title: Chromosomal Loop Domains Direct the Recombination of Antigen Receptor Genes.
abstract: RAG initiates antibody V(D)J recombination in developing lymphocytes by generating "on-target" DNA breaks at matched pairs of bona fide recombination signal sequences (RSSs). We employ bait RAG-generated breaks in endogenous or ectopically inserted RSS pairs to identify huge numbers of RAG "off-target" breaks. Such breaks occur at the simple CAC motif that defines the RSS cleavage site and are largely confined within convergent CTCF-binding element (CBE)-flanked loop domains containing bait RSS pairs. Marked orientation dependence of RAG off-target activity within loops spanning up to 2 megabases implies involvement of linear tracking. In this regard, major RAG off-targets in chromosomal translocations occur as convergent RSS pairs at enhancers within a loop. Finally, deletion of a CBE-based IgH locus element disrupts V(D)J recombination domains and, correspondingly, alters RAG on- and off-target distributions within IgH. Our findings reveal how RAG activity is developmentally focused and implicate mechanisms by which chromatin domains harness biological processes within them.
authors.affiliation: Howard Hughes Medical Institute; Program in Cellular and Molecular Medicine, Boston Children's Hospital, Boston, MA 02115, USA; Department of Genetics, Harvard Medical School, Boston, MA 02115, USA.
authors.forename: Jiazhi
authors.initials: J
authors.lastname: Hu
authors.affiliation: Howard Hughes Medical Institute; Program in Cellular and Molecular Medicine, Boston Children's Hospital, Boston, MA 02115, USA; Department of Genetics, Harvard Medical School, Boston, MA 02115, USA.
authors.forename: Yu
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Howard Hughes Medical Institute; Program in Cellular and Molecular Medicine, Boston Children's Hospital, Boston, MA 02115, USA; Department of Genetics, Harvard Medical School, Boston, MA 02115, USA.
authors.forename: Lijuan
authors.initials: L
authors.lastname: Zhao
authors.affiliation: Howard Hughes Medical Institute; Program in Cellular and Molecular Medicine, Boston Children's Hospital, Boston, MA 02115, USA; Department of Genetics, Harvard Medical School, Boston, MA 02115, USA.
authors.forename: Richard L
authors.initials: RL
authors.lastname: Frock
authors.affiliation: Howard Hughes Medical Institute; Program in Cellular and Molecular Medicine, Boston Children's Hospital, Boston, MA 02115, USA; Department of Genetics, Harvard Medical School, Boston, MA 02115, USA.
authors.forename: Zhou
authors.initials: Z
authors.lastname: Du
authors.affiliation: Howard Hughes Medical Institute; Program in Cellular and Molecular Medicine, Boston Children's Hospital, Boston, MA 02115, USA; Department of Genetics, Harvard Medical School, Boston, MA 02115, USA.
authors.forename: Robin M
authors.initials: RM
authors.lastname: Meyers
authors.affiliation: Howard Hughes Medical Institute; Program in Cellular and Molecular Medicine, Boston Children's Hospital, Boston, MA 02115, USA; Department of Genetics, Harvard Medical School, Boston, MA 02115, USA.
authors.forename: Fei-long
authors.initials: FL
authors.lastname: Meng
authors.affiliation: Howard Hughes Medical Institute; Department of Immunobiology, Yale University School of Medicine, 300 Cedar Street, Box 208011, New Haven, CT 06520-8011, USA.
authors.forename: David G
authors.initials: DG
authors.lastname: Schatz
authors.affiliation: Howard Hughes Medical Institute; Program in Cellular and Molecular Medicine, Boston Children's Hospital, Boston, MA 02115, USA; Department of Genetics, Harvard Medical School, Boston, MA 02115, USA. Electronic address: alt@enders.tch.harvard.edu.
authors.forename: Frederick W
authors.initials: FW
authors.lastname: Alt
mesh_terms: D000818:Animals; D000076246:CCCTC-Binding Factor; D033481:Chromosomes, Mammalian / Q000737:chemistry / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism; D016259:Genes, myc; D016678:Genome; D059014:High-Throughput Nucleotide Sequencing; D018398:Homeodomain Proteins / Q000378:metabolism; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000235:genetics; D008223:Lymphoma / Q000235:genetics; D051379:Mice; D059372:Nucleotide Motifs; D012045:Regulatory Sequences, Nucleic Acid; D012097:Repressor Proteins / Q000378:metabolism; D017422:Sequence Analysis, DNA; D014178:Translocation, Genetic; D060152:V(D)J Recombination
chemical_list: D000076246:CCCTC-Binding Factor; C000619697:CTCF protein, human; C000619698:Ctcf protein, mouse; D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; D007143:Immunoglobulin Heavy Chains; C499308:Rag2 protein, mouse; D012097:Repressor Proteins; C064658:RAG-1 protein