Kinetochore Specification and Function

着丝粒规格及功能

• 批准号: 9214331
• 负责人: Arshad Desai
• 金额: $ 37.62万
• 依托单位: LUDWIG INSTITUTE FOR CANCER RES LTD
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-06-01 至 2018-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9214331
• 关键词: Address; Attenuated; Basic Science; Binding; Biological Assay; Caenorhabditis elegans; Cancer cell line; Cell Cycle Progression; Cell division; Cells; Centromere; Chemotherapy-Oncologic Procedure; Chromosomal Instability; Chromosome Segregation; Chromosomes; Complex; Congenital Abnormality; Coupled; Data; Development; Dynein ATPase; Embryonic Development; Engineering; Ensure; Exhibits; Fatigue; Fertility Disorders; Genetic; Genome; Goals; Human; In Vitro; Kinetochores; Lateral; Lead; MXD1 gene; Mad1 protein; Malignant Neoplasms; Measures; Mechanics; Mediating; Microtubules; Mitosis; Mitotic Chromosome; Molecular; Motor; Mutation; Nuclear Pore; Organism; Pathway interactions; Phenotype; Phosphotransferases; Process; Proteins; Recruitment Activity; Role; Set protein; Signal Pathway; Signal Transduction; Sister Chromatid; Site; Work; anaphase-promoting complex; base; cancer cell; chromosome loss; cohesion; daughter cell; defined contribution; design; in vivo; mutant; novel therapeutic intervention; novel therapeutics; prevent; protein complex; public health relevance; scaffold; segregation; therapy resistant; tumorigenesis

DESCRIPTION (provided by applicant): Accurate partitioning of the replicated genome to daughter cells during cell division is essential for the development and propagation of all living organisms. Errors in genome distribution are a hallmark of cancer cells and the machinery involved in this process is targeted in cancer chemotherapy. Thus, elucidating the mechanisms ensuring accurate genome distribution will inform our understanding of the genesis of cancer and guide development of new therapeutic strategies. A major player in genome distribution is the kinetochore, the machine built on the centromere regions of chromosomes during mitosis to generate a dynamic end- coupled interface with spindle microtubules. The mechanics at this interface are tightly integrated with a signaling pathway, termed the spindle checkpoint, which prevents cell cycle progression until all chromosomes are connected to the spindle. Microtubule binding and checkpoint signaling are coordinated at the kinetochore by the conserved Knl1/Mis12 complex/Ndc80 complex (KMN) protein network. Within this protein set, the Ndc80 complex interacts directly with microtubule ends. The work proposed here has three goals: 1) to elucidate the mechanisms ensuring accurate formation of Ndc80-mediated kinetochore-microtubule attachments, 2) to define kinetochore-independent essential functions of core checkpoint pathway components, and 3) to determine if enhancing cohesion fatigue is a viable strategy to target cancer cell division. These goals will be addressed in 4 specific aims. Aim 1 will focus on understanding the formation of Ndc80-mediated end-coupled attachments, which are accelerated by lateral capture of microtubules by kinetochore-localized dynein, the major minus end-directed microtubule motor in cells. The proposed work will investigate crosstalk we discovered between the kinetochore dynein module and the Ndc80 complex and determine its significance to chromosome segregation fidelity. Aim 2 will address poorly understood essential functions of the conserved Bub1 kinase and the Ndc80 complex subunit Nuf2 in the formation of kinetochore- microtubule attachments. Aim 3 will focus on the provocative idea, based on our recent findings, that there are essential functions of the spindle checkpoint proteins Mad1 and Mad2 independent of their role in kinetochore- based checkpoint signaling. This aim exploits mechanism-based engineering of the Mad1/Mad2 complex in an organismal context to elucidate the basis for the severe developmental and fertility defects associated with loss of Mad1/Mad2 in C. elegans, which we have shown are independent of kinetochore-localization dependent checkpoint signaling. Finally, Aim 4 builds on our observation that sister chromatids of chromosomes attached to the spindle ultimately become unglued in an uncoordinated manner when mitosis is prolonged; this traps the cell in an aberrant state, termed cohesion fatigue, from which it cannot recover. We will broadly assess if the propensity for cohesion fatigue is increased in cancer cell lines, as suggested by preliminary data, and determine whether enhancing cohesion fatigue could serve as a strategy to target cancer cell division.

描述（由申请人提供）：在细胞分裂过程中将复制的基因组准确分配给子细胞对于所有活生物体的发育和繁殖至关重要。基因组分布的错误是癌细胞的一个标志，而参与这一过程的机制是癌症化疗的目标。因此，阐明确保准确的基因组分布的机制将有助于我们了解癌症的起源并指导新治疗策略的开发。基因组分布的主要参与者是着丝粒，它是在有丝分裂期间建立在染色体着丝粒区域上的机器，用于与纺锤体微管产生动态末端耦合界面。该界面的机制与信号通路紧密集成，称为纺锤体检查点，它阻止细胞周期进展，直到所有染色体都连接到纺锤体。微管结合和检查点信号传导在动粒处由保守的 Knl1/Mis12 复合物/Ndc80 复合物 (KMN) 蛋白网络协调。在该蛋白质组中，Ndc80 复合物直接与微管末端相互作用。这里提出的工作有三个目标：1）阐明确保 Ndc80 介导的着丝粒-微管附着的准确形成的机制，2）定义核心检查点通路成分的独立于着丝粒的基本功能，3）确定增强内聚疲劳是否是针对癌细胞分裂的可行策略。这些目标将通过 4 个具体目标来实现。目标 1 将重点了解 Ndc80 介导的末端偶联附着物的形成，这种附着物通过动粒定位动力蛋白（细胞中主要的负末端定向微管马达）横向捕获微管而加速。拟议的工作将研究我们在着丝粒动力蛋白模块和 Ndc80 复合体之间发现的串扰，并确定其对染色体分离保真度的重要性。目标 2 将解决保守的 Bub1 激酶和 Ndc80 复合体亚基 Nuf2 在着丝粒-微管附着物形成中的基本功能知之甚少的问题。根据我们最近的发现，目标 3 将重点关注一个令人兴奋的想法，即纺锤体检查点蛋白 Mad1 和 Mad2 的基本功能与其在基于动粒的检查点信号传导中的作用无关。这一目标利用在有机体背景下对 Mad1/Mad2 复合物进行基于机制的工程，以阐明与秀丽隐杆线虫中 Mad1/Mad2 缺失相关的严重发育和生育缺陷的基础，我们已经证明，这种缺陷独立于动粒定位依赖的检查点信号传导。最后，目标 4 建立在我们的观察之上，即当有丝分裂延长时，附着在纺锤体上的染色体的姐妹染色单体最终会以不协调的方式脱胶；这会使细胞陷入一种异常状态，称为内聚疲劳，并且无法从中恢复。我们将广泛评估癌细胞系中的内聚疲劳倾向是否增加（如初步数据所示），并确定增强内聚疲劳是否可以作为针对癌细胞分裂的策略。