Abstract
The immunoglobulin heavy chain locus (Igh) features higher-order chromosomal interactions to facilitate stage-specific assembly of the Ig molecule. Cohesin, a ring-like protein complex required for sister chromatid cohesion, shapes chromosome architecture and chromatin interactions important for transcriptional regulation and often acts together with CTCF. Cohesin is likely involved in B cell activation and Ig class switch recombination. Hence, binding profiles of cohesin in resting mature murine splenic B lymphocytes and at two stages after cell activation were elucidated by chromatin immunoprecipitation and deep sequencing. Comparative genomic analysis revealed cohesin extensively changes its binding to transcriptional control elements after 48 h of stimulation with LPS/IL-4. Cohesin was clearly underrepresented at switch regions regardless of their activation status, suggesting that switch regions need to be cohesin-poor. Specific binding changes of cohesin at B-cell specific gene loci Pax5 and Blimp-1 indicate new cohesin-dependent regulatory pathways. Together with conserved cohesin/CTCF sites at the Igh 3′RR, a prominent cohesin/CTCF binding site was revealed near the 3′ end of Cα where PolII localizes to 3′ enhancers. Our study shows that cohesin likely regulates B cell activation and maturation, including Ig class switching.
Highlights
Cohesin is a chromosome-associated multi-protein complex, conserved from yeast to man, is essential for sister chromatid cohesion, and is involved in DNA repair and recombination
To obtain insights into the potential role of cohesin during class switch recombination (CSR), we traced the differential binding of the cohesin subunit RAD21 to chromatin in purified B cells, which were left unstimulated or were stimulated with IL-4 and LPS to induce the switch to IgG1
Peak calling with a stringent cut-off criteria of 0.05 false discovery rate (FDR) identified 5687 cohesin binding sites in resting B cells (Fig. 1A), and 5547 binding sites in cells stimulated for 48 h with LPS and IL-4
Summary
Cohesin is a chromosome-associated multi-protein complex, conserved from yeast to man, is essential for sister chromatid cohesion, and is involved in DNA repair and recombination The kleisin connects the ATPase heads of the two SMC proteins, thereby forming a ring-like structure, with which the SA protein associates This ring or multiple rings connects two double-stranded DNA molecules. In addition to its role in sister chromatid cohesion, the cohesin protein complex facilitates several kinds of chromatin interactions, some of which are cell type-specific [7,8,9,10,11]. In T cells, Rad deficiency led to reduced promoter-enhancer looping at the TCRa locus associated with transcriptional changes [16]
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