CENTROSOME PROTEIN PERICENTRIN

中心体蛋白 中心周蛋白

• 批准号: 2749998
• 负责人: STEPHEN J DOXSEY
• 金额: $ 19.85万
• 依托单位: UNIV OF MASSACHUSETTS MED SCH WORCESTER
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-08-01 至 2000-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2749998
• 关键词: Xenopus; binding proteins; cell cycle; centrosome; cytoskeleton; fluorescence microscopy; immunoprecipitation; microtubules; protein biosynthesis; protein sequence; protein structure function; tissue /cell culture

The long term objective of this work is to develop a molecular understanding of the process by which centrosomes organize microtubules. The spatial orientation of microtubules is crucial for their function and is controlled by the centrosome. The centrosome has been recalcitrant to investigation so the basis for its function and regulation are unknown. We have discovered a highly conserved centrosome protein called pericentrin that is essential for cell division and organization of the microtubule cytoskeleton. Pericentrin provides us with a unique opportunity to understand centrosome function at the molecular level. A recent advance in our laboratory suggests that we will be able to identify and reconstitute the components of nucleation. We are now in an excellent position to: 1. Test the hypothesis that pericentrin is involved in centrosome assembly and define the sequence requirements for function. 2. Test the hypothesis that pericentrin assembles onto centrosomes in a cell cycle-specific phosphorylation-dependent manner. 3. Test the hypothesis that pericentrin binding proteins are involved in microtuble nucleation. Significant progress has been made toward th goals of this proposal. We have raised antibodies that block pericentrin function, produced mutant and wild type pericentrin cDNAs and fusion proteins and developed a novel in vitro assay for reconstituting centrosome function. Since that first submission of this proposal, we identified a pericentrin binding partner, perturbed centrosome structure using mutant pericentrin, and made the remarkable observation that pericentrin antibody-beads promote assembly of microtubules. These recent advances, molecular tools, and assays will allow us to make rapid progress in understanding the properties and function of pericentrin and centrosomes in general. Understanding centrosome function is important for several reasons. As the organizer of the microtubule cytoskeleton, the centrosome is a pivotal player in many fundamental cellular processes. Given its importance in the assembly of the mitotic spindle during cell division, the centrosome is a potential regulator of cell growth and a coordinator of morphogenesis. Centrosome- mediated microtubule polarity is critical for organizing the cytoplasm for intracellular vesicular transport and positioning of organelles. Despite its central position at the focus of microtubules and its central role in the life of a cell, the centrosome is poorly understood. Our preliminary results suggest that pericentrin will allow us to make important contributions in an area of biology that has been difficult to study. We are especially excited about the future prospect of realizing our long term goal of reconstituting centrosome function in vitro from individual components.

这项工作的长期目标是开发一种分子 理解中心体组织微管的过程。 微管的空间取向对其功能和功能至关重要 由中心体控制。中心体一直顽固地 因此，其功能和调控的依据尚不清楚。我们 发现了一种高度保守的中心体蛋白，称为周中心素 这对细胞分裂和微管的组织是必不可少的 细胞骨架。百菌素为我们提供了一个独特的机会 在分子水平上理解中心体的功能。最近在…方面的进展 我们的实验室表明，我们将能够识别和重建 成核的成分。我们现在处于一个极好的位置，可以： 1.检验周中心素参与中心体组装的假说 并定义了功能的顺序要求。 2.检验周中心素与中心体结合的假设。 细胞周期特异性磷酸化依赖方式。 3.检验围集素结合蛋白参与的假说 微管成核。 在实现这一提案的目标方面取得了重大进展。我们 产生了阻断周中心素功能的抗体，产生了突变和 野生型外周蛋白的cDNA和融合蛋白，并开发了一种新的 中心体功能重建的体外实验。从第一次开始 在提交这项提案时，我们确定了一个外周中心素结合伙伴， 利用突变的周丝点蛋白干扰中心体结构，并使 周球蛋白抗体微球促进集落形成的显著观察 微管。这些最新的进展、分子工具和分析将 使我们能够在理解这些属性和 周丝点蛋白和中心体的一般功能。理解 中心体功能之所以重要，有几个原因。作为本次活动的组织者 作为中心体的微管细胞骨架在许多生物系统中起着关键作用 基本的细胞过程。鉴于它在大会上的重要性 有丝分裂的纺锤体在细胞分裂过程中，中心体是一个潜在的 细胞生长的调节者和形态发生的协调者。中心体- 介导的微管极性是组织细胞质的关键 细胞器的胞内囊泡运输和定位。尽管 它在微管中心的位置及其在体内的中心作用 细胞的生命，即中心体，人们对此知之甚少。我们的预赛 结果表明，围集素将使我们能够使重要的 在一个一直难以研究的生物学领域做出了贡献。我们 对实现我们的长期目标的未来前景特别兴奋 体外从个体重建中心体功能的目标 组件。