Regulation of mitosis by kinetochore-dependent mechanisms

有丝分裂的调控依赖于动粒的机制酿酒酵母环，Stu1，通过定位到微管和(通过它的第一个TOGL结构域)以一种阶段依赖的方式协调有丝分裂。在中期，Stu1定位于动粒，并以张力依赖的方式稳定动粒微管。此外，Stu1定位于极间微管的晶格上，并通过其第二个TOGL结构域的活性和极间微管的交联来稳定中期纺锤体。在后期，Stu1从动粒解离，因此可能允许动粒微管缩短，这是后期A的典型现象。Stu1也从MT晶格解离，并定位于纺锤体中部。这仍然允许微管通过第二个TOGL结构域稳定，并最有可能减轻微管的交联性，从而允许后期滑动。在早中期，Stu1被隔离在独立的动点上。这防止了早熟纺锤体的形成，并可能有助于捕捉这些动点。Stu1定位对独立的动点的依赖使人想起那些促进纺锤体组装检查点(SAC)蛋白在独立的动点定位的依赖。此外，Stu1通过第一个TOGL结构域与SAC蛋白Mad1相互作用。在未来，我们计划研究Stu1定位的原理：1)中期与后期动粒和微管定位的调节。2)与Stu1的第一个TOGL结构域相互作用的动粒蛋白的鉴定。3)蛋白激酶Mps1的底物和磷酸化位点，它是STU1在独立的动粒上隔离所必需的。4)导致独立动粒隔离效应(诱导构象变化？)的机制。此外，我们计划研究Stu1在有丝分裂中的表现和推测的作用：1)当Stu1定位于中期动粒时，促进动粒微管稳定的机制。2)Stu1促进独立动点捕获的机制。3)Stu1-Mad1相互作用的作用。

Regulation of mitosis by kinetochore-dependent mechanisms The S. cerevisiae CLASP, Stu1, orchestrates mitosis by localizing to microtubules and (via its first TOGL domain) to kinetochores in a phase-dependent manner. In metaphase Stu1 localizes to kinetochores and stabilizes kinetochore microtubules in a tension-dependent manner. Furthermore, Stu1 localizes to the lattice of interpolar microtubules and stabilizes the metaphase spindle via the activity of its second TOGL domain and most likely by crosslinking the interpolar microtubules. In anaphase Stu1 dissociates from kinetochores and thus possibly allows the shortening of kinetochore microtubules typical for anaphase A. Stu1 also dissociates from the MT lattice and localizes to the spindle midzone. This still allows microtubule stabilization via the second TOGL domain and most likely alleviates microtubule crosslinking to allow gliding in anaphase. In prometaphase Stu1 gets sequestered at unattached kinetochores. This prevents precocious spindle formation and possibly facilitates capturing of these kinetochores. The dependencies of the Stu1 localization to unattached kinetochores are reminiscent of those that facilitate the localization of spindle assembly checkpoint (SAC) proteins at unattached kinetochores. Moreover Stu1 interacts with the SAC protein Mad1 via the first TOGL domain. In the future we plan to investigate the principles of Stu1 localization: 1) The regulation of kinetochore and microtubule localization in metaphase versus anaphase. 2) The identification of the kinetochore proteins that interact with the first TOGL domain of Stu1. 3) The substrates and phosphorylation sites of the protein kinase Mps1 that is essential for the sequestering of Stu1 at unattached kinetochores. 4) The mechanism that leads to the sequestering effect at unattached kinetochores (induction of a conformational change?). Furthermore we plan to investigate the manifested and putative roles of Stu1 in mitosis: 1) The mechanism that facilitates stabilization of kinetochore microtubules when Stu1 localizes to metaphase kinetochores. 2) The mechanism by which Stu1 promotes capturing of unattached kinetochores. 3) The role of the Stu1-Mad1 interaction.