oct 15;15(20):2675-86
references.pmid: 11641274
references.citation: Cell Growth Differ. 2001 Jul;12(7):363-9
references.pmid: 11457733
references.citation: Am J Physiol. 1998 Feb;274(2):L212-9
references.pmid: 9486205
references.citation: J Cell Sci. 2005 Jan 15;118(Pt 2):301-11
references.pmid: 15615775
references.citation: Blood. 2011 May 19;117(20):5485-93
references.pmid: 21346252
references.citation: Nat Rev Mol Cell Biol. 2001 Nov;2(11):827-37
references.pmid: 11715049
references.citation: Mol Cell. 2008 May 23;30(4):393-402
references.pmid: 18498744
references.citation: Biochim Biophys Acta. 2006 Aug;1766(1):79-87
references.pmid: 16546322
references.citation: Proc Natl Acad Sci U S A. 2008 Mar 25;105(12):4745-50
references.pmid: 18347341
references.citation: FASEB J. 2007 Mar;21(3):700-7
references.pmid: 17197389
references.citation: Epigenetics. 2010 Jan 1;5(1):9-15
references.pmid: 20110770
references.citation: Nat Rev Cancer. 2008 Jul;8(7):497-511
references.pmid: 18563191
references.citation: Genes Dev. 1999 Jan 1;13(1):64-75
references.pmid: 9887100
references.citation: Proc Natl Acad Sci U S A. 2007 Aug 7;104(32):12955-61
references.pmid: 17670940
references.citation: Cancer Res. 2006 Jul 1;66(13):6683-91
references.pmid: 16818642
references.citation: Genes Dev. 2002 Jun 15;16(12):1466-71
references.pmid: 12080085
references.citation: Mol Cell Biol. 2006 Nov;26(22):8336-46
references.pmid: 16966370
references.citation: Oncogene. 2011 Feb 3;30(5):548-60
references.pmid: 20856199
references.citation: J Biol Chem. 2009 Jun 19;284(25):16767-16775
references.pmid: 19386601
references.citation: Development. 2010 Jun;137(12):1975-9
references.pmid: 20463033
references.citation: Oncol Rep. 2006 Jul;16(1):153-8
references.pmid: 16786139
references.citation: J Biol Chem. 1996 Dec 20;271(51):32529-37
references.pmid: 8955077
references.citation: Genes Dev. 1992 Mar;6(3):481-96
references.pmid: 1547944
references.citation: J Biol Chem. 2009 Sep 4;284(36):24204-12
references.pmid: 19564335
references.citation: J Biol Chem. 1997 Apr 25;272(17):11205-14
references.pmid: 9111021
references.citation: Proc Natl Acad Sci U S A. 1996 Nov 12;93(23):12969-73
references.pmid: 8917528
references.citation: J Biol Chem. 2002 Jul 19;277(29):26351-5
references.pmid: 12042299
references.citation: J Biol Chem. 2005 Jan 14;280(2):887-98
references.pmid: 15466854
references.citation: Mol Cell Biol. 2002 May;22(9):2984-92
references.pmid: 11940656
references.citation: Nature. 2001 May 3;411(6833):107-10
references.pmid: 11333987
references.citation: Dev Biol. 2009 Jan 1;325(1):106-21
references.pmid: 18977342
references.citation: Oncogene. 2010 Mar 25;29(12):1810-20
references.pmid: 20062082
references.citation: Oncogene. 1997 Sep 25;15(13):1513-25
references.pmid: 9380403
references.citation: Nature. 2011 May 15;474(7351):403-6
references.pmid: 21572435
references.citation: Sci STKE. 2005 Oct 18;2005(306):re12
references.pmid: 16234508
references.citation: Neuron. 2002 Aug 29;35(5):877-92
references.pmid: 12372283
references.citation: J Biol Chem. 2001 Jun 8;276(23):20108-15
references.pmid: 11274170
references.citation: Mol Cell Biol. 2007 May;27(10):3758-68
references.pmid: 17353276
references.citation: Cell. 2009 Mar 6;136(5):839-851
references.pmid: 19217150
references.citation: EMBO Rep. 2011 Jan;12(1):63-70
references.pmid: 21109780