Orderly termination of sister-chromatid cohesion during mitosis is critical for accurate chromosome segregation. During prophase, mitotic kinases phosphorylate cohesin and its protector sororin, triggering Wapl-dependent cohesin release from chromosome arms. The shugoshin (Sgo1)–PP2A complex protects centromeric cohesin until its cleavage by separase at anaphase onset. Here, we report the crystal structure of a human cohesin subcomplex comprising SA2 and Scc1. Multiple HEAT repeats of SA2 form a dragon-shaped structure. Scc1 makes extensive contacts with SA2, with one binding hotspot. Sgo1 and Wapl compete for binding to a conserved site on SA2–Scc1. Mutations of SA2 residues at this site that disrupt Wapl binding bypass Sgo1 requirement in cohesion protection. Thus, besides recruiting PP2A to dephosphorylate cohesin and sororin, Sgo1 physically shields cohesin from Wapl. This unexpected, direct antagonism between Sgo1 and Wapl augments centromeric cohesion protection.

SUMMARY The ring-shaped cohesin complex regulates transcription, DNA repair, and chromosome segregation by dynamically entrapping chromosomes to promote chromosome compaction and sister-chromatid cohesion. The cohesin ring needs to open and close to allow its loading to and release from chromosomes. Cohesin dynamics are controlled by the releasing factors Pds5 and Wapl and the cohesin stabilizer Sororin. Here, we report the crystal structure of human Pds5B bound to a conserved peptide motif found in both Wapl and Sororin. Our structure establishes the basis for how Wapl and Sororin antagonistically influence cohesin dynamics. The structure further reveals that Pds5 can bind inositol hexakisphosphate (IP6). The IP6-binding segment of Pds5B is shaped like the jaw of a plier lever and inhibits the binding of Scc1 to Smc3. We propose that Pds5 stabilizes a transient, open state of cohesin to promote its release from chromosomes.

Cohesin, along with positive regulators, establishes sister-chromatid cohesion by forming a ring to circle chromatin. The wings apart-like protein (Wapl) is a key negative regulator of cohesin and forms a complex with precocious dissociation of sisters protein 5 (Pds5) to promote cohesin release from chromatin. Here we report the crystal structure and functional characterization of human Wapl. Wapl contains a flexible, variable N-terminal region (Wapl-N) and a conserved C-terminal domain (Wapl-C) consisting of eight HEAT (Huntingtin, Elongation factor 3, A subunit, and target of rapamycin) repeats. Wapl-C folds into an elongated structure with two lobes. Structure-based mutagenesis maps the functional surface of Wapl-C to two distinct patches (I and II) on the N lobe and a localized patch (III) on the C lobe. Mutating critical patch I residues weaken Wapl binding to cohesin and diminish sister-chromatid resolution and cohesin release from mitotic chromosomes in human cells and Xenopus egg extracts. Surprisingly, patch III on the C lobe does not contribute to Wapl binding to cohesin or its known regulators. Although patch I mutations reduce Wapl binding to intact cohesin, they do not affect Wapl-Pds5 binding to the cohesin subcomplex of sister chromatid cohesion protein 1 (Scc1) and stromal antigen 2 (SA2) in vitro, which is instead mediated by Wapl-N. Thus, Wapl-N forms extensive interactions with Pds5 and Scc1-SA2. Wapl-C interacts with other cohesin subunits and possibly unknown effectors to trigger cohesin release from chromatin.chromosome segregation | crystallography | genomic stability | mitosis | protein-protein interaction P roper chromosome segregation during mitosis maintains genomic stability. Errors in this process cause aneuploidy, which contributes to tumorigenesis under certain contexts (1). Timely establishment and dissolution of sister-chromatid cohesion are critical for accurate chromosome segregation and require the cell-cycle-regulated interactions between cohesin and its regulators (2-4).In human cells, cohesin consists of four core subunits: Structural maintenance of chromosomes 1 (Smc1), Smc3, sister chromatid cohesion protein 1 (Scc1), and stromal antigen 1 or 2 (SA1/2). Smc1 and Smc3 are related ATPases, and each contains an ATPase head domain, a long coiled-coil domain, and a hinge domain that mediates Smc1-Smc3 heterodimerization. The Smc1-Smc3 heterodimer associates with the Scc1-SA1/2 heterodimer to produce the intact cohesin. Specifically, the N-and C-terminal winged helix domains (WHDs) of Scc1 connect the ATPase domains of Smc3 and Smc1, respectively, forming a ring (4).Cohesin is loaded onto chromatin in telophase/G1, but the chromatin-bound cohesin at this stage is highly dynamic and is actively removed from chromatin by the cohesin inhibitor Wings apart-like protein (Wapl) (5-7). During DNA replication in S phase, the ATPase head domain of Smc3 is acetylated by the acetyltransferase establishment of cohesion protein 1 (Eco1) (8-13). In vertebrates, replication-coupled Smc3 acet...

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