title: Chromatin insulator factors involved in long-range DNA interactions and their role in the folding of the Drosophila genome.
abstract: Chromatin insulators are genetic elements implicated in the organization of chromatin and the regulation of transcription. In Drosophila, different insulator types were characterized by their locus-specific composition of insulator proteins and co-factors. Insulators mediate specific long-range DNA contacts required for the three dimensional organization of the interphase nucleus and for transcription regulation, but the mechanisms underlying the formation of these contacts is currently unknown. Here, we investigate the molecular associations between different components of insulator complexes (BEAF32, CP190 and Chromator) by biochemical and biophysical means, and develop a novel single-molecule assay to determine what factors are necessary and essential for the formation of long-range DNA interactions. We show that BEAF32 is able to bind DNA specifically and with high affinity, but not to bridge long-range interactions (LRI). In contrast, we show that CP190 and Chromator are able to mediate LRI between specifically-bound BEAF32 nucleoprotein complexes in vitro. This ability of CP190 and Chromator to establish LRI requires specific contacts between BEAF32 and their C-terminal domains, and dimerization through their N-terminal domains. In particular, the BTB/POZ domains of CP190 form a strict homodimer, and its C-terminal domain interacts with several insulator binding proteins. We propose a general model for insulator function in which BEAF32/dCTCF/Su(HW) provide DNA specificity (first layer proteins) whereas CP190/Chromator are responsible for the physical interactions required for long-range contacts (second layer). This network of organized, multi-layer interactions could explain the different activities of insulators as chromatin barriers, enhancer blockers, and transcriptional regulators, and suggest a general mechanism for how insulators may shape the organization of higher-order chromatin during cell division.
authors.affiliation: Centre National de la Recherche Scientifique, Unité Mixte de Recherche 5048, Centre de Biochimie Structurale, Montpellier, France; Institut National de la Santé et la Recherche Médicale, Unité 1054, Montpellier, France; Universités Montpellier I et II, Montpellier, France.
authors.forename: Jutta
authors.initials: J
authors.lastname: Vogelmann
authors.affiliation: Centre National de la Recherche Scientifique, Unité Mixte de Recherche 5048, Centre de Biochimie Structurale, Montpellier, France; Institut National de la Santé et la Recherche Médicale, Unité 1054, Montpellier, France; Universités Montpellier I et II, Montpellier, France.
authors.forename: Antoine
authors.initials: A
authors.lastname: Le Gall
authors.affiliation: Centre National de la Recherche Scientifique, Unité Mixte de Recherche 5048, Centre de Biochimie Structurale, Montpellier, France; Institut National de la Santé et la Recherche Médicale, Unité 1054, Montpellier, France; Universités Montpellier I et II, Montpellier, France.
authors.forename: Stephanie
authors.initials: S
authors.lastname: Dejardin
authors.affiliation: Centre National de la Recherche Scientifique, Unité Mixte de Recherche 5048, Centre de Biochimie Structurale, Montpellier, France; Institut National de la Santé et la Recherche Médicale, Unité 1054, Montpellier, France; Universités Montpellier I et II, Montpellier, France.
authors.forename: Frederic
authors.initials: F
authors.lastname: Allemand
authors.affiliation: Laboratoire de Biologie Moléculaire Eucaryote, CNRS and Université de Toulouse, Toulouse; France.
authors.forename: Adrien
authors.initials: A
authors.lastname: Gamot
authors.affiliation: Centre National de la Recherche Scientifique, Unité Mixte de Recherche 5048, Centre de Biochimie Structurale, Montpellier, France; Institut National de la Santé et la Recherche Médicale, Unité 1054, Montpellier, France; Universités Montpellier I et II, Montpellier, France.
authors.forename: Gilles
authors.initials: G
authors.lastname: Labesse
authors.affiliation: Laboratoire de Biologie Moléculaire Eucaryote, CNRS and Université de Toulouse, Toulouse; France.
authors.forename: Olivier
authors.initials: O
authors.lastname: Cuvier
authors.affiliation: Laboratoire Diversité, Génomes & Interactions Microorganismes-Insectes, INRA UMR1333, Université de Montpellier 2, Montpellier, France.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Nègre
authors.affiliation: Centre National de la Recherche Scientifique, Unité Mixte de Recherche 5048, Centre de Biochimie Structurale, Montpellier, France; Institut National de la Santé et la Recherche Médicale, Unité 1054, Montpellier, France; Universités Montpellier I et II, Montpellier, France.
authors.forename: Martin
authors.initials: M
authors.lastname: Cohen-Gonsaud
authors.affiliation: Centre National de la Recherche Scientifique, Unité Mixte de Recherche 5048, Centre de Biochimie Structurale, Montpellier, France; Institut National de la Santé et la Recherche Médicale, Unité 1054, Montpellier, France; Universités Montpellier I et II, Montpellier, France.
authors.forename: Emmanuel
authors.initials: E
authors.lastname: Margeat
authors.affiliation: Centre National de la Recherche Scientifique, Unité Mixte de Recherche 5048, Centre de Biochimie Structurale, Montpellier, France; Institut National de la Santé et la Recherche Médicale, Unité 1054, Montpellier, France; Universités Montpellier I et II, Montpellier, France.
authors.forename: Marcelo
authors.initials: M
authors.lastname: Nöllmann
mesh_terms: D000818:Animals; D002843:Chromatin / Q000235:genetics*; D004247:DNA / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics; D029721:Drosophila Proteins / Q000235:genetics; D004331:Drosophila melanogaster / Q000235:genetics*; D005136:Eye Proteins / Q000235:genetics; D053263:Gene Regulatory Networks; D049750:Genome, Insect; D038101:Insulator Elements / Q000235:genetics*; D008869:Microtubule-Associated Proteins / Q000235:genetics; D034521:Nuclear Matrix-Associated Proteins / Q000235:genetics; D009687:Nuclear Proteins / Q000235:genetics
chemical_list: C452919:BEAF-32 protein, Drosophila; C107964:CP190 protein, Drosophila; C498868:Chro protein, Drosophila; D002843:Chromatin; D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; D005136:Eye Proteins; D008869:Microtubule-Associated Proteins; D034521:Nuclear Matrix-Associated Proteins; D009687:Nuclear Proteins; D004247:DNA