Hsu, K.-S. (2012) Molecular mechanism of the kinetochore-microtubule attachment in fission yeast. Doctoral thesis, UCL (University College London).
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Kinetochore capture by the spindle microtubule is a crucial step required for chromosome biorientation. Any errors in this process would result in production of aneuploid progenies, a hallmark of human cancer. The Ndc80 complex, a conserved outer kinetochore complex, comprising four components, Ndc80/Hec1, Nuf2, Spc24 and Spc25, constitutes one of the core microtubulebinding sites within the kinetochore. Despite this knowledge, molecular mechanism by which this complex contributes to establishment of correct bipolar attachment of the kinetochore to the spindle microtubule remains largely elusive. Here I show that the conserved internal loop of fission yeast Ndc80 directly binds the Dis1/TOG microtubule-associated protein, thereby coupling spindle microtubule dynamics with kinetochore capture. Ndc80 loop mutant proteins fail to recruit Dis1 to kinetochores, imposing unstable attachment and frequent spindle collapse. In these mutants, mitotic progression is halted attributable to spindle assembly checkpoint activation, and chromosomes remain in the vicinity of the spindle poles without congression. dis1 deletion precisely phenocopies the loop mutants. Furthermore, forced targeting of Dis1 to the Ndc80 complex rescues loop mutant’s defects. I propose that Ndc80 comprises two microtubule-interacting interfaces; the N-terminal region directly binds the microtubule lattice, whilst the internal loop interacts with the plus end of microtubules via Dis1/TOG. Therefore, my results provide a crucial insight into how the Ndc80 complex establishes stable bipolar attachment to the spindle microtubule. In addition, integrity of the Ndc80 complex is also required for the kinetochore tethering to the SPB during interphase. Loss of Ndc80 results in kinetochore declustering in the nucleus, and those scattered kinetochores are found to colocalise with cytoplasmic microtubules and the tubulin nucleation factor γ- TuC. Further investigation will be needed to understand how Ndc80 affects the γ-TuC distribution and/or how Ndc80 and the γ-TuC participate in the SPBkinetochore anchorage in interphase.
|Title:||Molecular mechanism of the kinetochore-microtubule attachment in fission yeast|
|Open access status:||An open access version is available from UCL Discovery|
|Additional information:||A copyright restricted article with supplementary material has been removed from the digital copy of this thesis|
|UCL classification:||UCL > School of Life and Medical Sciences > Faculty of Life Sciences|
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