Cdc6 is sequentially regulated by PP2A-Cdc55, Cdc14, and Sic1 for origin licensing in <i>S. cerevisiae</i>.

Philip, Jasmin; Örd, Mihkel; Silva, Andriele; Singh, Shaneen; Diffley, John Fx; Remus, Dirk; Loog, Mart; Ikui, Amy E · Elife · 2022

basic_science · Level V

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Abstract

Cdc6, a subunit of the pre-replicative complex (pre-RC), contains multiple regulatory cyclin-dependent kinase (Cdk1) consensus sites, SP or TP motifs. In <i>Saccharomyces cerevisiae</i>, Cdk1 phosphorylates Cdc6-T7 to recruit Cks1, the Cdk1 phospho-adaptor in S phase, for subsequent multisite phosphorylation and protein degradation. Cdc6 accumulates in mitosis and is tightly bound by Clb2 through N-terminal phosphorylation in order to prevent premature origin licensing and degradation. It has been extensively studied how Cdc6 phosphorylation is regulated by the cyclin-Cdk1 complex. However, a detailed mechanism on how Cdc6 phosphorylation is reversed by phosphatases has not been elucidated. Here, we show that PP2A<sup>Cdc55</sup> dephosphorylates Cdc6 N-terminal sites to release Clb2. Cdc14 dephosphorylates the C-terminal phospho-degron, leading to Cdc6 stabilization in mitosis. In addition, Cdk1 inhibitor Sic1 releases Clb2·Cdk1·Cks1 from Cdc6 to load Mcm2-7 on the chromatin upon mitotic exit. Thus, pre-RC assembly and origin licensing are promoted by phosphatases through the attenuation of distinct Cdk1-dependent Cdc6 inhibitory mechanisms.

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