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Mol Cell Biol. Oct 1996; 16(10): 5221–5231.
PMCID: PMC231522

Functional interference between hypoxia and dioxin signal transduction pathways: competition for recruitment of the Arnt transcription factor.


Hypoxia-inducible factor 1 alpha (HIF-1 alpha) and the intracellular dioxin receptor mediate hypoxia and dioxin signalling, respectively. Both proteins are conditionally regulated basic helix-loop-helix (bHLH) transcription factors that, in addition to the bHLH motif, share a Per-Arnt-Sim (PAS) region of homology and form heterodimeric complexes with the common bHLH/PAS partner factor Arnt. Here we demonstrate that HIF-1 alpha required Arnt for DNA binding in vitro and functional activity in vivo. Both the bHLH and PAS motifs of Arnt were critical for dimerization with HIF-1 alpha. Strikingly, HIF-1 alpha exhibited very high affinity for Arnt in coimmunoprecipitation assays in vitro, resulting in competition with the ligand-activated dioxin receptor for recruitment of Arnt. Consistent with these observations, activation of HIF-1 alpha function in vivo or overexpression of HIF-1 alpha inhibited ligand-dependent induction of DNA binding activity by the dioxin receptor and dioxin receptor function on minimal reporter gene constructs. However, HIF-1 alpha- and dioxin receptor-mediated signalling pathways were not mutually exclusive, since activation of dioxin receptor function did not impair HIF-1 alpha-dependent induction of target gene expression. Both HIF-1 alpha and Arnt mRNAs were expressed constitutively in a large number of human tissues and cell lines, and these steady-state expression levels were not affected by exposure to hypoxia. Thus, HIF-1 alpha may be conditionally regulated by a mechanism that is distinct from induced expression levels, the prevalent model of activation of HIF-1 alpha function. Interestingly, we observed that HIF-1 alpha was associated with the molecular chaperone hsp90. Given the critical role of hsp90 for ligand binding activity and activation of the dioxin receptor, it is therefore possible that HIF-1 alpha is regulated by a similar mechanism, possibly by binding an as yet unknown class of ligands.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Andersson S, Davis DL, Dahlbäck H, Jörnvall H, Russell DW. Cloning, structure, and expression of the mitochondrial cytochrome P-450 sterol 26-hydroxylase, a bile acid biosynthetic enzyme. J Biol Chem. 1989 May 15;264(14):8222–8229. [PubMed]
  • Antonsson C, Arulampalam V, Whitelaw ML, Pettersson S, Poellinger L. Constitutive function of the basic helix-loop-helix/PAS factor Arnt. Regulation of target promoters via the E box motif. J Biol Chem. 1995 Jun 9;270(23):13968–13972. [PubMed]
  • Antonsson C, Whitelaw ML, McGuire J, Gustafsson JA, Poellinger L. Distinct roles of the molecular chaperone hsp90 in modulating dioxin receptor function via the basic helix-loop-helix and PAS domains. Mol Cell Biol. 1995 Feb;15(2):756–765. [PMC free article] [PubMed]
  • Berghard A, Gradin K, Pongratz I, Whitelaw M, Poellinger L. Cross-coupling of signal transduction pathways: the dioxin receptor mediates induction of cytochrome P-450IA1 expression via a protein kinase C-dependent mechanism. Mol Cell Biol. 1993 Jan;13(1):677–689. [PMC free article] [PubMed]
