Fundamental roles of chromatin loop extrusion in antibody class switching

Antibody class switch recombination (CSR) in B lymphocytes replaces immunoglobulin heavy chain locus ( Igh ) Cμ constant region exons (C H s) with one of six C H s lying 100–200 kb downstream 1 . Each C H is flanked upstream by an I promoter and long repetitive switch (S) region 1 . Cytokines and activators induce activation-induced cytidine deaminase (AID) 2 and I-promoter transcription, with 3′ IgH regulatory region (3′ IgHRR) enhancers controlling the latter via I-promoter competition for long-range 3′ IgHRR interactions 3 – 8 . Transcription through donor Sμ and an activated downstream acceptor S-region targets AID-generated deamination lesions at, potentially, any of hundreds of individual S-region deamination motifs 9 – 11 . General DNA repair pathways convert these lesions to double-stranded breaks (DSBs) and join an Sμ-upstream DSB-end to an acceptor S-region-downstream DSB-end for deletional CSR 12 . AID-initiated DSBs at targets spread across activated S regions routinely participate in such deletional CSR joining 11 . Here we report that chromatin loop extrusion underlies the mechanism 11 by which IgH organization in cis promotes deletional CSR. In naive B cells, loop extrusion dynamically juxtaposes 3′ IgHRR enhancers with the 200-kb upstream Sμ to generate a CSR centre (CSRC). In CSR-activated primary B cells, I-promoter transcription activates cohesin loading, leading to generation of dynamic subdomains that directionally align a downstream S region with Sμ for deletional CSR. During constitutive Sα CSR in CH12F3 B lymphoma cells, inversional CSR can be activated by insertion of a CTCF-binding element (CBE)-based impediment in the extrusion path. CBE insertion also inactivates upstream S-region CSR and converts adjacent downstream sequences into an ectopic S region by inhibiting and promoting their dynamic alignment with Sμ in the CSRC, respectively. Our findings suggest that, in a CSRC, dynamically impeded cohesin-mediated loop extrusion juxtaposes proper ends of AID-initiated donor and acceptor S-region DSBs for deletional CSR. Such a mechanism might also contribute to pathogenic DSB joining genome-wide. Chromatin loop extrusion has fundamental mechanistic roles in immunoglobulin heavy chain class switch recombination.