Date of Award
Doctor of Philosophy (PhD)
Medicinal Chemistry and Molecular Pharmacology
Committee Member 1
Committee Member 2
Committee Member 3
Kinetochores are conserved protein complexes that bind the centromeres in replicated chromosomes to the mitotic spindle and then direct their segregation. To better comprehend Saccharomyces cerevisiae kinetochore function, we investigated the phospho-regulated dynamic interaction between the conserved kinetochore protein Cnn1CENP-T, the centromere region and the Ndc80 complex through the cell cycle. Cnn1 localizes to kinetochores at basal levels from G1 through metaphase but accumulates abruptly at anaphase onset. How Cnn1 is recruited and which activities regulate its dynamic localization is unclear. We show that Cnn1 harbors two kinetochore-localization activities: a C-terminal histone-fold domain that associates with centromere region, and a N-terminal Spc24/25-interaction sequence that mediates linkage to the microtubule-binding Ndc80 complex. We demonstrate that a previously established Ndc80 binding site in the N-terminus of Cnn1, Cnn160-84, should be extended to include flanking residues, Cnn125-91, to allow near maximal binding affinity to Ndc80. Cnn1 localization was proposed to depend on Mps1 kinase activity at Cnn1-S74 based on in vitro experiments demonstrating the Cnn1-Ndc80 complex interaction. We demonstrate that in G1 through metaphase, Cnn1 localizes via the histone-fold domain or a N-terminal Spc24/25-
Thapa, Kriti Shrestha, "Dissecting the Spatiotemporal Regulation of Yeast Kinetochore Interactions" (2016). Open Access Dissertations. 1284.