title: Disruptions of topological chromatin domains cause pathogenic rewiring of gene-enhancer interactions.
abstract: Mammalian genomes are organized into megabase-scale topologically associated domains (TADs). We demonstrate that disruption of TADs can rewire long-range regulatory architecture and result in pathogenic phenotypes. We show that distinct human limb malformations are caused by deletions, inversions, or duplications altering the structure of the TAD-spanning WNT6/IHH/EPHA4/PAX3 locus. Using CRISPR/Cas genome editing, we generated mice with corresponding rearrangements. Both in mouse limb tissue and patient-derived fibroblasts, disease-relevant structural changes cause ectopic interactions between promoters and non-coding DNA, and a cluster of limb enhancers normally associated with Epha4 is misplaced relative to TAD boundaries and drives ectopic limb expression of another gene in the locus. This rewiring occurred only if the variant disrupted a CTCF-associated boundary domain. Our results demonstrate the functional importance of TADs for orchestrating gene expression via genome architecture and indicate criteria for predicting the pathogenicity of human structural variants, particularly in non-coding regions of the human genome.
authors.affiliation: Max Planck Institute for Molecular Genetics, RG Development & Disease, 14195 Berlin, Germany; Institute for Medical and Human Genetics, Charité Universitätsmedizin Berlin, 13353 Berlin, Germany.
authors.forename: Darío G
authors.initials: DG
authors.lastname: Lupiáñez
authors.affiliation: Max Planck Institute for Molecular Genetics, RG Development & Disease, 14195 Berlin, Germany; Institute for Medical and Human Genetics, Charité Universitätsmedizin Berlin, 13353 Berlin, Germany.
authors.forename: Katerina
authors.initials: K
authors.lastname: Kraft
authors.affiliation: Institute for Medical and Human Genetics, Charité Universitätsmedizin Berlin, 13353 Berlin, Germany.
authors.forename: Verena
authors.initials: V
authors.lastname: Heinrich
authors.affiliation: Max Planck Institute for Molecular Genetics, RG Development & Disease, 14195 Berlin, Germany; Institute for Medical and Human Genetics, Charité Universitätsmedizin Berlin, 13353 Berlin, Germany.
authors.forename: Peter
authors.initials: P
authors.lastname: Krawitz
authors.affiliation: Medical Genetics Unit, Policlinico Tor Vergata University Hospital, 00133 Rome, Italy.
authors.forename: Francesco
authors.initials: F
authors.lastname: Brancati
authors.affiliation: Institute of Human Genetics Biozentrum, Julius Maximilian University of Würzburg, 97070 Würzburg, Germany.
authors.forename: Eva
authors.initials: E
authors.lastname: Klopocki
authors.affiliation: Institute for Medical and Human Genetics, Charité Universitätsmedizin Berlin, 13353 Berlin, Germany.
authors.forename: Denise
authors.initials: D
authors.lastname: Horn
authors.affiliation: Medical Genetics Department, Istanbul Medical Faculty, Istanbul University, 34093 Istanbul, Turkey.
authors.forename: Hülya
authors.initials: H
authors.lastname: Kayserili
authors.affiliation: Department of Pediatrics, School of Medicine, University of Utah, Salt Lake City, UT 84108, USA.
authors.forename: John M
authors.initials: JM
authors.lastname: Opitz
authors.affiliation: Department of Pediatrics, School of Medicine, University of Utah, Salt Lake City, UT 84108, USA.
authors.forename: Renata
authors.initials: R
authors.lastname: Laxova
authors.affiliation: Instituto de Genética Médica y Molecular (INGEMM), IdiPAZ, Hospital Universitario La Paz, 28046 Madrid, Spain; U753 Centro de Investigación Biomédica en Red de Enfermedades Raras (CIBERER), Instituto de Salud Carlos III, 28046 Madrid, Spain.
authors.forename: Fernando
authors.initials: F
authors.lastname: Santos-Simarro
authors.affiliation: Service de Génétique, C.H.U. de Poitiers, 86021 Poitiers, France.
authors.forename: Brigitte
authors.initials: B
authors.lastname: Gilbert-Dussardier
authors.affiliation: Department Developmental Genetics, Max Planck Institute for Molecular Genetics, 14195 Berlin, Germany.
authors.forename: Lars
authors.initials: L
authors.lastname: Wittler
authors.affiliation: Max Planck Institute for Molecular Genetics, RG Development & Disease, 14195 Berlin, Germany.
authors.forename: Marina
authors.initials: M
authors.lastname: Borschiwer
authors.affiliation: Department of Computational Molecular Biology, Max Planck Institute for Molecular Genetics, 14195 Berlin, Germany.
authors.forename: Stefan A
authors.initials: SA
authors.lastname: Haas
authors.affiliation: Genomics Division, MS 84-171, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA.
authors.forename: Marco
authors.initials: M
authors.lastname: Osterwalder
authors.affiliation: Max Planck Institute for Molecular Genetics, RG Development & Disease, 14195 Berlin, Germany; Institute for Medical and Human Genetics, Charité Universitätsmedizin Berlin, 13353 Berlin, Germany.
authors.forename: Martin
authors.initials: M
authors.lastname: Franke
authors.affiliation: Max Planck Institute for Molecular Genetics, Sequencing Core Facility, 14195 Berlin, Germany.
authors.forename: Bernd
authors.initials: B
authors.lastname: Timmermann
authors.affiliation: Max Planck Institute for Molecular Genetics, RG Development & Disease, 14195 Berlin, Germany; Berlin-Brandenburg Center for Regenerative Therapies (BCRT), Charité Universitätsmedizin Berlin, 13353 Berlin, Germany.
authors.forename: Jochen
authors.initials: J
authors.lastname: Hecht
authors.affiliation: Max Planck Institute for Molecular Genetics, RG Development & Disease, 14195 Berlin, Germany; Institute for Medical and Human Genetics, Charité Universitätsmedizin Berlin, 13353 Berlin, Germany; Berlin-Brandenburg Center for Regenerative Therapies (BCRT), Charité Universitätsmedizin Berlin, 13353 Berlin, Germany.
authors.forename: Malte
authors.initials: M
authors.lastname: Spielmann
authors.affiliation: Genomics Division, MS 84-171, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA; U.S. Department of Energy Joint Genome Institute, Walnut Creek, CA 94598, USA; School of Natural Sciences, University of California, Merced, CA 95343, USA.
authors.forename: Axel
authors.initials: A
authors.lastname: Visel
authors.affiliation: Max Planck Institute for Molecular Genetics, RG Development & Disease, 14195 Berlin, Germany; Institute for Medical and Human Genetics, Charité Universitätsmedizin Berlin, 13353 Berlin, Germany; Berlin-Brandenburg Center for Regenerative Therapies (BCRT), Charité Universitätsmedizin Berlin, 13353 Berlin, Germany. Electronic address: mundlos@molgen.mpg.de.
authors.forename: Stefan
authors.initials: S
authors.lastname: Mundlos
mesh_terms: D000818:Animals; D004195:Disease Models, Animal; D004742:Enhancer Elements, Genetic; D005121:Extremities / Q000033:anatomy & histology / Q000254:growth & development; D005786:Gene Expression Regulation; D006801:Humans; D017880:Limb Deformities, Congenital / Q000235:genetics; D051379:Mice; D011401:Promoter Regions, Genetic; D022661:RNA, Untranslated / Q000235:genetics / Q000378:metabolism; D036122:Receptor, EphA4 / Q000235:genetics
chemical_list: D022661:RNA, Untranslated; D036122:Receptor, EphA4