Author ORCID Identifier
https://orcid.org/0000-0002-9258-5813
Date of Award
Winter 2023
Document Type
Thesis (Ph.D.)
Department or Program
Biochemistry and Cell Biology
First Advisor
Duane Compton
Second Advisor
Charles Barlowe
Third Advisor
Arminja Kettenbach
Abstract
The goal of mitosis is to achieve faithful chromosome segregation; ensuring that the daughter cells inherit equal numbers of chromosomes. This is vital to cell health and viability and if mis-regulated can result in birth defects and disease such as cancer. There are many intricately regulated processes that occur throughout mitosis to achieve proper chromosome segregation, and one such example is the dynamic attachments formed between cytoskeletal structures, known as microtubules, and chromosomes, the carriers of genetic material. These attachments occur at structures called kinetochores, and the microtubules attached here are referred to as kinetochore-microtubules (k-MTs). These k-MTs are inherently dynamically unstable. One parameter of this dynamic instability is depolymerization (shrinking of the microtubule), and in the following sections I have conducted experiments that explore the different processes that rely on this. In Chapter 2, I examine if chromosome poleward motion and the detachment of k-MTs are dependent on one another. Using live-cell imaging of three different cell lines and targeted manipulation of proteins involved in regulating the stability of k-MT attachments, I find that poleward chromosome motion and detachment for error correction are independent processes. In Chapter 3, I perform feasibility experiments to improve the resolution of our photoactivation protocol from a population-based measurement to a single kinetochore-fiber’s
(k-fiber) detachment rate. As established in the literature and discussed in detail in Chapter 2, one role of depolymerization is to promote detachment of microtubules from chromosomes. Our current photoactivation technique allows us to measure the detachment rate as an average of microtubules in a population. However, it remains unclear what variation exists among detachment rates of single k-fibers within that population, or if there are differences between correctly and incorrectly attached microtubules. Through an iterative process, I have improved the resolution of our photoactivation to be able to measure the half-life (our read-out for detachment rate) of single k-fibers. From these experiments we have learned more about the nature of how microtubule depolymerization contributes to poleward chromosome movement and error correction to ensure faithful chromosome segregation during cell division.
Recommended Citation
Parks, Melissa K., "Measuring how kinetochore-microtubule detachment contributes to chromosome movement and the correction of attachment errors" (2023). Dartmouth College Ph.D Dissertations. 123.
https://digitalcommons.dartmouth.edu/dissertations/123