Generation of Diverse Centrosomes, Cilia and Flagellae During Development

发育过程中多种中心体、纤毛和鞭毛的产生

• 批准号: 10590581
• 负责人: David M Glover
• 金额: $ 50.21万
• 依托单位: CALIFORNIA INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-05-15 至 2025-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10590581
• 关键词: Affect; Applied Genetics; Biology; Brain; Cell Cycle; Cell Line; Cell division; Cells; Centrioles; Centrosome; Cerebral cortex; Cilia; Cilium Microtubule; Complex; Cultured Cells; Cytoplasm; Defect; Development; Disease; Drosophila genus; Eukaryota; Flagella; Foundations; Functional disorder; Future; G2 Phase; Generations; Genetic; Germ Lines; Growth; Heritability; Human; Inherited; Interphase; Lead; Leg; Length; Light; Malignant Neoplasms; Meiosis; Microcephaly; Microtubule Triplet; Microtubules; Mitosis; Molecular; Molecular Biology; Motion; Mutagenesis; Mutation; Nomenclature; Organ; Organelles; Pathology; Pathway interactions; Patients; Phosphorylation; Physiology; Pilot Projects; Procentriole; Process; Prognosis; Proteins; Research; Role; Set protein; Signal Transduction; Somatic Cell; Specific qualifier value; Spermatocytes; Spermatogenesis; Structure; Testing; Tissues; Work; brain abnormalities; cell type; ciliopathy; cilium biogenesis; genetic approach; human disease; imaginal disc; kinetosome; male; man; neurosensory; organizational structure; protein function; recruit; sperm cell; stem cells; structural biology

PROJECT SUMMARY Centrioles are required to template cilia and flagella and to organize centrosomes that nucleate cytoplasmic microtubules in interphase and dividing cells. Centrosomes lose their ability to organise cytoplasmic microtubules when they become basal bodies, which can template the formation of primary cilia for signalling or flagellae for propelling sperm. To understand these diverse roles, it necessary to study centrioles in the development of different cell types. Yet, their replication, structure and function has been largely studied in cultured cells. Here we take advantage of the genetic and developmental advantages of Drosophila to study centriole function in two cell types; cells destined to make neurosensory cilia in the leg imaginal disc and male germ line cells destined to generate sperm. Our focus is on the roles of a network of centriole proteins in which Ana3, counterpart of human ROTATIN and Rcd4, PPP1R35 in human cells, interact with each other and with a set of proteins required for various steps of the elongation and maturation of the centriole necessary for it to become a basal body. Specifically, we will: (i) apply molecular and structural biology approaches to determine how the Ana3 and Rcd4 proteins interact with each other and with other centriole proteins to specify different aspects of centriole assembly and function and how this is regulated by phosphorylation. We will test the functions of these interactions through directed mutagenesis. (ii) use genetic approaches to determine inter-dependencies of the networks that regulate centriole elongation in the conventional cell division cycles of cultured cells and the leg imaginal disc and in cells that develop neurosensory cilia. (iii) determine the differential roles of Rcd4 and Ana3 and their interacting network of proteins in the multiple steps leading to formation of the giant centrioles of spermatocytes and their sequential roles in generating primary cilia and subsequently sperm flagellae., Because centriole duplication pathways are highly conserved between Drosophila and man, our findings will shed light onto a wide range of human diseases including cancer, where centrosome defects indicate poor prognosis; heritable ciliopathies, which affect centrioles and cilia in a wide range of tissues; and in microcephaly, where defects in proteins such as ROTATIN, whose counterpart we study here, lead to abnormal brain development.

项目概要 中心粒需要模板纤毛和鞭毛并组织成核的中心体 间期和分裂细胞中的细胞质微管。中心体失去组织能力 当细胞质微管变成基体时，它们可以模板形成 用于发出信号的初级纤毛或用于推动精子的鞭毛。要了解这些不同的角色， 研究不同细胞类型发育过程中的中心粒是必要的。然而，它们的复制、结构 其功能已在培养细胞中得到大量研究。在这里，我们利用遗传和 果蝇研究两种细胞类型中心粒功能的发育优势；细胞注定要 使腿部成虫盘中的神经感觉纤毛和注定要产生精子的男性生殖细胞。 我们的重点是中心粒蛋白网络的作用，其中 Ana3（人类的对应物） 人类细胞中的 ROTATIN 和 Rcd4、PPP1R35 彼此相互作用并与一组蛋白质相互作用 中心粒伸长和成熟的各个步骤所需的 一个基体。具体来说，我们将：（i）应用分子和结构生物学方法来确定 Ana3 和 Rcd4 蛋白如何相互作用以及如何与其他中心粒蛋白相互作用以指定 中心粒组装和功能的不同方面以及磷酸化如何对其进行调节。我们 将通过定向诱变测试这些相互作用的功能。 (ii) 使用遗传方法 确定传统方法中调节中心粒伸长的网络的相互依赖性 培养细胞和腿部成象盘以及发育神经感觉细胞的细胞分裂周期 纤毛。 (iii) 确定 Rcd4 和 Ana3 的不同作用及其相互作用的蛋白质网络 导致精母细胞巨大中心粒形成的多个步骤及其顺序 在产生初级纤毛和随后的精子鞭毛中的作用。, 由于中心粒复制途径在果蝇和人类之间高度保守，我们的发现 将揭示包括癌症在内的多种人类疾病，其中中心体缺陷表明 预后不良；遗传性纤毛病，影响多种组织中的中心粒和纤毛；并在 小头畸形，例如 ROTATIN（我们在这里研究的对应蛋白）等蛋白质的缺陷会导致 大脑发育异常。