Doctor of Philosophy, The Ohio State University, 2013, Molecular, Cellular and Developmental Biology
Studies from our lab demonstrated that a cold-sensitive gamma-tubulin mutant allele, mipAD159, causes defects in the coordination of late mitotic events at restrictive temperatures with observed phenotypes such as abnormal chromosome segregation and inhibition of anaphase A (Prigozhina et al., 2004). These abnormalities are not a result of defects in microtubule nucleation since gamma-tubulin localized normally to the spindle pole body (SPB), microtubules were abundant and mitotic spindle formation and elongation appeared to be normal (Prigozhina et al., 2004). Nayak et al. (2010) further examined gamma-tubulin's role in mitotic regulation and determined that, at restrictive temperatures, mipAD159 caused a failure of accumulation of cyclin B, cyclin dependent kinase 1 (Cdk1) and the phosphatase, Ancdc14, in a subset of nuclei. These nuclei were removed from the cell cycle while other nuclei in the same cell accumulated these proteins and cycled normally. Extensive analysis revealed that this failure of accumulation was due to a nuclear autonomous failure of inactivation of the anaphase promoting complex/cyclosome (APC/C) sometime between late mitosis and S phase.
The two projects I have focused on are directed toward further elucidating gamma-tubulin's role in cell cycle regulation. Many mitotic regulatory proteins are known to localize to the SPB, or the centrosome, its functional equivalent in higher organisms, in mitosis. Therefore, gamma-tubulin might be interacting with such proteins and such interactions might be altered in strains carrying mipAD159. I decided to focus on the spindle assembly checkpoint (SAC) proteins Mad2, Mps1, Bub3, BubR1 and Cdc20. I identified the A. nidulans homolog of each, created fluorescent protein fusions, and observed them in vivo by spinning disk confocal microscopy. I found that these proteins are physically separate from each other in interphase, keeping the SAC inactive until mitosis, when they are all at the S (open full item for complete abstract)
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Committee: Harold Fisk (Advisor); Berl Oakley (Advisor); Stephen Osmani (Committee Member); Hay-Oak Park (Committee Member)
Subjects: Biology; Cellular Biology; Genetics; Molecular Biology