Dissecting alternate modes and regulation of ciliary motility

项目总结 这项研究的总体目标是通过解剖分子来增加我们对纤毛如何跳动的理解。 纤毛运动的分子水平机制及其调控。纤毛和鞭毛很保守，随处可见。 微管为基础的细胞器在细胞运动、液体运输、感觉、细胞信号和 发育，这是许多真核细胞生存和正常功能的关键过程 纸巾。在人类中，纤毛运动和组装的缺陷是导致许多先天性 疾病，如原发纤毛运动障碍，慢性呼吸系统疾病，生育能力受损，大脑发育 缺陷、先天性心脏病和左右身体轴的随机化。纤毛运动是由 数千个动力蛋白分子的协调活动，由多种异构体组成。我们之前的研究 野生型和突变型纤毛，以及主动击打纤毛，打开了进入功能组织的新窗口 活动的纤毛。然而，监管信号如何变化仍是一个长期存在的根本性问题 动力蛋白在分子水平上的活性，不同的调节复合体在纤毛过程中的作用是什么 运动性，以及动力蛋白是如何在空间和时间上协调以产生典型的 纤毛。基于已公布和初步的新数据这一强有力的前提，该提案直接解决了 这些通过三个具体目标的关键差距旨在(目标1)揭示 动力蛋白的作用被调节为启动和传播睫状波，(目标2)决定动力蛋白的模式 产生不同纤毛波形的活动，以及(目标3)表征组装的纤毛组件 仅在特定的二联体上询问其固有的不对称分布是否有助于产生纤毛跳动。 我们使用包括冷冻电子断层扫描在内的现代方法的强大和创新组合来 用分子分辨率、遗传学和蛋白质组学成像突变纤毛和标记蛋白，另一种模式 研究纤毛的有机体，以及比以前更深入地观察细胞内部的最先进的“切割”技术 有可能。我们期待我们的联合研究将提供重要的新的概念和机制的见解。 转化为睫毛的运动和调节，这也会影响我们对睫状体疾病的认识。

PROJECT SUMMARY The overall goal of this research is to increase our understanding of how cilia beat by dissecting the molecular mechanisms and regulation of ciliary motility on a molecular level. Cilia and flagella are conserved and ubiquitous microtubule-based organelles with important roles in cell locomotion, fluid transport, sensation, cell signaling and development, which are critical processes for the survival and proper function of many eukaryotic cells and tissues. In humans, defects in the motility and assembly of cilia are responsible for numerous congenital diseases, such as primary ciliary dyskinesia, chronic respiratory disease, impaired fertility, brain developmental defects, congenital heart disease and randomization of the left-right body axis. Cilia motility is driven by the coordinated activities of thousands of dynein molecules, comprised of multiple isoforms. Our previous studies of wild type and mutant cilia, and actively beating cilia have opened a new window into the functional organization of motile cilia. However, long-standing fundamental questions remain about how regulatory signals change dynein’s activity on a molecular level, what are the roles of the different regulatory complexes during ciliary motility, and how dyneins are spatially and temporally coordinated to generate the oscillatory beating typical for cilia. Building on a strong premise of both published and preliminary new data, this proposal directly addresses these critical gaps through three specific aims that are directed at (Aim 1) revealing mechanisms by which dynein’s action is regulated to initiate and propagate ciliary waves, (Aim 2) determine the patterns of dynein activity that generate different ciliary waveforms, and (Aim 3) characterizing ciliary components that assemble only on specific doublets to ask if their inherently asymmetric distribution contributes to producing ciliary beating. We use a powerful and innovative combination of modern approaches that include cryo-electron tomography to image mutant cilia and tagged proteins with molecular resolution, genetics and proteomics, an alternate model organism to study cilia, and a state-of-the-art “cutting” technique to look “deeper inside” cells than previously possible. We expect that our combined studies will provide important new conceptual and mechanistic insights into ciliary motility and regulation, which will also impact our understanding of ciliary diseases.