Characterizing the molecular mechanisms of centriole duplication, growth and maturation

表征中心粒复制、生长和成熟的分子机制

• 批准号: 10640273
• 负责人: Gregory Charles Rogers
• 金额: $ 56.24万
• 依托单位: UNIVERSITY OF ARIZONA
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-06-01 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10640273
• 关键词: Behavior Control; Biological Models; Cell Cycle; Cells; Centrioles; Centrosome; Cilia; Congenital Abnormality; Daughter; Drosophila genus; Drosophila polo protein; Elements; Equipment and supply inventories; Etiology; Eukaryotic Cell; Explosion; Funding; Goals; Growth; Image; Knowledge; Length; Malignant Neoplasms; Microtubules; Mission; Mitotic; Mitotic spindle; Molecular; Mothers; Organelles; PLK1 gene; Pathology; Phosphorylation; Phosphotransferases; Procentriole; Process; Proteins; Proteomics; Public Health; Publishing; Regulation; Research; Series; Shapes; Site; Structure; Ubiquitination; United States National Institutes of Health; cancer cell; ciliopathy; fascinate; functional genomics; human disease; overexpression; prevent; programs; protein complex; recruit; restraint; tumorigenesis

PROJECT SUMMARY/ABSTRACT Centrosomes are organelles used to build microtubule-based protein machines, including mitotic spindles and cilia. At the centrosome core lies a pair of `mother-daughter' centrioles, barrel-shaped structures that act as the duplicating elements of the organelle. Normally, the centriole pair duplicates only once each cell cycle and, during mitotic entry, centrioles recruit a shell of pericentriolar material (PCM) – a process called `maturation' – from which microtubules grow. Not only are they one of the largest protein complexes in eukaryotic cells but one of the most ancient of organelles, and have fascinated cell biologists since their discovery in the late 19th century. During the past 20 years, advances in imaging, proteomics and functional genomic screens have led to an explosion of discoveries in the centrosome field. At present, we have a complete inventory of the proteins comprising centrosomes. In our model system, Drosophila, centrosomes assemble from a surprisingly small number of components (approximately 20). Despite these advances, many important questions remain unanswered. Although only two conserved master-regulators, Polo kinase and Polo-like kinase 4 (Plk4), initiate centriole maturation and duplication, respectively, it is not known how they are activated specifically on centrioles. Also, what are the phosphorylation targets of these kinases and how do they promote centriole duplication and maturation? How are mother centrioles restrained to spawn only a single daughter once per cell cycle? How is centriole length controlled? Understanding these processes at the molecular level is important because alterations in centrosome function or number cause a number of serious pathologies, including birth defects, ciliopathies and cancer. Plk4 has been the centerpiece of our research program because it is both necessary and sufficient to induce centrosome overduplication (amplification) when overexpressed, a scenario observed in cancer cells. We have published a series of studies that have defined Plk4 regulation and identified several of its substrates. Notably, Plk4 utilizes multiple mechanisms of control to restrain its activity and prevent rampant centrosome overduplication, using an elaborate combination of autophosphorylation, ubiquitination and autoinhibition. We continue to pursue two overarching goals: 1) identifying the molecular mechanisms that suppress centrosome amplification (funded by R01 GM110166) and 2) characterizing the inherent mechanisms that govern centrosome function and duplication (funded by R01 GM126035). Building on our progress during the past five years, we propose to extend our studies that will define the mechanisms underlying the five sequential steps in the assembly process. Specifically, we will determine (i) how a single site of daughter centriole assembly is selected on mother centrioles, (ii) the composition of the pre-procentrioles and how it forms, (iii) how nascent daughter centrioles assemble, (iv) how centriole growth is controlled, and (v) the initial steps in centrosome maturation.

项目摘要/摘要 中心体是用于构建基于微管的蛋白质机器的细胞器，包括有丝分裂的纺锤体和 纤毛。在中心体的核心是一对“母女”中心，桶形的结构充当 细胞器的重复元素。通常，中心对仅重复一次，每个细胞周期和 在有丝分裂进入期间，Centrioles招募了围肠里材料（PCM）的外壳 - 一种称为“成熟”的过程 - 微管从中生长。它们不仅是真核细胞中最大的蛋白质复合物之一，而且是 自19世纪末发现以来，最古老的细胞器之一是他们着迷的细胞生物学家 世纪。在过去的20年中，成像，蛋白质组学和功能性基因组筛选的进展已导致 在中心体场的发现爆炸。目前，我们有蛋白质的完整库存 包括中心体。在我们的模型系统中，果蝇，中心体从一个令人惊讶的小 组件数量（约20）。尽管有这些进展，但仍有许多重要的问题 虽然只有两个保守的主调节剂Polo激酶和类似Polo样激酶4（PLK4）启动 Centriole成熟和重复，尚不清楚它们是如何被专门激活的 中心。同样，这些激酶的磷酸化靶标是什么？它们如何促进中心 重复和成熟？每个细胞只有一个女儿，如何仅将一个女儿限制为单个女儿 循环？中心长度如何控制？在分子水平上了解这些过程很重要 因为中心体功能或数量的改变会导致许多严重的病理，包括出生 缺陷，纤毛病和癌症。 PLK4一直是我们研究计划的核心 当过表达时，必要且足以诱导中心体过多（放大） 在癌细胞中观察到。我们发表了一系列研究，这些研究定义了PLK4调节并确定了 它的几个底物。值得注意的是，PLK4利用多种控制机制来限制其活性并防止 使用自磷酸化，泛素化和 自身抑制。我们继续追求两个总体目标：1）确定分子机制 抑制中心体扩增（由R01 GM110166资助）和2）表征继承机制 控制中心体功能和复制（由R01 GM126035资助）。基于我们在 在过去的五年中，我们建议扩展我们的研究，以定义五个的机制 组装过程中的顺序步骤。具体来说，我们将确定（i）女儿的单个站点 中心装配是在母中心列中选择的，（ii）孕前三环的组成及其形成方式， （iii）新生的女儿中央核心如何组装，（iv）中心元如何控制和（v）的初始步骤 中心体成熟。