Dynein function at the vertebrate kinetochore

细胞分裂（有丝分裂）是最基本的生物过程之一。这是值得注意的，因为每一次分裂都必须极其忠实地进行。这个过程的不忠会导致各种各样的痛苦，包括细胞死亡。每个细胞复制的遗传信息——包含在染色体中——必须在两个子细胞之间平等地（并且忠实地）分配，这一过程的高度准确性是一个复杂的安全机制网络的结果，该网络确保在有丝分裂完成之前纠正错误。确保染色体从母体向子细胞高保真遗传的分子机制包括精细排列的丝状结构（称为微管）和组装在染色体上的大型蛋白质结构（称为着丝点）。遗传物质的正常分裂需要所有复制的染色体通过着丝点与微管物理连接，从而使染色体在细胞中心排列整齐，为分裂做准备。着丝点必须在机械上稳定地附着在微管上，正是通过这些稳定的连接，复制的姐妹染色体都被驱动到细胞中间，然后在有丝分裂结束时被拉开。细胞包含一个监控系统（一个“检查点”），它阻止细胞退出有丝分裂，直到所有着丝点都正确地附着在微管上，这样它们就可以忠实地分裂染色体。虽然已知着丝点监测和调节它们自己的附着状态，但每个着丝点的附着状态如何传递给检查点机制是未知的。本研究项目的目标是确定一种称为动力蛋白的分子马达如何影响和促进：(1)染色体排列，(2)有丝分裂检查点信号传导。这个项目的结果将对我们对有丝分裂细胞分裂的理解产生重大影响，以及潜在的分子过程如何确保它以高保真度发生。这项工作的更广泛影响包括这一进程对地球上所有多细胞生命的内在重要性，以及将在社区一级和对小学生开展的外展工作。本项目的目的是了解微管运动蛋白动力蛋白如何在着丝点上起作用，以促进细胞分裂过程中染色体的忠实分离。细胞具有复杂的机制，以确保染色体分离发生非常高的保真度。在细胞分裂过程中，组成有丝分裂纺锤体的微管通过直接附着在着丝点上促进姐妹染色单体的分离，着丝点是建立在着丝粒DNA上的大分子组装体。细胞至少采用两种关键机制来减少这一过程中的错误：(A)纺锤体组装检查点阻止有丝分裂进程，直到所有染色体都建立了适当的着丝点-微管附着。这个检查点的效应器积聚在不正确的或未连接的动着点上，并因此传递“等待后期”信号。只有在建立适当的附着物后，这些蛋白质才会从着丝点中被排出，从而使抑制信号沉默，从而促进后期的发生。(B)纠错途径促进不正确的着丝点-微管附着体的释放，从而允许它们“重置”并形成新的、正确的附着体。这两个过程的关键效应物是微管运动蛋白动力蛋白（dynein），它(1)在适当的微管附着时将检查点效应物从着丝点转运，(2)将错误附着的染色体转运到纺锤极，在那里它们有很高的被纠正的可能性。研究人员将结合体外和细胞内的方法来了解动力蛋白在这两个关键的有丝分裂过程中的作用。本研究由生物科学理事会分子和细胞生物科学部的细胞动力学和功能项目资助。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

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.