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In budding yeast, if the spindle becomes mispositioned, cells prevent exit from mitosis by inhibiting the mitotic exit network (MEN). The MEN is a signaling cascade that localizes to spindle pole bodies (SPBs) and activates the phosphatase Cdc14. There are two competing models that explain MEN regulation by spindle position. In the 'zone model', exit from mitosis occurs when a MEN-bearing SPB enters the bud. The 'cMT-bud neck model' posits that cytoplasmic microtubule (cMT)-bud neck interactions prevent MEN activity. Here we find that 1) eliminating cMT- bud neck interactions does not trigger exit from mitosis and 2) loss of these interactions does not precede Cdc14 activation. Furthermore, using binucleate cells, we show that exit from mitosis occurs when one SPB enters the bud despite the presence of a mispositioned spindle. We conclude that exit from mitosis is triggered by a correctly positioned spindle rather than inhibited by improper spindle position.

Summary: Background: Budding yeast cells that enter the developmental path of meiosis do not commit to finishing meiosis until after prophase I and the realization of such meiosis-specific events as pairing of homologous chromosomes and initiation of recombination. If the meiosis-inducing signal is withdrawn prior to commitment, cells exit meiosis and return to mitosis. The timing of this transition poses a singular problem for maintaining genome integrity. Cells in meiotic prophase have already replicated their DNA, but they have not undergone the morphological changes intrinsic to mitosis, including budding. Successful re-entry into mitosis requires that these cells bud but not rereplicate their DNA, reversing the normal order of mitosis. This study focuses on the cellular mechanisms that permit this dramatically altered order of cell-cycle events. Results: By developing a microfluidics assay to monitor individual cells, we show that the successful transition from meiotic prophase to mitosis requires the modulation of Cdk1 activity to coordinate cell-cycle events. The S. cerevisiae Wee1 homolog Swe1 prevents the formation of multinucleate cells by restraining M phase CDK activity to allow bud formation prior to nuclear division. The remaining S phase CDK activity promotes bud formation and prevents origin licensing so that DNA cannot rereplicate between bud formation and nuclear division. Once a bud has formed, M phase CDK drives cells through a normal mitotic division. Conclusions: Our study uncovers the essential requirement of Swe1 to modulate CDK activity to coordinate cell-cycle events and maintain genome integrity during the transition from meiotic prophase to mitosis. [Copyright &y& Elsevier]