ABSTRACT
In budding yeast, the nucleolus serves as the site to sequester Cdc14, a phosphatase essential for mitotic exit. Nucleolar proteins Tof2, Net1, and Fob1 are required for this sequestration. Although it is known that these nucleolar proteins are SUMOylated, how SUMOylation regulates their activity remains unknown. Here, we show that Tof2 exhibits cell-cycle-regulated nucleolar delocalization and turnover. Depletion of the nuclear small ubiquitin-like modifier (SUMO) protease Ulp2 not only causes Tof2 polySUMOylation, nucleolar delocalization, and degradation but also leads to Cdc14 nucleolar release and activation. This outcome depends on polySUMOylation and the activity of downstream enzymes, including SUMO-targeted ubiquitin ligase and Cdc48/p97 segregase. We further developed a system to tether SUMO machinery to Tof2 and generated a SUMO-deficient tof2 mutant, and the results indicate that Tof2 polySUMOylation is necessary and sufficient for its nucleolar delocalization and degradation. Together, our work reveals a polySUMO-dependent mechanism that delocalizes Tof2 from the nucleolus to facilitate mitotic exit.
ABSTRACT
The conserved chromosomal passenger complex (CPC) consists of Ipl1Aurora-B, Sli15INCENP, Bir1Survivin, and Nbl1Borealin, and localizes at the kinetochore/centromere to correct kinetochore attachment errors and to prevent checkpoint silencing. After anaphase entry, the CPC moves from the kinetochore/centromere to the spindle. In budding yeast, CPC subunit Sli15 is phosphorylated by both cyclin-dependent kinase (CDK) and Ipl1 kinase. Following anaphase onset, activated Cdc14 phosphatase reverses Sli15 phosphorylation imposed by CDK to promote CPC translocation. Although abolished Sli15 phosphorylation imposed by Ipl1 also causes CPC translocation, the regulation of Ipl1-imposed Sli15 phosphorylation remains unclear. In addition to Sli15, Cdc14 also dephosphorylates Fin1, a regulatory subunit of protein phosphatase 1 (PP1), to enable kinetochore localization of Fin1-PP1. Here, we present evidence supporting the notion that kinetochore-localized Fin1-PP1 likely reverses Ipl1-imposed Sli15 phosphorylation to promote CPC translocation from the kinetochore/centromere to the spindle. Importantly, premature Fin1 kinetochore localization or phospho-deficient sli15 mutation causes checkpoint defects in response to tensionless attachments, resulting in chromosome missegregation. In addition, our data indicate that reversion of CDK- and Ipl1-imposed Sli15 phosphorylation shows an additive effect on CPC translocation. Together, these results reveal a previously unidentified pathway to regulate CPC translocation, which is important for accurate chromosome segregation.