  • Bohen SP, Kralli A, Yamamoto KR. Hold 'em and fold 'em: chaperones and signal transduction. Science. 1995 Jun 2;268(5215):1303–1304. [PubMed]
  • Burbach KM, Poland A, Bradfield CA. Cloning of the Ah-receptor cDNA reveals a distinctive ligand-activated transcription factor. Proc Natl Acad Sci U S A. 1992 Sep 1;89(17):8185–8189. [PMC free article] [PubMed]
  • Carver LA, Hogenesch JB, Bradfield CA. Tissue specific expression of the rat Ah-receptor and ARNT mRNAs. Nucleic Acids Res. 1994 Aug 11;22(15):3038–3044. [PMC free article] [PubMed]
  • Carver LA, Jackiw V, Bradfield CA. The 90-kDa heat shock protein is essential for Ah receptor signaling in a yeast expression system. J Biol Chem. 1994 Dec 2;269(48):30109–30112. [PubMed]
  • Chomczynski P, Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 1987 Apr;162(1):156–159. [PubMed]
  • Coumailleau P, Poellinger L, Gustafsson JA, Whitelaw ML. Definition of a minimal domain of the dioxin receptor that is associated with Hsp90 and maintains wild type ligand binding affinity and specificity. J Biol Chem. 1995 Oct 20;270(42):25291–25300. [PubMed]
  • Dahmane N, Charron G, Lopes C, Yaspo ML, Maunoury C, Decorte L, Sinet PM, Bloch B, Delabar JM. Down syndrome-critical region contains a gene homologous to Drosophila sim expressed during rat and human central nervous system development. Proc Natl Acad Sci U S A. 1995 Sep 26;92(20):9191–9195. [PMC free article] [PubMed]
  • Fernandez-Salguero P, Pineau T, Hilbert DM, McPhail T, Lee SS, Kimura S, Nebert DW, Rudikoff S, Ward JM, Gonzalez FJ. Immune system impairment and hepatic fibrosis in mice lacking the dioxin-binding Ah receptor. Science. 1995 May 5;268(5211):722–726. [PubMed]
  • Firth JD, Ebert BL, Pugh CW, Ratcliffe PJ. Oxygen-regulated control elements in the phosphoglycerate kinase 1 and lactate dehydrogenase A genes: similarities with the erythropoietin 3' enhancer. Proc Natl Acad Sci U S A. 1994 Jul 5;91(14):6496–6500. [PMC free article] [PubMed]
  • Goldberg MA, Schneider TJ. Similarities between the oxygen-sensing mechanisms regulating the expression of vascular endothelial growth factor and erythropoietin. J Biol Chem. 1994 Feb 11;269(6):4355–4359. [PubMed]
  • Gradin K, Wilhelmsson A, Poellinger L, Berghard A. Nonresponsiveness of normal human fibroblasts to dioxin correlates with the presence of a constitutive xenobiotic response element-binding factor. J Biol Chem. 1993 Feb 25;268(6):4061–4068. [PubMed]
  • Gregor PD, Sawadogo M, Roeder RG. The adenovirus major late transcription factor USF is a member of the helix-loop-helix group of regulatory proteins and binds to DNA as a dimer. Genes Dev. 1990 Oct;4(10):1730–1740. [PubMed]
  • Hankinson O. The aryl hydrocarbon receptor complex. Annu Rev Pharmacol Toxicol. 1995;35:307–340. [PubMed]
  • Hirose K, Morita M, Ema M, Mimura J, Hamada H, Fujii H, Saijo Y, Gotoh O, Sogawa K, Fujii-Kuriyama Y. cDNA cloning and tissue-specific expression of a novel basic helix-loop-helix/PAS factor (Arnt2) with close sequence similarity to the aryl hydrocarbon receptor nuclear translocator (Arnt). Mol Cell Biol. 1996 Apr;16(4):1706–1713. [PMC free article] [PubMed]
  • Hoffman EC, Reyes H, Chu FF, Sander F, Conley LH, Brooks BA, Hankinson O. Cloning of a factor required for activity of the Ah (dioxin) receptor. Science. 1991 May 17;252(5008):954–958. [PubMed]
