Mechanism and Regulation of Axonemal Dynein Arm Assembly in Motile Ciliated Epithelial Cells

运动纤毛上皮细胞轴丝动力蛋白臂组装的机制和调控

• 批准号: 10930194
• 负责人: ZHAOXIA SUN
• 金额: $ 57.65万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-23 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10930194
• 关键词: Acceleration; Amino Acids; Biochemical; Biological Assay; Biological Process; Biology; Candidate Disease Gene; Cell physiology; Cells; Cellular Membrane; Cellular biology; Characteristics; Cilia; Crowding; Cytoplasm; Cytosol; Data; Defect; Disease; Dynein ATPase; Environment; Epithelial Cells; Etiology; Extracellular Fluid; Fertility; Foundations; Future; Genes; Genetic; Genetic Diseases; Goals; Health; Human; Image; Infection; Intervention; Investigation; Knock-in; Left; Light; Liquid substance; LoxP-flanked allele; Lung diseases; Macromolecular Complexes; Mediating; Membrane; Messenger RNA; Microtubules; Modeling; Molecular; Molecular Chaperones; Motor; Mucous body substance; Mus; N-terminal; Names; Organism; Patients; Periodicity; Phase; Physiological; Primary Ciliary Dyskinesias; Proteins; Publishing; RNA; Recurrence; Regulation; Reproduction; Research; Respiratory Tract Infections; Ribosomes; Role; Shapes; Skin; System; Testing; Trachea; Translating; Translations; Xenopus; Zebrafish; appendage; arm; cell motility; ciliopathy; cilium biogenesis; cilium motility; fluid flow; insight; mouse genetics; novel; prevent; pulmonary function decline; symptom management; tool

Motile cilia beat rhythmically to propel cell movement or drive extracellular fluid flow. The functional importance of cilia motility in human health is highlighted by primary ciliary dyskinesia (PCD), a genetic disease caused by cilia motility defects. Patients with PCD display left-right asymmetry defect, reduced fertility and progressive lung disease. Currently there is no specific therapy for PCD and management of symptoms has been the main approach. The dynein arms that power cilia motility comprise multiple components that are pre-assembled in the cytosol and many genes associated with PCD encode components of these dynein arms. Intriguingly, however, a separate group includes proteins that reside in the cytosol and appear to be involved in the assembly of dynein arm subunits and they are called dynein arm assembly factors (DNAAFs). Interestingly, multiple DNAAFs are localized in droplet shaped cytosolic foci. However, the precise function of these foci and the precise molecular function of most DNAAFs remain poorly understood. In addition to the assembly of protein components, how mRNAs and the translation machinery are coordinated to supply dynein arm components stoichiometrically is also largely unknown. Based on extensive preliminary and published data, our central hypothesis is that the co- chaperone proteins Pontin and Reptin are core components of a novel membraneless cytosolic assemblage distinct from droplets formed through LLPS; they function to coordinate the translation, folding and assembly of axonemal dynein arm components and prevent aggregate formation. In this project, we will combine zebrafish genetics, mouse genetics and cultured tracheal cells to test our central hypothesis. We propose two specific aims to achieve this goal. In the first aim, we will dissect the molecular and cellular function of Pontin-Reptin foci in discrete steps in dynein arm assembly. In the second aim, we will characterize Pontin-Reptin foci and investigate the mechanism of foci formation. Successful completion of this project will not only provide a molecular framework for dynein arm assembly and the etiology of PCD, but also lay the foundation for future investigation into the regulation, and possible intervention, of dynein arm assembly and cilia motility under diverse physiological and disease conditions.

活动纤毛有节奏地跳动以推动细胞运动或驱动细胞外液流动。功能重要性 原发性纤毛运动障碍 (PCD) 突显了纤毛运动对人类健康的影响，这是一种由以下原因引起的遗传病 纤毛运动缺陷。 PCD 患者表现出左右不对称缺陷、生育能力下降和肺部进展 疾病。目前PCD尚无特效疗法，主要是症状管理 方法。为纤毛运动提供动力的动力蛋白臂由多个组件组成，这些组件预先组装在 胞浆和许多与 PCD 相关的基因编码这些动力蛋白臂的成分。然而有趣的是， 一个单独的组包括驻留在细胞质中并且似乎参与动力蛋白组装的蛋白质 臂亚基，它们被称为动力蛋白臂组装因子（DNAAF）。有趣的是，多个 DNAAF 定位于液滴状胞质病灶。然而，这些焦点的精确功能和精确的分子 大多数 DNAAF 的功能仍知之甚少。除了蛋白质成分的组装之外，如何 mRNA 和翻译机制相互协调，以化学计量提供动力蛋白臂成分 也很大程度上不为人所知。基于广泛的初步和已发表的数据，我们的中心假设是 伴侣蛋白 Pontin 和 Reptin 是新型无膜胞质组合的核心成分 与 LLPS 形成的液滴不同；它们的功能是协调平移、折叠和组装 轴丝动力蛋白臂成分并防止聚集体形成。在这个项目中，我们将结合斑马鱼 遗传学、小鼠遗传学和培养的气管细胞来检验我们的中心假设。我们提出两个具体方案 旨在实现这一目标。第一个目标是剖析 Pontin-Reptin 灶的分子和细胞功能 动力蛋白臂组装中的离散步骤。在第二个目标中，我们将描述 Pontin-Reptin 病灶的特征和 研究病灶形成的机制。该项目的成功完成不仅将为 动力蛋白臂组装的分子框架和PCD的病因学，也为未来奠定了基础 研究动力蛋白臂组装和纤毛运动的调节和可能的干预 不同的生理和疾病状况。