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bioRxiv · 10.1101/2024.11.01.621564

A coordinated kinase and phosphatase network regulates Stu2 recruitment to yeast kinetochores

Stewart, M. G.·Carrier, J. S.·Zahm, J. A.·Harrison, S.·Miller, M. P.
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Abstract

Cells coordinate diverse events at anaphase onset, including separase activation, cohesin cleavage, chromosome separation, and spindle reorganization. Regulation of the XMAP215 family member and microtubule polymerase, Stu2, at the metaphase-anaphase transition determines a specific redistribution from kinetochores to spindle microtubules. We show that cells modulate Stu2 kinetochore-microtubule localization by Polo-like kinase1/Cdc5-mediated phosphorylation of T866, near the Stu2 C-terminus, thereby promoting dissociation from the kinetochore Ndc80 complex. Cdk/Cdc28 likely primes Cdc5:Stu2 interaction. Cdc28 activity is also required for Stu2 nuclear import. PP2ACdc55 actively opposes Cdc5 activity on Stu2T866 during metaphase. This counter-regulation allows for switchlike redistribution of Stu2pT866 at anaphase onset when separase inhibits PP2ACdc55. Blocking Stu2T866 phosphorylation disrupts anaphase spindle progression, and we infer that PP2ACdc55 regulates the mitotic spindle by dephosphorylating Stu2 and other MAPs. These data support a model in which increased phosphorylation at anaphase onset results from phosphatase inhibition and point to a larger regulatory network that facilitates rapid cytoskeletal modulation required for anaphase spindle maintenance. SUMMARYStu2 displays dynamic localization patterns in the cell cycle, with different kinetochore and microtubule distribution during distinct phases. Phosphorylation near Stu2s C-terminus reduces its attachment to kinetochores to promote its microtubule activity in anaphase. Cdc5 and PP2ACdc55 play counteracting roles in this pathway to promote proper timing of Stu2 phosphorylation.

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Stewart, M. G., Carrier, J. S., Zahm, J. A., Harrison, S., Miller, M. P.. 2024-11-03. A coordinated kinase and phosphatase network regulates Stu2 recruitment to yeast kinetochores. https://doi.org/10.1101/2024.11.01.621564

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