  • Hord NG, Perdew GH. Physicochemical and immunocytochemical analysis of the aryl hydrocarbon receptor nuclear translocator: characterization of two monoclonal antibodies to the aryl hydrocarbon receptor nuclear translocator. Mol Pharmacol. 1994 Oct;46(4):618–626. [PubMed]
  • Ikeda E, Achen MG, Breier G, Risau W. Hypoxia-induced transcriptional activation and increased mRNA stability of vascular endothelial growth factor in C6 glioma cells. J Biol Chem. 1995 Aug 25;270(34):19761–19766. [PubMed]
  • Isaac DD, Andrew DJ. Tubulogenesis in Drosophila: a requirement for the trachealess gene product. Genes Dev. 1996 Jan 1;10(1):103–117. [PubMed]
  • Jain S, Dolwick KM, Schmidt JV, Bradfield CA. Potent transactivation domains of the Ah receptor and the Ah receptor nuclear translocator map to their carboxyl termini. J Biol Chem. 1994 Dec 16;269(50):31518–31524. [PubMed]
  • Kleman MI, Poellinger L, Gustafsson JA. Regulation of human dioxin receptor function by indolocarbazoles, receptor ligands of dietary origin. J Biol Chem. 1994 Feb 18;269(7):5137–5144. [PubMed]
  • Kvietikova I, Wenger RH, Marti HH, Gassmann M. The transcription factors ATF-1 and CREB-1 bind constitutively to the hypoxia-inducible factor-1 (HIF-1) DNA recognition site. Nucleic Acids Res. 1995 Nov 25;23(22):4542–4550. [PMC free article] [PubMed]
  • Levy AP, Levy NS, Goldberg MA. Post-transcriptional regulation of vascular endothelial growth factor by hypoxia. J Biol Chem. 1996 Feb 2;271(5):2746–2753. [PubMed]
  • Levy AP, Levy NS, Wegner S, Goldberg MA. Transcriptional regulation of the rat vascular endothelial growth factor gene by hypoxia. J Biol Chem. 1995 Jun 2;270(22):13333–13340. [PubMed]
  • Lindebro MC, Poellinger L, Whitelaw ML. Protein-protein interaction via PAS domains: role of the PAS domain in positive and negative regulation of the bHLH/PAS dioxin receptor-Arnt transcription factor complex. EMBO J. 1995 Jul 17;14(14):3528–3539. [PMC free article] [PubMed]
  • Littlewood TD, Evan GI. Transcription factors 2: helix-loop-helix. Protein Profile. 1995;2(6):621–702. [PubMed]
  • Liu Y, Cox SR, Morita T, Kourembanas S. Hypoxia regulates vascular endothelial growth factor gene expression in endothelial cells. Identification of a 5' enhancer. Circ Res. 1995 Sep;77(3):638–643. [PubMed]
  • Ma PC, Rould MA, Weintraub H, Pabo CO. Crystal structure of MyoD bHLH domain-DNA complex: perspectives on DNA recognition and implications for transcriptional activation. Cell. 1994 May 6;77(3):451–459. [PubMed]
  • Mangelsdorf DJ, Evans RM. The RXR heterodimers and orphan receptors. Cell. 1995 Dec 15;83(6):841–850. [PubMed]
  • Mason GG, Witte AM, Whitelaw ML, Antonsson C, McGuire J, Wilhelmsson A, Poellinger L, Gustafsson JA. Purification of the DNA binding form of dioxin receptor. Role of the Arnt cofactor in regulation of dioxin receptor function. J Biol Chem. 1994 Feb 11;269(6):4438–4449. [PubMed]
  • Matsushita N, Sogawa K, Ema M, Yoshida A, Fujii-Kuriyama Y. A factor binding to the xenobiotic responsive element (XRE) of P-4501A1 gene consists of at least two helix-loop-helix proteins, Ah receptor and Arnt. J Biol Chem. 1993 Oct 5;268(28):21002–21006. [PubMed]
  • McGuire J, Coumailleau P, Whitelaw ML, Gustafsson JA, Poellinger L. The basic helix-loop-helix/PAS factor Sim is associated with hsp90. Implications for regulation by interaction with partner factors. J Biol Chem. 1995 Dec 29;270(52):31353–31357. [PubMed]
