Post-replicative formation of cohesion is required for repair and induced by a single DNA break

Sister chromatid cohesion, established during replication by the protein complex Cohesin, is essential for both chromosome segregation and double-strand break (DSB) repair. Normally cohesion formation is strictly limited to the S-phase of the cell cycle, but DSBs can trigger cohesion also after DNA replication has been completed. The function of this damage-induced cohesion remains unknown. In this investigation we show that it is essential for repair in post-replicative cells in yeast. Furthermore, it is established genome-wide after induction of a single DSB, and controlled by the DNA damage response and Cohesin regulating factors. We thus define a cohesion establishment pathway that is independent of DNA duplication and acts together with cohesion formed during replication in sister chromatid-based DSB repair. Keywords: ChIP-chip analysis Scc2, Scc1, and rH2AX distribution with or without DSB in wild type and mutant strains. DSB was induced by galactose promoter driven HO. All yeast strains were haploid and of W303 origin (ade2-1, trp1-1, can1-100, leu2-3, 112, his3-11, 15, ura3, RAD5). DNA was purified and amplified as previously described by Katou et al. (nature, 2003), and hybridized to SC3456a 52005F or S.cerevisiae Tiling 1.0F arrays, Part#520286 (Affymetrix). Scc2 was C- terminally tagged with 6HIS-3xFLAG, while Mcd1UNCL was marked with 3HA epitopes. Anti-FLAG antibody M2 (Sigma), anti-HA antibody (Babco), or anti γ-H2A-antobodies kindly provided by A. Verreault were used for immuno-precipitations.