title: CTCF Binding Polarity Determines Chromatin Looping.
abstract: CCCTC-binding factor (CTCF) is an architectural protein involved in the three-dimensional (3D) organization of chromatin. In this study, we assayed the 3D genomic contact profiles of a large number of CTCF binding sites with high-resolution 4C-seq. As recently reported, our data also suggest that chromatin loops preferentially form between CTCF binding sites oriented in a convergent manner. To directly test this, we used CRISPR/Cas9 genome editing to delete core CTCF binding sites in three loci, including the CTCF site in the Sox2 super-enhancer. In all instances, CTCF and cohesin recruitment were lost, and chromatin loops with distal, convergent CTCF sites were disrupted or destabilized. Re-insertion of oppositely oriented CTCF recognition sequences restored CTCF and cohesin recruitment, but did not re-establish chromatin loops. We conclude that CTCF binding polarity plays a functional role in the formation of higher-order chromatin structure.
authors.affiliation: Hubrecht Institute-KNAW and University Medical Center Utrecht, Uppsalalaan 8, 3584 CT Utrecht, the Netherlands.
authors.forename: Elzo
authors.initials: E
authors.lastname: de Wit
authors.affiliation: Hubrecht Institute-KNAW and University Medical Center Utrecht, Uppsalalaan 8, 3584 CT Utrecht, the Netherlands.
authors.forename: Erica S M
authors.initials: ES
authors.lastname: Vos
authors.affiliation: Hubrecht Institute-KNAW and University Medical Center Utrecht, Uppsalalaan 8, 3584 CT Utrecht, the Netherlands.
authors.forename: Sjoerd J B
authors.initials: SJ
authors.lastname: Holwerda
authors.affiliation: Hubrecht Institute-KNAW and University Medical Center Utrecht, Uppsalalaan 8, 3584 CT Utrecht, the Netherlands.
authors.forename: Christian
authors.initials: C
authors.lastname: Valdes-Quezada
authors.affiliation: Hubrecht Institute-KNAW and University Medical Center Utrecht, Uppsalalaan 8, 3584 CT Utrecht, the Netherlands.
authors.forename: Marjon J A M
authors.initials: MJ
authors.lastname: Verstegen
authors.affiliation: Hubrecht Institute-KNAW and University Medical Center Utrecht, Uppsalalaan 8, 3584 CT Utrecht, the Netherlands.
authors.forename: Hans
authors.initials: H
authors.lastname: Teunissen
authors.affiliation: Hubrecht Institute-KNAW and University Medical Center Utrecht, Uppsalalaan 8, 3584 CT Utrecht, the Netherlands.
authors.forename: Erik
authors.initials: E
authors.lastname: Splinter
authors.affiliation: Hubrecht Institute-KNAW and University Medical Center Utrecht, Uppsalalaan 8, 3584 CT Utrecht, the Netherlands.
authors.forename: Patrick J
authors.initials: PJ
authors.lastname: Wijchers
authors.affiliation: Hubrecht Institute-KNAW and University Medical Center Utrecht, Uppsalalaan 8, 3584 CT Utrecht, the Netherlands.
authors.forename: Peter H L
authors.initials: PH
authors.lastname: Krijger
authors.affiliation: Hubrecht Institute-KNAW and University Medical Center Utrecht, Uppsalalaan 8, 3584 CT Utrecht, the Netherlands. Electronic address: w.delaat@hubrecht.eu.
authors.forename: Wouter
authors.initials: W
authors.lastname: de Laat
mesh_terms: D000818:Animals; D001665:Binding Sites; D000076246:CCCTC-Binding Factor; D064113:CRISPR-Cas Systems; D018797:Cell Cycle Proteins / Q000378:metabolism; D002460:Cell Line; D002843:Chromatin / Q000737:chemistry*; D002868:Chromosomal Proteins, Non-Histone / Q000378:metabolism; D053595:Embryonic Stem Cells / Q000166:cytology; D051379:Mice; D011485:Protein Binding; D012097:Repressor Proteins / Q000737:chemistry* / Q000378:metabolism*; D000097722:Cohesins
chemical_list: D000076246:CCCTC-Binding Factor; D018797:Cell Cycle Proteins; D002843:Chromatin; D002868:Chromosomal Proteins, Non-Histone; D012097:Repressor Proteins; D000097722:Cohesins; C000619698:Ctcf protein, mouse