  • McGuire J, Whitelaw ML, Pongratz I, Gustafsson JA, Poellinger L. A cellular factor stimulates ligand-dependent release of hsp90 from the basic helix-loop-helix dioxin receptor. Mol Cell Biol. 1994 Apr;14(4):2438–2446. [PMC free article] [PubMed]
  • Miyata Y, Yahara I. The 90-kDa heat shock protein, HSP90, binds and protects casein kinase II from self-aggregation and enhances its kinase activity. J Biol Chem. 1992 Apr 5;267(10):7042–7047. [PubMed]
  • Nordeen SK. Luciferase reporter gene vectors for analysis of promoters and enhancers. Biotechniques. 1988 May;6(5):454–458. [PubMed]
  • Perdew GH, Whitelaw ML. Evidence that the 90-kDa heat shock protein (HSP90) exists in cytosol in heteromeric complexes containing HSP70 and three other proteins with Mr of 63,000, 56,000, and 50,000. J Biol Chem. 1991 Apr 15;266(11):6708–6713. [PubMed]
  • Picard D, Khursheed B, Garabedian MJ, Fortin MG, Lindquist S, Yamamoto KR. Reduced levels of hsp90 compromise steroid receptor action in vivo. Nature. 1990 Nov 8;348(6297):166–168. [PubMed]
  • Poellinger L, Göttlicher M, Gustafsson JA. The dioxin and peroxisome proliferator-activated receptors: nuclear receptors in search of endogenous ligands. Trends Pharmacol Sci. 1992 Jun;13(6):241–245. [PubMed]
  • Pollenz RS, Sattler CA, Poland A. The aryl hydrocarbon receptor and aryl hydrocarbon receptor nuclear translocator protein show distinct subcellular localizations in Hepa 1c1c7 cells by immunofluorescence microscopy. Mol Pharmacol. 1994 Mar;45(3):428–438. [PubMed]
  • Pongratz I, Mason GG, Poellinger L. Dual roles of the 90-kDa heat shock protein hsp90 in modulating functional activities of the dioxin receptor. Evidence that the dioxin receptor functionally belongs to a subclass of nuclear receptors which require hsp90 both for ligand binding activity and repression of intrinsic DNA binding activity. J Biol Chem. 1992 Jul 5;267(19):13728–13734. [PubMed]
  • Pratt WB. The role of heat shock proteins in regulating the function, folding, and trafficking of the glucocorticoid receptor. J Biol Chem. 1993 Oct 15;268(29):21455–21458. [PubMed]
  • Rannug U, Rannug A, Sjöberg U, Li H, Westerholm R, Bergman J. Structure elucidation of two tryptophan-derived, high affinity Ah receptor ligands. Chem Biol. 1995 Dec;2(12):841–845. [PubMed]
  • Reyes H, Reisz-Porszasz S, Hankinson O. Identification of the Ah receptor nuclear translocator protein (Arnt) as a component of the DNA binding form of the Ah receptor. Science. 1992 May 22;256(5060):1193–1195. [PubMed]
  • Semenza GL, Roth PH, Fang HM, Wang GL. Transcriptional regulation of genes encoding glycolytic enzymes by hypoxia-inducible factor 1. J Biol Chem. 1994 Sep 23;269(38):23757–23763. [PubMed]
  • Semenza GL, Wang GL. A nuclear factor induced by hypoxia via de novo protein synthesis binds to the human erythropoietin gene enhancer at a site required for transcriptional activation. Mol Cell Biol. 1992 Dec;12(12):5447–5454. [PMC free article] [PubMed]
  • Shima DT, Deutsch U, D'Amore PA. Hypoxic induction of vascular endothelial growth factor (VEGF) in human epithelial cells is mediated by increases in mRNA stability. FEBS Lett. 1995 Aug 21;370(3):203–208. [PubMed]
  • Smith DF, Toft DO. Steroid receptors and their associated proteins. Mol Endocrinol. 1993 Jan;7(1):4–11. [PubMed]
  • Sogawa K, Nakano R, Kobayashi A, Kikuchi Y, Ohe N, Matsushita N, Fujii-Kuriyama Y. Possible function of Ah receptor nuclear translocator (Arnt) homodimer in transcriptional regulation. Proc Natl Acad Sci U S A. 1995 Mar 14;92(6):1936–1940. [PMC free article] [PubMed]
