Regulation of mitosis by kinetochore-dependent mechanisms

• Applicant: Privatdozent Dr. Johannes Lechner
• Subject Area: Cell Biology
• Term: from 2014 to 2022
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 268663799

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.

• DFG Programme: Research Grants