Dynein function at the vertebrate kinetochore

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Cell division (mitosis) is one of the most fundamental biological processes. It is remarkable as every division must take place with exceedingly high fidelity. Infidelity of the process leads to various afflictions, including cell death. Each cell’s duplicated genetic information – contained within chromosomes – must be equally (and faithfully) divided between the two daughter cells and the high degree of accuracy in this process is the consequence of a complex network of safety mechanisms that ensures mistakes are corrected prior to the completion of mitosis. The molecular machinery that ensures high-fidelity chromosome inheritance from mother to daughter cells includes an elaborate arrangement of filamentous structures called microtubules, and large protein-based structures assembled upon the chromosomes called kinetochores. Proper division of the genetic material requires that all duplicated chromosomes physically connect to microtubules through their kinetochores, allowing the chromosomes to become organized and aligned at the center of the cell in preparation for division. Kinetochores must make mechanically stable attachments to microtubules, and it is through these stable connections that duplicated sister chromosomes are both driven to the middle of the cell, then pulled apart towards the end of mitosis. Cells contain a monitoring system (a “checkpoint”) that prevents cells from exiting mitosis until all kinetochores are properly attached to microtubules such that they are poised to faithfully divide the chromosomes. While it is known that kinetochores monitor and regulate their own attachment status, how the attachment status of each kinetochore is relayed to the checkpoint machinery is unknown. The goals of this research project are to determine how a molecular motor, called dynein, affects and facilitates: (1) chromosome alignment, and (2) mitotic checkpoint signaling. The results from this project will have a significant impact on our understanding of mitotic cell division, and how the underlying molecular processes ensure it takes place with high fidelity. The Broader Impacts of the work include the inherent importance of this process to all multi-cellular life on the planet, together with outreach work that will be carried out at the community level and to elementary school students. The goal of this project is to understand how the microtubule motor protein dynein functions at kinetochores to promote faithful segregation of chromosomes during cell division. Cells possess complex mechanisms that ensure chromosome segregation occurs with remarkably high fidelity. During cell division, microtubules that comprise the mitotic spindle facilitate separation of sister chromatids through direct attachments to kinetochores, large macromolecular assemblies built upon centromeric DNA. Cells employ at least two critical mechanisms to minimize errors during this process: (A) The spindle assembly checkpoint prevents mitotic progression until all chromosomes have established proper kinetochore-microtubule attachments. Effectors of this checkpoint accumulate on improperly or unattached kinetochores, and consequently transmit a “wait anaphase” signal. Only upon establishment of proper attachments are these proteins evicted from kinetochores, which silences the inhibitory signal, thereby promoting anaphase onset. (B) The error correction pathway promotes the release of incorrect kinetochore-microtubule attachments, thereby allowing them to “reset” and form new, correct attachments. A key effector of both these processes is the microtubule motor protein dynein, which (1) transports checkpoint effectors away from kinetochores upon proper microtubule attachment, and (2) transports erroneously attached chromosomes to spindle poles, where they have a high likelihood of being corrected. The researchers will use a combination of in vitro and in-cell approaches to understand the role for dynein in both of these critical mitotic processes.This research is funded by the Cellular Dynamics and Function program in the Division of Molecular and Cellular Biosciences in the Directorate of Biological Sciences.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.