Towards Understanding Substrate Recognition and Regulatory Protein Complexes of Human Separase - A Molecular Scissor initiating Chromosome Segregation

Diplôme Thèse Towards Understanding Substrate Recognition and Regulatory Protein Complexes of Human Separase - A Molecular Scissor initiating Chromosome Segregation eng 2026-04-16 2026-05-10 author Schmidt Sophia 985626 director Boland Andreas 912539 Département de biologie moléculaire et cellulaire (MOCEL) 177 10.13097/archive-ouverte/unige:194226 urn:nbn:ch:unige-1942268 Sc. Vie 396 The transition from metaphase to anaphase within the cell cycle is a no-return process. It is initiated when the protease separase cleaves sister chromatid cohesion 1 / RAD21 homolog (SCC1), a subunit of the ring-shaped protein cohesin that holds the sister chromatids together. Once chromosome cohesion is released, the sister chromatids are pulled towards the spindle poles. To avoid segregation errors, human separase is held inactive by securin and cyclin-dependent kinases (CDK)1-cyclin B1 until the chromosomes are properly attached to the spindle microtubules. The formation of the complex separase-CDK1-cyclin B1 requires the separase residue serine 1126 to be phosphorylated (pS1126separase). pS1126separase is suggested to be dephosphorylated by the phosphatase protein phosphatase 2 (PP2A). Besides PP2A, also the cohesin-associated protein sororin is suggested to regulate separase and was shown to bind to separase in vivo. This PhD thesis addresses the questions how separase recognizes its substrate SCC1 and how separase is regulated through interactions with PP2A and sororin. Among the two previously reported SCC1 cleavage sites, we validate the first cleavage site and correct the second cleavage site. Consistent with in vivo studies with human SCC1 we show that SCC1 cleavage is stimulated if it is phosphorylated. We also identify conserved separase substrate phosphorylation sites that interact with five phosphate-binding pockets on separase. Furthermore, we observe that the cohesin subunit stromal antigen 2 (SA2) promotes SCC1 cleavage by recruiting SCC1 to separase. Moreover, we show that securin and sororin compete with each other for binding to separase, and that sororin can only bind to securin-free separase. It was previously proposed that the conserved substrate pocket of the regulatory subunit of PP2A binds to the PP2A substrate motif on the flexible insert 2 of separase. Then, PP2A would dissociate from active separase, because active separase undergoes autocleavage, destroying the PP2A substrate motif. Supporting this hypothesis, we can only form a complex of PP2A and inactive separase that cannot do autocleavage. We observe that the main interaction between separase and PP2A is mediated by the conserved substrate pocket of PP2A. Overall, our study demonstrates for the first time that PP2A and sororin bind to separase in vitro. Separase PP2A Sororin Chromosome Segregation Cell cycle control Dewey 570 7daed2ee-1311-4824-9df9-2e51260a966a Département de biologie moléculaire et cellulaire 16484 false 1277 Groupe Boland 7d151375-ec20-443e-b242-f7d9520a35dc Thesis Public PhD_Sophia_Schmidt.pdf 119716782 application/pdf Private 2027-06-11 99303dd9d1d7fccf370d6f3a6a2a929664609b7453c5d7bc982c3f67233e5769 9b0269e8-582c-41dc-a6b8-4763c3cb6cc6 Imprimatur Public Imprimatur_Sophia_Schmidt.pdf 434816 application/pdf 1039acb98986da6c65631d44ac7dab78c3b1a43db41cadcff053bd9a4f83c7c1 embargoedAccess Faculté des sciences / Section de biologie / Département de biologie moléculaire et cellulaire

Title	Published in	Access level	OA Policy	Year	Views	Downloads

Towards Understanding Substrate Recognition and Regulatory Protein Complexes of Human Separase - A Molecular Scissor initiating Chromosome Segregation				2026	14	0

Archive ouverte UNIGE

Schmidt, Sophia

account_balance Affiliation entities

Research groups

Affiliation entities