Abstract
Cohesion between sister chromatids is established during DNA replication but needs to be maintained to enable proper chromosome–spindle attachments in mitosis or meiosis. Cohesion is mediated by cohesin, but also depends on cohesin acetylation and sororin. Sororin contributes to cohesion by stabilizing cohesin on DNA. Sororin achieves this by inhibiting WAPL, which otherwise releases cohesin from DNA and destroys cohesion. Here we describe mouse models which enable the controlled depletion of sororin by gene deletion or auxin‐induced degradation. We show that sororin is essential for embryonic development, cohesion maintenance, and proper chromosome segregation. We further show that the acetyltransferases ESCO1 and ESCO2 are essential for stabilizing cohesin on chromatin, that their only function in this process is to acetylate cohesin's SMC3 subunit, and that DNA replication is also required for stable cohesin–chromatin interactions. Unexpectedly, we find that sororin interacts dynamically with the cohesin complexes it stabilizes. This implies that sororin recruitment to cohesin does not depend on the DNA replication machinery or process itself, but on a property that cohesin acquires during cohesion establishment.
Highlights
The accurate segregation of replicated chromosomes during mitosis depends on the prior attachment of sister kinetochores to microtubules from opposing spindle poles
Our results indicate that in human cells the SMC3 acetyltransferases ESCO1 and ESCO2 are required for stabilization of cohesin on chromatin during DNA replication, as one would predict if SMC3 acetylation leads to interactions between cohesin and sororin and to inhibition of WAPL
We depleted ESCO1 and ESCO2 by RNA interference (RNAi) from HeLa cells expressing SMC3 fused to greenfluorescent protein (GFP), synchronized these cells in G2-phase by release from a thymidine-induced DNA replication arrest (Fig 1A, Appendix Fig S1A), and measured cohesin–chromatin interactions in inverse fluorescence recovery after photobleaching experiments (Fig 1B)
Summary
The accurate segregation of replicated chromosomes during mitosis depends on the prior attachment of sister kinetochores to microtubules from opposing spindle poles. Cohesin associates with chromatin already before DNA replication (Losada et al, 1998; Sumara et al, 2000) in a manner that depends on integrity of the cohesin ring (Pauli et al, 2008; Huis in ‘t Veld et al, 2014), implying that cohesin interacts with unreplicated DNA via topological entrapment. These cohesin–DNA interactions can be reversed by the cohesinassociated protein WAPL (Gandhi et al, 2006; Kueng et al, 2006; Tedeschi et al, 2013), which releases cohesin from DNA by opening a DNA “gate” between SMC3 and SCC1 (Chan et al, 2012; Buheitel & Stemmann, 2013; Eichinger et al, 2013; Huis in ‘t Veld et al, 2014). Similar cohesin–DNA interactions might exist in post-mitotic cells (Wendt et al, 2008) in which cohesin has roles in chromatin structure and gene regulation (reviewed in Seitan & Merkenschlager, 2012)
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