Centrosome biogenesis and copy number control

中心体生物发生和拷贝数控制

• 批准号: 9242655
• 负责人: Andrew Jon Holland
• 金额: $ 31.19万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-01 至 2020-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9242655
• 关键词: Acute; Address; Architecture; Biogenesis; Cell division; Cell physiology; Cells; Cellular Structures; Centrosome; Chromosome Segregation; Chromosomes; Cilia; Data; Data Reporting; Disease; Dose; Ensure; Environment; Feedback; Flagella; Genome Stability; Genomic Instability; Goals; Health; Human; Interphase Cell; Intracellular Transport; Laboratories; Lead; Light; Malignant - descriptor; Malignant Neoplasms; Mediating; Methodology; Methods; Microtubule-Organizing Center; Microtubules; Mitotic spindle; Molecular; Organelles; PLK1 gene; Pharmacology; Phase I Clinical Trials; Phosphorylation; Phosphorylation Site; Phosphotransferases; Play; Production; Proteins; Recruitment Activity; Regulation; Research; Role; S Phase; Signal Transduction; Structure; System; Testing; Work; cancer cell; cancer therapy; cell growth; cell motility; in vivo; kinase inhibitor; kinetosome; novel; overexpression; public health relevance; scaffold; spatiotemporal; tool; tumor

 DESCRIPTION (provided by applicant): The long-term goal of our research is to understand the molecular mechanisms that control centrosome biogenesis. Centrosomes are microtubule-nucleating organelles that play a central role in most microtubule- related functions, including cell motility, intracellular transport and chromosome segregation. In addition, the centrosome forms the basis of the basal body, which is required for the formation of cilia and flagella, and i therefore crucial for cells to both sense their environment and transduce signals. Centrosomes are present as a single copy at the beginning of the cycle and duplicate once during S phase to ensure only two copies are present to organize the poles of the mitotic spindle. Abnormalities in centrosome number are commonly observed in human cancer cells, where extra centrosomes lead to chromosome segregation errors that are thought to drive tumor formation. Understanding the mechanism by which cells achieve the once per cycle duplication of the centrosome is therefore an important fundamental question of considerable relevance to human health. Polo-like kinase 4 (Plk4) has emerged as the central, dose-dependent regulator of centrosome duplication. Suppressing Plk4 inhibits centrosome duplication, while Plk4 overexpression leads to the production of too many centrosomes. However, we understand little about how this kinase functions; and in particular, the critical Plk4 targets that control centrosome biogenesis remain to be identified. Our proposed research seeks to establish the mechanisms through which Plk4 orchestrates and coordinates centrosome biogenesis. Previous efforts to study Plk4 have been hampered because tools to specifically and rapidly manipulate Plk4 function have not been available. In this application we have overcome this limitation by developing two complementary methodologies that allow us to regulate Plk4 levels and activity in living cells. Using these tools we will establish the direct effect of altering Plk4 levels/actiity and distinguish between kinase-dependent and scaffolding functions of Plk4 in centrosome biogenesis. In Aim 1, we will study the effect of rapid loss/inhibition of Plk4 on cell growth and centrosome structure. In our preliminary data we have identified a highly conserved centrosome protein as a key Plk4 substrate required for centrosome duplication. In Aim 2, we propose to characterize how Plk4-mediated phosphorylation of this substrate contributes to centrosome assembly. These studies are relevant for understanding the normal regulation of centrosome assembly and for furthering ongoing efforts to target Plk4 in cancer therapy.

 描述（由申请人提供）：我们研究的长期目标是了解控制中心体生物发生的分子机制。中心体是微管成核细胞器，在大多数微管相关功能中发挥核心作用，包括细胞运动、细胞内运输和染色体分离。此外，中心体形成基体的基础，这是形成纤毛和鞭毛所必需的，因此对于细胞感知环境和转导信号至关重要。中心体在周期开始时以单个副本存在，并在 S 期复制一次，以确保仅存在两个副本来组织有丝分裂纺锤体的两极。在人类癌细胞中经常观察到中心体数量的异常，其中额外的中心体导致染色体分离错误，这被认为是驱动肿瘤形成的原因。因此，了解细胞实现中心体每个周期复制一次的机制是与人类健康密切相关的重要基本问题。 Polo 样激酶 4 (Plk4) 已成为中心体复制的中心、剂量依赖性调节因子。抑制 Plk4 会抑制中心体复制，而 Plk4 过度表达会导致产生过多中心体。然而，我们对这种激酶的功能知之甚少。特别是，控制中心体生物发生的关键 Plk4 靶标仍有待确定。我们提出的研究旨在建立 Plk4 协调和协调中心体生物发生的机制。之前研究 Plk4 的努力受到了阻碍，因为还没有专门且快速操纵 Plk4 功能的工具。在此应用中，我们通过开发两种互补的方法克服了这一限制，这两种方法使我们能够调节活细胞中的 Plk4 水平和活性。使用这些工具，我们将确定改变 Plk4 水平/活性的直接影响，并区分 Plk4 在中心体生物发生中的激酶依赖性功能和支架功能。在目标 1 中，我们将研究 Plk4 快速丢失/抑制对细胞生长和中心体结构的影响。在我们的初步数据中，我们已经确定了一种高度保守的中心体蛋白作为中心体复制所需的关键 Plk4 底物。在目标 2 中，我们建议描述 Plk4 介导的该底物磷酸化如何促进中心体组装。这些研究对于了解中心体组装的正常调节以及进一步推进癌症治疗中针对 Plk4 的持续努力相关。