Cohesin regulates homology search during recombinational DNA repair [ChIP-Seq]

Homologous recombination (HR) is an ubiquitous DNA double-strand break (DSB) repair mechanism. It entails a homology search step, carried out along a conserved RecA/Rad51-ssDNA nucleoprotein filament (NPF) assembled on each DSB ends. In contrast to the extensive knowledge of DNA damage checkpoint (DDC)-induced changes in chromatin composition and mobility,  the questions of if, how, and to what extent a DSB impacts the spatial organization of chromatin, and whether this organization in turn influences the homology search process, remain ill-defined. Here we characterize two layers of spatial chromatin reorganization following DSB formation in S. cerevisiae. While cohesin folds chromosomes into cohesive arrays of ~20 kb-long chromatin loops as cells arrest in G2/M, the DSB-flanking regions interact locally in a resection- and 9-1-1 clamp-dependent manner, independently of cohesin, Mec1ATR, Rad52 and Rad51. This local structure blocks cohesin progression, constraining the DSB region at the base of a loop. Functionally, cohesin promotes DSB-dsDNA interactions and donor identification in cis, while inhibiting them in trans. This study identifies multiple direct and indirect ways by which cohesin regulates homology search during HR repair. Scc1-Pk/HA ChIP-seq analysis were performed on yeast pellets collected 4 hours after DSB induced by galactose addition on several samples WT and mutants. Scc1-Pk ChIP-seq IP and input in WT (no DSB + nocodazole), WT (DSB 4hrs), WT and rad17 (DSB 4 hrs + nocodazole) were performed once. Scc1-HA ChIP-seq IP and input in WT, rad17, pds5 and pds5 rad17 (DSB 4 hrs + nocodazole) were performed twice and only once for WT (no DSB + nocodazole) and untagged control. In order to compare quantitatively the samples, the ChIP-seq experiments are calibrated with Candida glabrata as described in Hu et al. 2015.