IDENTIFICATION OF INTERACTING PROTEINS OF SPINDLY

Spindly 相互作用蛋白的鉴定

• 批准号: 8171402
• 负责人: Arshad Desai
• 金额: $ 0.24万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-01 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8171402
• 关键词: Aneuploidy; Binding; Biology; Characteristics; Chromosome Segregation; Chromosomes; Complex; Computer Retrieval of Information on Scientific Projects Database; Cultured Cells; Dynein ATPase; Ensure; Funding; Goals; Grant; Immunoprecipitation; Institution; Kinetochores; Laboratories; Mass Spectrum Analysis; Microtubules; Mitosis; Mitotic; Mitotic Spindle Apparatus; Nocodazole; Proteins; Recruitment Activity; Research; Research Personnel; Resources; Role; Source; Transgenes; United States National Institutes of Health; Work; daughter cell; dynactin; human tissue; insight; mutant; neoplastic cell; tissue culture

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The kinetochore facilitates chromosome segregation in mitosis by interacting with microtubules of the mitotic spindle apparatus. Proper functioning of the kinetochore is essential to avoid uneven distribution of chromosomes to daughter cells, which leads to aneuploidy, a common characteristic of tumors cells. The dynein/dynactin complex localizes to the kinetochore in mitosis and has been implicated in both kinetochore-microtubule attachment and in the silencing of the spindle checkpoint, which ensures that mitotic exit does not occur in the presence of unattached kinetochores. Work in our laboratory has identified an outer kinetochore component, called Spindly, which has a key role in recruiting the dynein/dynactin complex to unattached kinetochores, but the molecular interactions involved are not known. Our goal is to gain insight into the mechanism of dynein/dynactin recruitment to kinetochores by identifying the dynein/dynactin subunits that interact with Spindly. We will use mass spectrometry after immunoprecipitation of Myc-tagged Spindly transgenes from nocodazole-arrested human tissue culture cells to identify candidate Spindly binding partners. As a negative control, we will use a Spindly point mutant which is unable to recruit dynein/dynactin to kinetochores, but whose own kinetochore recruitment is unaffected. In addition to dynein/dynactin subunits, we expect the mass spectrometric analysis to reveal the components that recruit Spindly itself to kinetochores. Mass spectrometry analysis is therefore crucial to answer fundamental questions in kinetochore biology.

该副本是利用众多研究子项目之一 由NIH/NCRR资助的中心赠款提供的资源。子弹和 调查员（PI）可能已经从其他NIH来源获得了主要资金， 因此可以在其他清晰的条目中代表。列出的机构是 对于中心，这不一定是调查员的机构。 动力学通过与有丝分裂纺锤体的微管相互作用，促进有丝分裂的染色体分离。 动力学的正确功能对于避免染色体对子细胞的分布不均匀至关重要，该细胞导致非整倍性（肿瘤细胞的常见特征）。 Dynein/dynactin复合物在有丝分裂中定位于动力学，并与动力学微动物的附着和纺锤体检查点的沉默有关，这确保在没有附属的动物学体的存在下不会发生有丝分裂的出口。 我们实验室中的工作已经确定了一种称为Spindly的外部动力学成分，该组件在将动力蛋白/dynactin络合物募集到未连接的动力学方面具有关键作用，但是所涉及的分子相互作用尚不清楚。 我们的目标是通过识别与Spindly相互作用的Dynein/Dynactin亚基来深入了解动力蛋白/Dynactin募集到动力学的机制。 我们将使用质谱法对诺科唑逮捕的人类组织培养细胞的MYC标记的晶状体转基因进行免疫沉淀后，以鉴定候选候选伴侣。 作为一个负面对照，我们将使用一个无法募集动力蛋白/dynactin的固定点突变体到动物学，但其自己的动脉募集不受影响。 除了Dynein/dynactin亚基外，我们预计质谱分析可以揭示募集本身为动力学的组成部分。 因此，质谱分析对于回答动物学生物学中的基本问题至关重要。