| Mitosis is a process that cells utilize to accurately and faithfully separate genetic information into two daughter cells. A successful mitosis requires that the chromosomes, the genetic information carrier, attach to a properly organized bipolar microtubule-based spindle. The attachment of the chromosomes is mediated by the sister kinetochores at the chromosome constriction, which help contribute to accurate chromosome segregation. After the nuclear contents are segregated, the cell must also divide its cytoplasm in a process called cytokinesis. The organization of functional microtubule arrays in different mitotic stages and the movement of chromosomes are both mainly driven by motor proteins, which utilize chemical energy from ATP to produce physical forces. This thesis focuses on understanding how mitotic spindle organization contributes to proper chromosome motility. One subfamily of motor proteins, the Kinesin-14 (HSET/XCTK2), cross-links and slides between microtubules, and plays distinct roles during spindle assembly, chromosome congression, and cytokinesis. We found that HSET perturbation affected spindle morphology through its sliding activity in a K-fiber independent way. This function is regulated by the association of its N-terminal tail domain with nuclear import factors importin alpha/beta. Meanwhile, the activity of HSET is also important for chromosome congression. In spindles with disrupted K-fibers and compromised HSET, chromosomes are still able to align at the metaphase plate in a CENP-E dependent manner, demonstrating that chromosomes have the ability to congress in the absence of k-fibers. However, chromosomes in this condition cannot be stably maintained at metaphase plate, demonstrating that K-fibers are not needed for congression but are needed to maintain chromosome alignment. Furthermore, we found that HSET localized on the midzone microtubules and midbody during early cytokinesis, and knockdown of HSET disrupted midzone microtubule organization, which resulted in the mislocalization of various midbody proteins. Our results demonstrate that the sliding and cross-linking activity of HSET is critical to maintain proper spindle organization, which in turn is critical for both proper chromosome congression and segregation. |
