3D Structure and Function of the Mammalian Kinetochore

哺乳动物动粒的 3D 结构和功能

• 批准号: 0110821
• 负责人: Bruce McEwen
• 金额: $ 65.64万
• 依托单位: Health Research Incorporated/New York State Department of Health
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-09-01 至 2006-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0110821&HistoricalAwards=false
• 关键词: 3D; Structure; Function; Mammalian; Kinetochore

In eukaryotic cell division, the partitioning of daughter chromosomes into the daughter cells is achieved by attachment of the chromosomes to microtubules which then move the chromosomes apart prior to cytokinesis. The site of attachment of the microtubules to the chromosomes is a morphologically and biochemically distinctive chromosomal region called the kinetochore. Recent work by Dr. McEwen, employing state-of-the-art preservation and high resolution imaging technologies, has challenged the "trilaminar" morphological model of the kinetochore that has been accepted in the field for the past thiry years, and which was probably inaccurate due to fixation artifacts inherent in conventional fixation protocols. Now,Dr. McEwen's long-range goal is to define a high-resolution structural map of the mammalian kinetochore that explains and predicts functional mechanisms. The research combines state-of-the-art methods of light and electron microscopy with molecular approaches to compute the structural map and obtain functional characterization of key kinetochore components. The current project has four objectives, listed below.Research Objective 1. Establish a prototype high-resolution 3D structural map of the mammalian kinetochore. 1a: Identify key features of the kinetochore and look for evidence of a subunit structure. 1b. Test the hypothesis that the kinetochore has discrete structural domains that dynamically respond to microtubule attachment and deprivation, and that an enlarged outer domain is correlated with promiscuous microtubule capture and a high incidence of aneuploidy.Research Objective 2. Test three hypotheses concerning the molecular role of CENP-E in mitosis. 2a: CENP-E has a role in maintaining the stability of kinetochore microtubule attachments. 2b: CENP-E prevents monooriented chromosomes from over-traveling poleward into the spindle pole. 2c: CENP-E is not required for release of the anaphase onset checkpoint.Research objective 3. Test the hypotheses that CENP-E function is redundant with ZW10/dynein in binding kinetochore microtubules, and opposes dynein in poleward motion.Research objective 4. Test the hypothesis that the kinetochore is sequentially assembled, with kinetochore proteins CENP-C and BUB1 being required for complete assembly.This research project will provide important and significant new information on the complexities of kinetochore structure and function. It will fill a major gap in our present knowledge of how molecular components of the kinetochore are arranged and how they interact with other components of the cell division apparatus.

在真核细胞分裂中，子代染色体分配到子代细胞中是通过将染色体附着到微管上来实现的，然后微管在胞质分裂之前将染色体移开。微管与染色体的附着位点是形态学和生物化学上独特的染色体区域，称为动粒。 McEwen 博士最近的工作采用了最先进的保存和高分辨率成像技术，对过去三十年来该领域所接受的动粒“三层”形态模型提出了挑战，该模型可能由于传统固定方案中固有的固定伪影而不准确。 现在，博士。麦克尤恩的长期目标是定义哺乳动物动粒的高分辨率结构图，以解释和预测功能机制。该研究将最先进的光学和电子显微镜方法与分子方法相结合，计算结构图并获得关键动粒成分的功能表征。当前项目有四个目标，如下所列。 研究目标 1. 建立哺乳动物动粒高分辨率 3D 结构图原型。 1a：识别着丝粒的关键特征并寻找亚基结构的证据。 1b.检验以下假设：动粒具有离散的结构域，可动态响应微管附着和剥夺，并且扩大的外部结构域与混杂的微管捕获和非整倍性的高发生率相关。研究目标 2. 检验有关 CENP-E 在有丝分裂中的分子作用的三个假设。 2a：CENP-E 具有维持着丝粒微管附着稳定性的作用。 2b：CENP-E 防止单向染色体过度向极移动进入纺锤体极。 2c：后期开始检查点的释放不需要 CENP-E。研究目标 3。测试 CENP-E 功能在结合着丝粒微管方面与 ZW10/动力蛋白冗余的假设，并且在极向运动中对抗动力蛋白。研究目标 4。测试着丝粒蛋白 CENP-C 和 BUB1 顺序组装的假设 完整组装所需的。该研究项目将为动粒结构和功能的复杂性提供重要且重要的新信息。 它将填补我们目前关于动粒分子成分如何排列以及它们如何与细胞分裂装置其他成分相互作用的知识的一个主要空白。