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Document Type

Campus-Only Access for Five (5) Years

Degree Name

Doctor of Philosophy (PhD)

Degree Program

Molecular and Cellular Biology

Year Degree Awarded

2018

Month Degree Awarded

May

First Advisor

Wei-Lih Lee

Second Advisor

Patricia Wadsworth

Subject Categories

Biochemistry | Biochemistry, Biophysics, and Structural Biology | Biology | Biophysics | Cell Biology | Genetics | Genetics and Genomics | Life Sciences | Molecular Biology | Molecular Genetics

Abstract

During development, metaphase spindles undergo large movement and/or rotation to determine the cell division axis. While it has been shown that spindle translocation is achieved by astral microtubules pulling and/or pushing the cortex, how metaphase spindle stability is maintained during translocation remains not fully understood. In budding yeast, our lab has previously proposed a model for spindle orientation wherein the mitotic spindle protein She1 promotes spindle translocation across the bud neck by polarizing cortical dynein pulling activity on the astral microtubules. Intriguingly, She1 exhibits dominant spindle localization throughout the cell cycle. However, whether She1 has any additional role on the spindle during translocation is unknown. Using function-separating alleles, live-cell fluorescence microscopy, biochemical and biophysical assays, I found that She1 ensures metaphase spindle integrity by crosslinking and stabilizing interpolar microtubules. Loss of this stabilizing activity leads to spindle defects characterized by an increase in interpolar microtubule turnover rate and a pronounced spindle bending/collapse phenotype. She1 binds microtubules via its C-terminal domain, and crosslinks microtubules possibly by self-association. The N-terminal domain contains a tandem newly identified PISH1/2 motif that can regulate Ipl1/Aurora targeting to the mitotic spindle, which, in turn, negatively regulates She1 crosslinking activity and localization on the spindle. My work reveals a new role for She1 in maintaining spindle integrity and offers insight into how metaphase spindles could be stabilized by a non-motor crosslinking microtubule-associated protein (MAP) during dynein-dependent spindle positioning in higher organisms.

Comments

The research manuscript related to this thesis work is published in the Journal of Cell Biology on Aug 9th, 2017.

http://jcb.rupress.org/content/216/9/2759

Available for download on Sunday, November 11, 2018

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