A study on kinetochore-spindle microtubule attachment : Ndc80 and TACC-TOG/MAPs

During mitosis, spindle microtubules nucleate from the spindle pole body (SPB; centrosome in humans) and capture the chromosomes at the kinetochores. Many players, including the Ndc80/Hec1 outer kinetochore complex, the TACC-TOG microtubule-associated proteins, and various motor molecules, have been implicated in this process. Mis-regulation (e.g. overexpression) of these proteins leads to chromosome segregation errors and aneuploidy, a hallmark of human cancer. Despite this knowledge, the underlying molecular mechanisms by which these proteins function and interact remain largely elusive. Targeting of Alp7/TACC to the SPB is essential for mitotic spindle assembly. Using Schizosaccharomyces pombe as a model system, I have identified the SPB- targeting sequence within Alp7/TACC and the conserved pericentrin-like protein Pcp1 as one of the loading platforms for Alp7/TACC at the SPB. Upon successful spindle assembly, the spindle microtubules elongate and capture the kinetochores, which is a crucial step for faultless chromosome segregation. Having isolated and characterised a novel ndc80 mutant (ndc80-NH12), I have uncovered the roles of the Ndc80 internal loop in binding to the Alp7/TACC-Alp14/TOG complex. This binding ensures proper kinetochore-microtubule attachment and subsequently recruits the kinesin-8-protein phosphatase I (Klp5-Klp6-PP1) complex to the kinetochores. This recruitment in turn silences the spindle assembly checkpoint (SAC) and promotes timely and rapid chromosome movement during anaphase A. These results unveil the importance of the Ndc80 internal loop and its binding partners in regulating kinetochore-microtubule attachment and controlling ordered mitotic progression. Intriguingly, by overexpressing different Ndc80 constructs, I have found a possible causal link between the Ndc80 internal loop and tumourigenesis. I propose that the Ndc80 loop, when overexpressed, would sequester away its binding partners, thus disrupting their normal functions and leading to aneuploid formation. My results provide new insights into the molecular organisation at the kinetochore-microtubule interface, and have significant implications for future therapeutics and drug development.