In budding yeast and humans, cohesion establishment during S phase requires the acetyltransferase Eco1/Esco1-2, which acetylates the cohesin subunit Smc3 on two conserved lysine residues. Whether Smc3 is the sole Eco1/Esco1-2 effector and how Smc3 acetylation promotes cohesion are unknown. In fission yeast (Schizosaccharomyces pombe), as in humans, cohesin binding to G 1 chromosomes is dynamic and the unloading reaction is stimulated by Wpl1 (human ortholog, Wapl). During S phase, a subpopulation of cohesin becomes stably bound to chromatin in an Eso1 (fission yeast Eco1/Esco1-2)-dependent manner. Cohesin stabilization occurs unevenly along chromosomes. Cohesin remains largely labile at the rDNA repeats but binds mostly in the stable mode to pericentromere regions. This pattern is largely unchanged in eso1⌬ wpl1⌬ cells, and cohesion is unaffected, indicating that the main Eso1 role is counteracting Wpl1. A mutant of Psm3 (fission yeast Smc3) that mimics its acetylated state renders cohesin less sensitive to Wpl1-dependent unloading and partially bypasses the Eso1 requirement but cannot generate the stable mode of cohesin binding in the absence of Eso1. Conversely, nonacetylatable Psm3 reduces the stable cohesin fraction and affects cohesion in a Wpl1-dependent manner, but cells are viable. We propose that Psm3 acetylation contributes to Eso1 counteracting of Wpl1 to secure stable cohesin interaction with postreplicative chromosomes but that it is not the sole molecular event by which this occurs.Following DNA replication in S phase, sister DNA molecules are linked together by cohesin. Thereafter, cohesion between sister chromatids persists throughout the G 2 phase and until mitosis, where it allows chromosome biorientation on the mitotic spindle (16). Defects in this process have been linked to aneuploidy and tumor progression. Cohesin is a multisubunit protein complex made of a dimer of long, flexible Smc subunits, which form a ring-shaped structure stabilized by the binding of a kleisin subunit (Scc1/Rad21 in the mitotic cycle; Rec8 in meiosis) (2, 25). Kleisin cleavage by separase destroys the ring and allows chromatid separation at anaphase (28,43,58). The ring shape of the complex suggested that cohesin ensures cohesion by topological trapping of sister DNA molecules, and strong experimental evidence supports this model (24), although other modes of cohesin-DNA interaction might coexist (29,40).One key aspect of the cohesion cycle is how cohesion is made during S phase. Cohesin is first deposited on unreplicated chromosomes in a reaction requiring ATP hydrolysis by the Smc heads and the cohesin-loading complex Scc2/Scc4 (4,5,15,61). In an unperturbed cell cycle, cohesion is made exclusively during S phase, and except in the event of a DNA double-strand break (DSB), cohesin loading after DNA replication does not result in functional cohesion (26,34,51,60). Numerous studies have shown that mutations in nonessential factors associated with the replication fork machinery affect sister chromatid cohesion, leadin...

Epigenetic information is transmitted from mother to daughter cells through mitosis. Here, to identify factors that might play a role in conveying epigenetic memory through cell division, we report on the isolation of unfixed, native chromosomes from metaphase-arrested cells using flow cytometry and perform LC-MS/MS to identify chromosome-bound proteins. A quantitative proteomic comparison between metaphase-arrested cell lysates and chromosome-sorted samples reveals a cohort of proteins that were significantly enriched on mitotic ESC chromosomes. These include pluripotency-associated transcription factors, repressive chromatin-modifiers such as PRC2 and DNA methyl-transferases, and proteins governing chromosome architecture. Deletion of PRC2, Dnmt1/3a/3b or Mecp2 in ESCs leads to an increase in the size of individual mitotic chromosomes, consistent with decondensation. Similar results were obtained by the experimental cleavage of cohesin. Thus, we identify chromosome-bound factors in pluripotent stem cells during mitosis and reveal that PRC2, DNA methylation and Mecp2 are required to maintain chromosome compaction.