  • Swanson HI, Chan WK, Bradfield CA. DNA binding specificities and pairing rules of the Ah receptor, ARNT, and SIM proteins. J Biol Chem. 1995 Nov 3;270(44):26292–26302. [PubMed]
  • Wang GL, Jiang BH, Rue EA, Semenza GL. 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):5510–5514. [PMC free article] [PubMed]
  • Wang GL, Semenza GL. Characterization of hypoxia-inducible factor 1 and regulation of DNA binding activity by hypoxia. J Biol Chem. 1993 Oct 15;268(29):21513–21518. [PubMed]
  • Weintraub H. The MyoD family and myogenesis: redundancy, networks, and thresholds. Cell. 1993 Dec 31;75(7):1241–1244. [PubMed]
  • Wenger RH, Marti HH, Schuerer-Maly CC, Kvietikova I, Bauer C, Gassmann M, Maly FE. Hypoxic induction of gene expression in chronic granulomatous disease-derived B-cell lines: oxygen sensing is independent of the cytochrome b558-containing nicotinamide adenine dinucleotide phosphate oxidase. Blood. 1996 Jan 15;87(2):756–761. [PubMed]
  • Wenger RH, Rolfs A, Marti HH, Bauer C, Gassmann M. Hypoxia, a novel inducer of acute phase gene expression in a human hepatoma cell line. J Biol Chem. 1995 Nov 17;270(46):27865–27870. [PubMed]
  • Wenger RH, Rolfs A, Marti HH, Guénet JL, Gassmann M. Nucleotide sequence, chromosomal assignment and mRNA expression of mouse hypoxia-inducible factor-1 alpha. Biochem Biophys Res Commun. 1996 Jun 5;223(1):54–59. [PubMed]
  • Wharton KA, Jr, Franks RG, Kasai Y, Crews ST. Control of CNS midline transcription by asymmetric E-box-like elements: similarity to xenobiotic responsive regulation. Development. 1994 Dec;120(12):3563–3569. [PubMed]
  • Whitelaw ML, Göttlicher M, Gustafsson JA, Poellinger L. Definition of a novel ligand binding domain of a nuclear bHLH receptor: co-localization of ligand and hsp90 binding activities within the regulable inactivation domain of the dioxin receptor. EMBO J. 1993 Nov;12(11):4169–4179. [PMC free article] [PubMed]
  • Whitelaw ML, Gustafsson JA, Poellinger L. Identification of transactivation and repression functions of the dioxin receptor and its basic helix-loop-helix/PAS partner factor Arnt: inducible versus constitutive modes of regulation. Mol Cell Biol. 1994 Dec;14(12):8343–8355. [PMC free article] [PubMed]
  • Whitelaw ML, McGuire J, Picard D, Gustafsson JA, Poellinger L. Heat shock protein hsp90 regulates dioxin receptor function in vivo. Proc Natl Acad Sci U S A. 1995 May 9;92(10):4437–4441. [PMC free article] [PubMed]
  • Whitelaw M, Pongratz I, Wilhelmsson A, Gustafsson JA, Poellinger L. Ligand-dependent recruitment of the Arnt coregulator determines DNA recognition by the dioxin receptor. Mol Cell Biol. 1993 Apr;13(4):2504–2514. [PMC free article] [PubMed]
  • Wiech H, Buchner J, Zimmermann R, Jakob U. Hsp90 chaperones protein folding in vitro. Nature. 1992 Jul 9;358(6382):169–170. [PubMed]
  • Wilhelmsson A, Cuthill S, Denis M, Wikström AC, Gustafsson JA, Poellinger L. The specific DNA binding activity of the dioxin receptor is modulated by the 90 kd heat shock protein. EMBO J. 1990 Jan;9(1):69–76. [PMC free article] [PubMed]
  • Wilk R, Weizman I, Shilo BZ. trachealess encodes a bHLH-PAS protein that is an inducer of tracheal cell fates in Drosophila. Genes Dev. 1996 Jan 1;10(1):93–102. [PubMed]

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