Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Genome-wide localization of Smc1, Smc3 and CTCF in mouse B cells.

ABSTRACT: Immunoglobulin class switch recombination (CSR) is initiated by the transcription-coupled recruitment of activation induced cytidine deaminase (AID) to switch regions and by the subsequent generation and resolution of dsDNA breaks (DSBs). During, CSR the IgH locus undergoes dynamic three-dimensional structural changes in which promoters, enhancers and switch regions are brought to close proximity. Nevertheless, little is known about the underlying mechanisms. Here we show that during CSR, AID associates with subunits of cohesin, a complex previously implicated in DNA repair and in the formation of DNA loops between enhancers and promoters. By ChIP-Seq experiments, we find that Cohesin is dynamically recruited to the IgH locus during CSR and that knockdown of Cohesin or its regulatory subunits results in impaired CSR and abnormal DSB resolution. Our results are consistent with a model in which Cohesin controls the formation of long-range DNA loops at the IgH locus and the resolution of DSBs generated during CSR. Smc1, Smc3 and CTCF were immunoprecipitated from resting or in vitro activated B cells.

ORGANISM(S): Mus musculus

SUBMITTER: Anne-Sophie THOMAS

PROVIDER: E-GEOD-43594 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

The cohesin complex regulates immunoglobulin class switch recombination.

Thomas-Claudepierre Anne-Sophie AS Schiavo Ebe E Heyer Vincent V Fournier Marjorie M Page Adeline A Robert Isabelle I Reina-San-Martin Bernardo B

The Journal of experimental medicine 20131021 12

Immunoglobulin (Ig) class switch recombination (CSR) is initiated by the transcription-coupled recruitment of activation-induced cytidine deaminase (AID) to switch regions and by the subsequent generation of double-stranded DNA breaks (DSBs). These DNA breaks are ultimately resolved through the nonhomologous end joining (NHEJ) pathway. We show that during CSR, AID associates with subunits of cohesin, a complex previously implicated in sister chromatid cohesion, DNA repair, and the formation of D ...[more]

PMID: 24145512

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Project description:In a B lymphocyte immunoglobulin heavy chain locus (IgH), a developmentally assembled V(D)J exon encoding an antibody variable region lies upstream of exons encoding a m constant region (Cm), allowing generation of mIgH chain transcripts and IgM-class antibodies1. Mouse IgH class switch recombination (CSR) replaces Cmwith one of 6 sets of constant region exons (CHs) that lie 100-200kb downstream1. Each CH is flanked upstream by a promoter, non-coding I-exon, and long repetitive switch (S) region1,2. Cytokines/activators induce specific I-promoter transcription and activation-induced cytidine deaminase (AID)2,3. AID is transcriptionally-targeted to initiate DNA breaks in Sm and activated downstream acceptor S regions, which are joined in deletional orientation to complete CSR4,5. 3’IgH regulatory region (3’IgHRR) enhancers control upstream I promoters and, thereby, CSR via linear competition involving I promoter/3’IgHRR interactions6-11. Here, we report that synapsis of regulatory elements, S regions and DSBs for CSR is achieved by chromatin loop extrusion. In naive B cells, 3’IgHRR enhancers and adjacent 3’IgH CTCF-binding elements (CBEs) interact via loop extrusion with the upstream Igh intronic enhancer (iEm)/Sm locale to generate dynamic 200kb 3’Igh basal loop. In CSR-activated B cells, induced transcription from I-promoters within this basal loop generates dynamic sub-loops that directionally align Sm and target S regions near the 3’IgHRR for CSR. In CH12F3 B lymphoma cells, inactivation of the constitutively active Ia-promoter abrogates looping and CSR to Sa, while activating transcription, looping, and CSR to upstream S regions. CBEs inserted upstream of Ia in convergent orientation with 3’IgH CBEs generate sub-loops that activate inversional Sa CSR. In I-promoter-deleted CH12F3 cells, this ectopic CBE-based sub-loop inactivates upstream S region CSR, while transcriptionally activating non-S region sequences adjacent to the inserted CBEs for S synapsis and CSR. Together, our findings implicate chromatin loop extrusion in the “unprecedented mechanism”5 by which Igh organization in cis promotes orientation-specific CSR DSB joining.

Project description:In a B lymphocyte immunoglobulin heavy chain locus (IgH), a developmentally assembled V(D)J exon encoding an antibody variable region lies upstream of exons encoding a  constant region (C), allowing generation of IgH chain transcripts and IgM-class antibodies1. Mouse IgH class switch recombination (CSR) replaces Cwith one of 6 sets of constant region exons (CHs) that lie 100-200kb downstream1. Each CH is flanked upstream by a promoter, non-coding I-exon, and long repetitive switch (S) region1,2. Cytokines/activators induce specific I-promoter transcription and activation-induced cytidine deaminase (AID)2,3. AID is transcriptionally-targeted to initiate DNA breaks in S and activated downstream acceptor S regions, which are joined in deletional orientation to complete CSR4,5. 3’IgH regulatory region (3’IgHRR) enhancers control upstream I promoters and, thereby, CSR via linear competition involving I promoter/3’IgHRR interactions6-11. Here, we report that synapsis of regulatory elements, S regions and DSBs for CSR is achieved by chromatin loop extrusion. In naive B cells, 3’IgHRR enhancers and adjacent 3’IgH CTCF-binding elements (CBEs) interact via loop extrusion with the upstream Igh intronic enhancer (iE)/S locale to generate dynamic 200kb 3’Igh basal loop. In CSR-activated B cells, induced transcription from I-promoters within this basal loop generates dynamic sub-loops that directionally align S and target S regions near the 3’IgHRR for CSR. In CH12F3 B lymphoma cells, inactivation of the constitutively active I-promoter abrogates looping and CSR to S, while activating transcription, looping, and CSR to upstream S regions. CBEs inserted upstream of I in convergent orientation with 3’IgH CBEs generate sub-loops that activate inversional S CSR. In I-promoter-deleted CH12F3 cells, this ectopic CBE-based sub-loop inactivates upstream S region CSR, while transcriptionally activating non-S region sequences adjacent to the inserted CBEs for S synapsis and CSR. Together, our findings implicate chromatin loop extrusion in the “unprecedented mechanism”5 by which Igh organization in cis promotes orientation-specific CSR DSB joining.

Dataset Information

Genome-wide localization of Smc1, Smc3 and CTCF in mouse B cells.

Publications

The cohesin complex regulates immunoglobulin class switch recombination.

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