ROLE OF AUGMIN IN HUMAN MITOTIC SPINDLES

AUGMIN 在人有丝分裂纺锤体中的作用

• 批准号: 8362560
• 负责人: Charles Kamasaki
• 金额: $ 3.19万
• 依托单位: UNIVERSITY OF COLORADO
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-05-01 至 2012-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8362560
• 关键词: 3-Dimensional; Cell physiology; Cells; Cellular Structures; Centrosome; Chromosome Segregation; Data; Electron Microscopy; Functional Imaging; Funding; Grant; Human; Laboratories; Life; Metaphase; Microtubules; Minus End of the Microtubule; Mitosis; Mitotic spindle; Modeling; National Center for Research Resources; Play; Principal Investigator; Research; Research Infrastructure; Resources; Role; Source; Testing; United States National Institutes of Health; cost; electron tomography; protein complex; reconstruction

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. The mitotic spindle, a bipolar microtubule array, plays a critical role in chromosome segregation during mitosis. Conventionally, the centrosome is considered to be th emajor microtubule nucleation center in the spindle. However, through live imaging and functional studies, several laboratories including ours have recently proposed a new model in which many spindle microtubules are generated inside the spindle, and therefore, minus-ends of the microtubules are distributed throughout the spindle. Furthermore, we have identified a protein complex 'augmin' that is responsible for the intra-spindle microtubule nucleation, in which new microtubules are possibly nucleated from the existing microtubules ("branched nucleation") (Goshima et al. 2007, 2008; Uehara et al. 2009, 2010). However, these models have not been proven, because the distribution of minus-ends have not been determined by electron microscopy in human cells. In this study, we would like to test the models through directly visualizing the minus-ends of spindle microtubules using electron tomography. Specifically, we wish to perform 1) Tomographic reconstruction of 3-D data and 3-D modeling for the metaphase spindle of human U2OS cells. 2) the same analysis for the U2OS cells depleted of augmin, which resulted in reduced microtubule numbers in the metaphase spindle, and compare with the control cells.

该副本是利用资源的众多研究子项目之一 由NIH/NCRR资助的中心赠款提供。对该子弹的主要支持 而且，副投影的主要研究员可能是其他来源提供的 包括其他NIH来源。 列出的总费用可能 代表subproject使用的中心基础架构的估计量， NCRR赠款不直接向子弹或副本人员提供的直接资金。 两极微管阵列的有丝分裂纺锤体在有丝分裂过程中在染色体分离中起关键作用。通常，中心体被认为是主轴中的Emajor微管成核中心。但是，通过实时成像和功能研究，包括我们的一些实验室最近提出了一个新模型，其中许多纺锤微管在主轴内部产生，因此，微管的负端分布在整个纺锤体中。此外，我们已经确定了一种蛋白质复合物“ augmin”，该蛋白质复合物是主轴内微管成核的，其中可能与现有微管（“分支成核”）成核新的微管（Goshima et al。2007，2008; Uehara et etal。2009，2010年）。但是，这些模型尚未得到证明，因为尚未通过电子显微镜在人类细胞中确定负端的分布。在这项研究中，我们希望通过使用电子断层扫描直接可视化纺锤微管的负端来测试模型。具体而言，我们希望执行1）对人U2OS细胞的中期纺锤体的3-D数据的层析成像重建和3-D建模。 2）对Augmin耗尽的U2OS细胞的相同分析，从而导致中期纺锤体中的微管数减少，并与对照细胞进行比较。