title: Destruction of a distal hypoxia response element abolishes trans-activation of the PAG1 gene mediated by HIF-independent chromatin looping.
abstract: A crucial step in the cellular adaptation to oxygen deficiency is the binding of hypoxia-inducible factors (HIFs) to hypoxia response elements (HREs) of oxygen-regulated genes. Genome-wide HIF-1α/2α/β DNA-binding studies revealed that the majority of HREs reside distant to the promoter regions, but the function of these distal HREs has only been marginally studied in the genomic context. We used chromatin immunoprecipitation (ChIP), gene editing (TALEN) and chromosome conformation capture (3C) to localize and functionally characterize a 82 kb upstream HRE that solely drives oxygen-regulated expression of the newly identified HIF target gene PAG1. PAG1, a transmembrane adaptor protein involved in Src signalling, was hypoxically induced in various cell lines and mouse tissues. ChIP and reporter gene assays demonstrated that the -82 kb HRE regulates PAG1, but not an equally distant gene further upstream, by direct interaction with HIF. Ablation of the consensus HRE motif abolished the hypoxic induction of PAG1 but not general oxygen signalling. 3C assays revealed that the -82 kb HRE physically associates with the PAG1 promoter region, independent of HIF-DNA interaction. These results demonstrate a constitutive interaction between the -82 kb HRE and the PAG1 promoter, suggesting a physiologically important rapid response to hypoxia.
authors.affiliation: Institute of Physiology and Zürich Center for Integrative Human Physiology ZIHP, University of Zürich, CH-8057 Zürich, Switzerland.
authors.forename: Alexandra
authors.initials: A
authors.lastname: Schörg
authors.affiliation: Institute of Physiology and Zürich Center for Integrative Human Physiology ZIHP, University of Zürich, CH-8057 Zürich, Switzerland.
authors.forename: Sara
authors.initials: S
authors.lastname: Santambrogio
authors.affiliation: Henry Wellcome Building for Molecular Physiology, University of Oxford, Ox3 7BN, UK.
authors.forename: James L
authors.initials: JL
authors.lastname: Platt
authors.affiliation: Department of Nephrology and Hypertension, Friedrich-Alexander-University Erlangen-Nuremberg, D-91054 Erlangen, Germany.
authors.forename: Johannes
authors.initials: J
authors.lastname: Schödel
authors.affiliation: Institute of Physiology and Zürich Center for Integrative Human Physiology ZIHP, University of Zürich, CH-8057 Zürich, Switzerland.
authors.forename: Maja T
authors.initials: MT
authors.lastname: Lindenmeyer
authors.affiliation: Institute of Physiology and Zürich Center for Integrative Human Physiology ZIHP, University of Zürich, CH-8057 Zürich, Switzerland National Center of Competence in Research "Kidney.CH", Switzerland.
authors.forename: Clemens D
authors.initials: CD
authors.lastname: Cohen
authors.affiliation: Institute of Physiology, University of Duisburg-Essen, D-45122 Essen, Germany.
authors.forename: Katrin
authors.initials: K
authors.lastname: Schrödter
authors.affiliation: Henry Wellcome Building for Molecular Physiology, University of Oxford, Ox3 7BN, UK.
authors.forename: David R
authors.initials: DR
authors.lastname: Mole
authors.affiliation: Institute of Physiology and Zürich Center for Integrative Human Physiology ZIHP, University of Zürich, CH-8057 Zürich, Switzerland National Center of Competence in Research "Kidney.CH", Switzerland roland.wenger@access.uzh.ch.
authors.forename: Roland H
authors.initials: RH
authors.lastname: Wenger
authors.affiliation: Institute of Physiology and Zürich Center for Integrative Human Physiology ZIHP, University of Zürich, CH-8057 Zürich, Switzerland National Center of Competence in Research "Kidney.CH", Switzerland Institute of Physiology, University of Duisburg-Essen, D-45122 Essen, Germany david.hoogewijs@uni-due.de.
authors.forename: David
authors.initials: D
authors.lastname: Hoogewijs
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000096:biosynthesis / Q000235:genetics*; D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins / Q000378:metabolism*; D015687:Cell Hypoxia; D002460:Cell Line; D002843:Chromatin / Q000737:chemistry / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D051793:Hypoxia-Inducible Factor 1 / Q000378:metabolism*; D008565:Membrane Proteins / Q000096:biosynthesis / Q000235:genetics*; D051379:Mice; D008810:Mice, Inbred C57BL; D010750:Phosphoproteins / Q000096:biosynthesis / Q000235:genetics; D011401:Promoter Regions, Genetic; D020218:Response Elements; D015398:Signal Transduction; D015533:Transcriptional Activation; D019061:src-Family Kinases / Q000378:metabolism; D000099162:Endothelial PAS Domain-Containing Protein 1
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D051792:Basic Helix-Loop-Helix Proteins; D002843:Chromatin; D051793:Hypoxia-Inducible Factor 1; D008565:Membrane Proteins; D010750:Phosphoproteins; D019061:src-Family Kinases; D000099162:Endothelial PAS Domain-Containing Protein 1; C498710:PAG1 protein, human; C498709:Pag1 protein, mouse