title: IgH class switching exploits a general property of two DNA breaks to be joined in cis over long chromosomal distances.
abstract: Antibody class switch recombination (CSR) in B lymphocytes joins two DNA double-strand breaks (DSBs) lying 100-200 kb apart within switch (S) regions in the immunoglobulin heavy-chain locus (IgH). CSR-activated B lymphocytes generate multiple S-region DSBs in the donor Sμ and in a downstream acceptor S region, with a DSB in Sμ being joined to a DSB in the acceptor S region at sufficient frequency to drive CSR in a large fraction of activated B cells. Such frequent joining of widely separated CSR DSBs could be promoted by IgH-specific or B-cell-specific processes or by general aspects of chromosome architecture and DSB repair. Previously, we found that B cells with two yeast I-SceI endonuclease targets in place of Sγ1 undergo I-SceI-dependent class switching from IgM to IgG1 at 5-10% of normal levels. Now, we report that B cells in which Sγ1 is replaced with a 28 I-SceI target array, designed to increase I-SceI DSB frequency, undergo I-SceI-dependent class switching at almost normal levels. High-throughput genome-wide translocation sequencing revealed that I-SceI-generated DSBs introduced in cis at Sμ and Sγ1 sites are joined together in T cells at levels similar to those of B cells. Such high joining levels also occurred between I-SceI-generated DSBs within c-myc and I-SceI- or CRISPR/Cas9-generated DSBs 100 kb downstream within Pvt1 in B cells or fibroblasts, respectively. We suggest that CSR exploits a general propensity of intrachromosomal DSBs separated by several hundred kilobases to be frequently joined together and discuss the relevance of this finding for recurrent interstitial deletions in cancer.
authors.affiliation: Howard Hughes Medical Institute, Program in Cellular and Molecular Medicine, Boston Children's Hospital, and Department of Genetics, Harvard Medical School, Boston, MA 02115.
authors.forename: Monica
authors.initials: M
authors.lastname: Gostissa
authors.forename: Bjoern
authors.initials: B
authors.lastname: Schwer
authors.forename: Amelia
authors.initials: A
authors.lastname: Chang
authors.forename: Junchao
authors.initials: J
authors.lastname: Dong
authors.forename: Robin M
authors.initials: RM
authors.lastname: Meyers
authors.forename: Gregory T
authors.initials: GT
authors.lastname: Marecki
authors.forename: Vivian W
authors.initials: VW
authors.lastname: Choi
authors.forename: Roberto
authors.initials: R
authors.lastname: Chiarle
authors.forename: Ali A
authors.initials: AA
authors.lastname: Zarrin
authors.forename: Frederick W
authors.initials: FW
authors.lastname: Alt
mesh_terms: D001402:B-Lymphocytes / Q000276:immunology*; D053903:DNA Breaks, Double-Stranded; D017931:DNA Primers / Q000235:genetics; D004260:DNA Repair / Q000235:genetics / Q000502:physiology*; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D017578:Immunoglobulin Class Switching / Q000235:genetics*; D007143:Immunoglobulin Heavy Chains / Q000235:genetics*; D009369:Neoplasms / Q000235:genetics* / Q000276:immunology; D016133:Polymerase Chain Reaction; D011995:Recombination, Genetic / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism
chemical_list: D017931:DNA Primers; D007143:Immunoglobulin Heavy Chains; D029701:Saccharomyces cerevisiae Proteins; C037725:SCEI protein, S cerevisiae; D015252:Deoxyribonucleases, Type II Site-Specific
keywords: double-strand break synapsis; intrachromosomal joining; topological domains