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Uncorrected defects in the kinetochore-microtubule interaction lead to chromosome mis-segregation and aneuploidy, a phenotype found in over 90% of cancers. Therefore cells have developed different strategies to ensure chromosome bipolar attachment occurs prior to anaphase entry. We provide evidence to suggest a previously uncharacterized mechanism in which budding yeast clusters kinetochores to facilitate chromosome bipolar attachment. Additionally, we have conducted a screen for genes required in the response to syntelic attachments, wherein both sister kinetochores are attached by microtubules emanating from a single spindle pole. The budding yeast kinetochore protein Slk19 has been previously shown to play a role in mitotic exit and spindle stability, but its function at the kinetochore has remained a mystery. We have found that, independently of the spindle, Slk19 clusters kinetochores and provides centromeric cohesion. Althoughslk19deletion fails to show a dramatic cell cycle delay in an unperturbed cell cycle, we found that upon disruption of the kinetochore-microtubule interaction,slk19mutants take a longer time to re-capture the kinetochore and show delayed chromosome bipolar attachment. Syntelic attachment occurs when sister kinetochores are bound to microtubules emanating from the same spindle pole. This fails to generate tension on a chromosome and triggers the tension checkpoint. Unfortunately, an approach to induce syntelic attachment has not been made available. The Cik1-Kar3 complex is a minus-end microtubule motor complex which has been shown to be involved in chromosome transport. Interestingly, we have recently found that conditional disruption of the Cik1-Kar3 complex leads to a higher frequency of chromosomes with syntelic attachment. Importantly, this disruption is not lethal to wild type cells, making it possible to identify genes required for the response to syntelic attachment. Therefore we conducted a screen by conditionally disrupting the Cik1-Kar3 complex in mutants that are synthetically lethal tocik1and/orkar3deletion. We speculate that some of these genes could be involved in checkpoint control, bipolar attachment, or error correction in the response to syntelic attachment. Our focus is on genes responsible for the tension checkpoint.
A Dissertation submitted to the Department of Biomedical Sciences in partial fulfillment of the requirements for the degree of Doctor of Philosophy.
Includes bibliographical references.
Yanchang Wang, Professor Directing Dissertation; Thomas Keller, III, University Representative; Myra Hurt, Committee Member; Akash Gunjan, Committee Member.
Florida State University
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