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How does Cdc14 order exit from mitosis?

Kataria, M; (2017) How does Cdc14 order exit from mitosis? Doctoral thesis , UCL (University College London). Green open access

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Abstract

Accurate progress through the cell cycle necessitates robust order and timing of events, the molecular basis for which is incompletely understood. During exit from mitosis, it is vital for cells to execute events in a precise sequence. Completion of chromosome segregation must precede chromatin decondensation, or indeed, cytokinesis. In budding yeast, the ordering of mitotic exit events is imposed by the changing balance between Cdk kinase activity, which begins to decline upon anaphase entry, and increasing activity of the major Cdk-counteracting phosphatase, Cdc14. In early anaphase, Cdk activity is still high and little Cdc14 is active. Nonetheless, certain substrates, such as the spindle stabilizer Fin1, are dephosphorylated to facilitate early anaphase events, e.g. spindle elongation. It has been shown that Cdc14 has a high catalytic efficiency towards its early substrates, when compared to later substrates. In this study, we seek a molecular understanding of mitotic exit by probing the biochemical characteristics of early Cdc14 substrate recognition. Through complementary in vitro and in vivo approaches, we find that Cdc14 is a dimer and the non-conserved C-terminal domain is not important for dimerization or substrate dephosphorylation. Cdc14 prefers phosphoserines over phosphothreonines, as well as a basic residue in the +3 position, even when presented in the context of Fin1, a highly efficient substrate. Further, it engages with this substrate via its non-catalytic Nterminal domain. We also find that Fin1 possesses a higher affinity towards Cdc14 compared to other substrates, which is independent of its phosphorylation sites. Truncation analysis revealed that Fin1’s C-terminus is important for this binding, and has a small impact on catalysis. Further, we have also established that early S-phase cyclin degradation is partly responsible for ensuring early dephosphorylation of this protein. In summary, multiple mechanisms collaborate to ensure efficient early substrate dephosphorylation.

Type: Thesis (Doctoral)
Title: How does Cdc14 order exit from mitosis?
Event: University College London
Open access status: An open access version is available from UCL Discovery
Language: English
URI: http://discovery.ucl.ac.uk/id/eprint/1542428
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