Regulation of mitosis by kinetochore-dependent mechanisms

S. cerevisiae CLASP， Stu1以相位依赖的方式定位于微管和（通过其第一个TOGL结构域）着丝点，从而协调有丝分裂。在中期，Stu1定位于着丝点并以张力依赖的方式稳定着丝点微管。此外，Stu1定位于极间微管晶格，并通过其第二个TOGL结构域的活性稳定中期纺锤体，很可能是通过交联极间微管。在后期，Stu1与着丝点分离，从而可能使着丝点微管缩短，这是后期a的典型特征。Stu1也与MT晶格分离，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，并且很可能减轻微管交联，从而允许后期滑动。在前期中期，Stu1被隔离在独立的着丝点上。这可以防止纺锤体过早形成，并可能促进这些着丝点的捕获。Stu1定位对未附着着丝点的依赖性使人想起那些促进纺锤体组装检查点（SAC）蛋白在未附着着丝点定位的依赖性。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)中期和后期着丝点和微管定位的调控。2)与Stu1第一个TOGL结构域相互作用的着丝点蛋白的鉴定。3)蛋白激酶Mps1的底物和磷酸化位点，该蛋白激酶对Stu1在未附着的着丝点上的隔离至关重要。4)导致未附着着丝点隔离效应的机制（诱导构象变化？）。此外，我们计划进一步研究Stu1在有丝分裂中的作用：1)当Stu1定位于中期着丝点时，促进着丝点微管稳定的机制。2) Stu1促进非附着着丝点捕获的机制。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.