Regulation of the Bfa1p–Bub2p complex at spindle pole bodies by the cell cycle phosphatase Cdc14p

Abstract

The budding yeast mitotic exit network (MEN) is a GTPase-driven signal transduction cascade that controls the release of the phosphatase Cdc14p from the nucleolus in anaphase and thereby drives mitotic exit. We show that Cdc14p is partially released from the nucleolus in early anaphase independent of the action of the MEN components Cdc15p, Dbf2p, and Tem1p. Upon release, Cdc14p binds to the spindle pole body (SPB) via association with the Bfa1p–Bub2p GTPase activating protein complex, which is known to regulate the activity of the G protein Tem1p. Cdc14p also interacts with this GTPase. The association of the MEN component Mob1p with the SPB acts as a marker of MEN activation. The simultaneous binding of Cdc14p and Mob1p to the SPB in early anaphase suggests that Cdc14p initially activates the MEN. In a second, later step, which coincides with mitotic exit, Cdc14p reactivates the Bfa1p–Bub2p complex by dephosphorylating Bfa1p. This inactivates the MEN and displaces Mob1p from SPBs. These data indicate that Cdc14p activates the MEN in early anaphase but later inactivates it through Bfa1p dephosphorylation and so restricts MEN activity to a short period in anaphase.

The Role of the Polo Kinase Cdc5 in Controlling Cdc14 Localization

“…To distinguish between these possibilities, we compared the kinetics of Cdc14 release from the nucleolus and mitotic exit of esp1-1 or slk19⌬ mutants with that of esp1-1 or slk19⌬ mutants lacking SPO12 and BNS1. We could not test whether deletion of SPO12 and BNS1 enhanced the phenotype of cells lacking CDC5 as Cdc14 release from the nucleolus is completely abolished in such cells (Pereira et al, 2002;Stegmeier et al, 2002;. Inactivation of SPO12 and BNS1 enhanced the defect in Cdc14 release from the nucleolus and mitotic spindle disassembly of, both, esp1-1 and slk19⌬ mutants (Figure 3, A and B).…”

“…Cdc5, besides being a FEAR network component, is also a member of the Mitotic Exit Network (Shou et al, 1999;Visintin et al, 1999;Hu et al, 2001;Lee et al, 2001) that functions in parallel to the FEAR network to promote Cdc14 release from the nucleolus (Pereira et al, 2002;Stegmeier et al, 2002;. The finding that overexpression of CDC5 led to ectopic activation of Dbf2 kinase ( Figure 7A) raised the possibility that the suppression of FEAR network mutants by high levels of Cdc5 was due to Cdc5's parallel activating function in the MEN.…”

“…Inactivation of FEAR network components prevents Cdc14 release from the nucleolus during early anaphase, when the mitotic spindle is between 3 and 7 m in length. In contrast, inactivation of the MEN abolishes Cdc14 release from the nucleolus during late stages of anaphase, when the mitotic spindle measures 7-10 m in length (Pereira et al, 2002;Stegmeier et al, 2002;. MEN-dependent release of Cdc14 from the nucleolus is essential for exit from mitosis, because temperature-sensitive mutants in MEN components fail to exit from mitosis but arrest in telophase.…”

“…The Cdc Fourteen Early Anaphase Release (FEAR) network promotes release of Cdc14 from the nucleolus during early anaphase and the Mitotic Exit Network (MEN) maintains Cdc14 in this released state during late stages of anaphase and telophase (Shou et al, 1999;Visintin et al, 1999;Pereira et al, 2002;Stegmeier et al, 2002;. Inactivation of FEAR network components prevents Cdc14 release from the nucleolus during early anaphase, when the mitotic spindle is between 3 and 7 m in length.…”

“…CDC5 is also a component of the FEAR network, because the gene is not only required for the release of Cdc14 from the nucleolus during late stages of anaphase but also during early anaphase (Pereira et al, 2002;Stegmeier et al, 2002). Other components of the FEAR network include ESP1, SLK19, and SPO12 (Stegmeier et al, 2002).…”