RESUMEN
The spindle assembly checkpoint (SAC) is the major surveillance system that ensures that sister chromatids do not separate until all chromosomes are correctly bioriented during mitosis. Components of the checkpoint include Mad1, Mad2, Mad3 (BubR1), Bub3, and the kinases Bub1, Mph1 (Mps1), and Aurora B. Checkpoint proteins are recruited to kinetochores when individual kinetochores are not bound to spindle microtubules or not under tension. Kinetochore association of Mad2 causes it to undergo a conformational change, which promotes its association to Mad3 and Cdc20 to form the mitotic checkpoint complex (MCC). The MCC inhibits the anaphase-promoting complex/cyclosome (APC/C) until the checkpoint is satisfied. SAC silencing derepresses Cdc20-APC/C activity. This triggers the polyubiquitination of securin and cyclin, which promotes the dissolution of sister chromatid cohesion and mitotic progression. We, and others, recently showed that association of PP1 to the Spc7/Spc105/KNL1 family of kinetochore proteins is necessary to stabilize microtubule-kinetochore attachments and silence the SAC. We now report that phosphorylation of the conserved MELT motifs in Spc7 by Mph1 (Mps1) recruits Bub1 and Bub3 to the kinetochore and that this is required to maintain the SAC signal.
Asunto(s)
Proteínas de Ciclo Celular/metabolismo , Proteínas Cromosómicas no Histona/metabolismo , Cinetocoros/metabolismo , Puntos de Control de la Fase M del Ciclo Celular/fisiología , Proteínas Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas de Schizosaccharomyces pombe/metabolismo , Schizosaccharomyces/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas Cromosómicas no Histona/genética , Regulación Fúngica de la Expresión Génica , Fosforilación/fisiología , Proteínas Quinasas/genética , Proteínas Serina-Treonina Quinasas/genética , Schizosaccharomyces/genética , Proteínas de Schizosaccharomyces pombe/genéticaRESUMEN
Type 1 phosphatase (PP1) antagonizes Aurora B kinase to stabilize kinetochore-microtubule attachments and to silence the spindle checkpoint. We screened for factors that exacerbate the growth defect of Δdis2 cells, which lack one of two catalytic subunits of PP1 in fission yeast, and identified Nsk1, a novel protein required for accurate chromosome segregation. During interphase, Nsk1 resides in the nucleolus but spreads throughout the nucleoplasm as cells enter mitosis. Following dephosphorylation by Clp1 (Cdc14-like) phosphatase and at least one other phosphatase, Nsk1 localizes to the interface between kinetochores and the inner face of the spindle pole body during anaphase. In the absence of Nsk1, some kinetochores become detached from spindle poles during anaphase B. If this occurs late in anaphase B, then the sister chromatids of unclustered kinetochores segregate to the correct daughter cell. These unclustered kinetochores are efficiently captured, retrieved, bioriented, and segregated during the following mitosis, as long as Dis2 is present. However, if kinetochores are detached from a spindle pole early in anaphase B, then these sister chromatids become missegregated. These data suggest Nsk1 ensures accurate chromosome segregation by promoting the tethering of kinetochores to spindle poles during anaphase B.
Asunto(s)
Anafase , Proteínas de Ciclo Celular/metabolismo , Segregación Cromosómica , Cinetocoros/metabolismo , Proteínas de Schizosaccharomyces pombe/metabolismo , Schizosaccharomyces/citología , Schizosaccharomyces/genética , Proteínas de Ciclo Celular/genética , Dineínas/metabolismo , Microtúbulos/metabolismo , Fosfoproteínas Fosfatasas/genética , Fosfoproteínas Fosfatasas/metabolismo , Fosforilación , Estabilidad Proteica , Transporte de Proteínas , Proteínas Tirosina Fosfatasas/genética , Proteínas Tirosina Fosfatasas/metabolismo , Schizosaccharomyces/metabolismo , Proteínas de Schizosaccharomyces pombe/genética , Huso Acromático/genética , Huso Acromático/metabolismoRESUMEN
The spindle checkpoint is the prime cell-cycle control mechanism that ensures sister chromatids are bioriented before anaphase takes place. Aurora B kinase, the catalytic subunit of the chromosome passenger complex, both destabilizes kinetochore attachments that do not generate tension and simultaneously maintains the spindle checkpoint signal. However, it is unclear how the checkpoint is silenced following chromosome biorientation. We demonstrate that association of type 1 phosphatase (PP1(Dis2)) with both the N terminus of Spc7 and the nonmotor domains of the Klp5-Klp6 (kinesin-8) complex is necessary to counteract Aurora B kinase to efficiently silence the spindle checkpoint. The role of Klp5 and Klp6 in checkpoint silencing is specific to this class of kinesin and independent of their motor activities. These data demonstrate that at least two distinct pools of PP1, one kinetochore associated and the other motor associated, are needed to silence the spindle checkpoint.