Mitotic exit in budding yeast is dependent on correct orientation of the mitotic spindle along the cell polarity axis. When accurate positioning of the spindle fails, a surveillance mechanism named the Spindle Position Checkpoint (SPOC) prevents cells from exiting mitosis. Mutants with a defective SPOC become multinucleated and lose their genomic integrity. Yet, a comprehensive understanding of the SPOC mechanism is missing. In this study, we identified the type 1 protein phosphatase, Glc7, in association with its regulatory protein Bud14 as a novel checkpoint component. We further showed that Glc7-Bud14 promotes dephosphorylation of the SPOC effector protein Bfa1. Our results suggest a model in which two mechanisms act in parallel for a robust checkpoint response: first, the SPOC kinase Kin4 isolates Bfa1 away from the inhibitory kinase Cdc5 and second, Glc7-Bud14 dephosphorylates Bfa1 to fully activate the checkpoint effector.

Saccharomyces cerevisiae, also known as the budding yeast, orients and elongates its mitotic spindle along its polarity axis in order to segregate one copy of its genomic DNA to the daughter cell. When accurate positioning of the mitotic spindle fails, a surveillance mechanism, named the Spindle Position Checkpoint (SPOC), prevents cells from exiting mitosis unless the spindle orientation is corrected. Such mitotic arrest provides cells time to align their spindle correctly before cell division is completed. Mutants with a defective SPOC loss their genomic integrity, become multiploid and aneuploid. Thus, SPOC is a crucial checkpoint for the budding yeast. Yet, a comprehensive understanding of how SPOC mechanism works is budmissing. In this study, we identified Bud14 as a novel checkpoint protein. We showed that the mitotic exit inhibitory function Bud14 requires its association with the type 1 protein phosphatase, Glc7. Cells bearing versions of Bud14 that cannot interact with Glc7 or a temperature sensitive mutant of Glc7 were SPOC deficient. Our data indicate that Glc7-Bud14 inhibits mitotic exit by promoting dephosphorylation of Bfa1 during anaphase and limiting the levels of Bfa1 at the spindle pole bodies. Our results support a model in which Glc7-Bud14 works parallel to the SPOC kinase Kin4 in regulating Bfa1, the most downstream effector of SPOC that inhibits mitotic exit.

No abstract