Structural basis of cohesin cleavage by separase

Abstract

The crystal structures of the protease domain of separase are reported, showing how separase recognizes cohesin, and how phosphorylation of the cleavage site enhances separase activity. Paired chromosomes are held together by the cohesin complex. Although the paired chromosomes attach to the mitotic spindle prior to cell division, as cells divide they are pulled apart to separate poles. For this to happen, cohesin must be cleaved by a protease, separase. Hongtao Yu and colleagues have solved crystal structures of the protease domain of a fungal separase, both unbound and bound to peptides corresponding to the cleavage site of cohesin. The structures provide insights into how separase recognizes cohesin, and how phosphorylation of the cleavage site enhances separase activity. Accurate chromosome segregation requires timely dissolution of chromosome cohesion after chromosomes are properly attached to the mitotic spindle. Separase is absolutely essential for cohesion dissolution in organisms from yeast to man1,2. It cleaves the kleisin subunit of cohesin and opens the cohesin ring to allow chromosome segregation. Cohesin cleavage is spatiotemporally controlled by separase-associated regulatory proteins, including the inhibitory chaperone securin3,4,5,6, and by phosphorylation of both the enzyme and substrates7,8,9,10,11,12. Dysregulation of this process causes chromosome missegregation and aneuploidy, contributing to cancer and birth defects. Despite its essential functions, atomic structures of separase have not been determined. Here we report crystal structures of the separase protease domain from the thermophilic fungus Chaetomium thermophilum, alone or covalently bound to unphosphorylated and phosphorylated inhibitory peptides derived from a cohesin cleavage site. These structures reveal how separase recognizes cohesin and how cohesin phosphorylation by polo-like kinase 1 (Plk1) enhances cleavage. Consistent with a previous cellular study13, mutating two securin residues in a conserved motif that partly matches the separase cleavage consensus converts securin from a separase inhibitor to a substrate. Our study establishes atomic mechanisms of substrate cleavage by separase and suggests competitive inhibition by securin.