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The spindle assembly checkpoint (SAC) is an evolutionarily conserved surveillance mechanism that delays anaphase onset and mitotic exit in response to the lack of kinetochore attachment. The target of the SAC is the E3 ubiquitin ligase anaphase-promoting complex (APC) bound to its Cdc20 activator. The Cdc20/APC complex is in turn required for sister chromatid separation and mitotic exit through ubiquitin-mediated proteolysis of securin, thus relieving inhibition of separase that unties sister chromatids. Separase is also involved in the Cdc-fourteen early anaphase release (FEAR) pathway of nucleolar release and activation of the Cdc14 phosphatase, which regulates several microtubule-linked processes at the metaphase/anaphase transition and also drives mitotic exit. Here, we report that the SAC prevents separation of microtubule-organizing centers (spindle pole bodies [SPBs]) when spindle assembly is defective. Under these circumstances, failure of SAC activation causes unscheduled SPB separation, which requires Cdc20/APC, the FEAR pathway, cytoplasmic dynein, and the actin cytoskeleton. We propose that, besides inhibiting sister chromatid separation, the SAC preserves the accurate transmission of chromosomes also by preventing SPBs to migrate far apart until the conditions to assemble a bipolar spindle are satisfied.

Faithful chromosome transmission requires establishment of sister chromatid cohesion during S phase, followed by its removal at anaphase onset. Sister chromatids are tethered together by cohesin, which is displaced from chromosomes through cleavage of its Mcd1 subunit by the separase protease. Separase is in turn inhibited, up to this moment, by securin. Budding yeast cells respond to morphogenetic defects by a transient arrest in G2 with high securin levels and unseparated chromatids. We show that neither securin elimination nor forced cohesin cleavage is sufficient for anaphase in these conditions, suggesting that other factors contribute to cohesion maintainance in G2. We find that the protein phosphatase PP2A bound to its regulatory subunit Cdc55 plays a key role in this process, uncovering a new function for PP2ACdc55 in controlling a noncanonical pathway of chromatid cohesion removal.

We report the characterization of the dominant-negative CLA4t allele of the budding yeast CLA4 gene, encoding a member of the p21-activated kinase (PAK) family of protein kinases, which, together with its homologue STE20, plays an essential role in promoting budding and cytokinesis. Overproduction of the Cla4t protein likely inhibits both endogenous Cla4 and Ste20 and causes a delay in the onset of anaphase that correlates with inactivation of Cdc20/anaphase-promoting complex (APC)–dependent proteolysis of both the cyclinB Clb2 and securin. Although the precise mechanism of APC inhibition by Cla4t remains to be elucidated, our results suggest that Cla4 and Ste20 may regulate the first wave of cyclinB proteolysis mediated by Cdc20/APC, which has been shown to be crucial for activation of the mitotic exit network (MEN). We show that the Cdk1-inhibitory kinase Swe1 is required for the Cla4t-dependent delay in cell cycle progression, suggesting that it might be required to prevent full Cdc20/APC and MEN activation. In addition, inhibition of PAK kinases by Cla4t prevents mitotic exit also by a Swe1-independent mechanism impinging directly on the MEN activator Tem1.