Molecular dissection of the ciliary gate

项目摘要 纤毛是真核细胞表面的感觉器官，在细胞发育过程中起着重要作用。 通过鞭毛内转运（IFT）和感觉转导能力的纤毛组装在哺乳动物中高度保守。 所有纤毛生物。随着人类疾病基因定位克隆技术的快速发展， 十年来，各种各样的疾病，如常染色体显性多囊肾病（ADPKD）， 在分子上被描述为纤毛病与纤毛的普遍存在相一致，许多 纤毛病作为影响多个器官的综合征性疾病发生，包括肾、肝、肢体、眼和 中枢神经系统纤毛生物学中的一个中心问题是纤毛门如何在功能上分离 纤毛从细胞体中分离出来，使其成为一个独立的感觉细胞器。在纤毛发生过程中， 母亲中心粒的附属物转变为过渡纤维（TF），形成一个9叶螺旋桨 连接基体和纤毛基膜的结构。TF的不同亚细胞定位 使其成为睫状门的良好候选者。尽管如此，最大的挑战是， 关于TF作为睫状门的结构或功能的建立的见解仍然不清楚。 由于纤毛在哺乳动物胚胎发育中的重要作用， 纤毛和疾病在哺乳动物模型中是极其困难的。因此，可替代的实验系统是 必要秀丽隐杆线虫已被建立为表征 纤毛蛋白在其天然细胞环境中的生理作用， 哺乳动物生物学由于高度保守的纤毛组成和信号。我们率先应用了 梭线虫作为模型来研究TF的生物学重要性。我们的初步研究表明，DYF-19 身体上与不同的球员，以调节不同的纤毛门控：与DYF-19-TALPID-3-ANK-26 功能模块用于调节IFT输入，而DYF-19-CCDC-85模块用于调节门控 膜蛋白另一方面，HYLS 1与GAS 8协调，以规范 睫状门我们还检索了新的蠕虫突变体，可能破坏TF的完整性，在正向遗传 筛选此外，我们的初步研究表明，在C。优雅的高度 在哺乳动物细胞中是保守的在本提案中，我们将确定 潜在的途径，使睫状门在纤毛和纤毛病的背景下的基本作用是 更好地理解。我们计划通过追求三个具体问题来实现这一目标：i）纤毛门控是如何 实现？（二）睫状门是如何建立的？以及iii）如果睫状门的核心通路是 在哺乳动物细胞中是保守的结合C.哺乳动物系统，我们有信心， 提供有关睫状门的分子身份和核心保守途径的开创性信息， 并大大扩展了我们对纤毛生物学以及人类纤毛病发病机理的理解。

Project Summary Cilia serve as sensory devices on most eukaryotic cells surface and play an essential role in development. Ciliary assembly via intraflagellar transport (IFT) and sensory transduction capabilities are highly conserved in all ciliated organisms. With rapid advancements in the positional cloning of human disease genes in the past decade, a wide variety of disorders such as autosomal dominant polycystic kidney disease (ADPKD) have been characterized molecularly as ciliopathies. Consistent with the ubiquitous presence of cilia, many ciliopathies occur as syndromic disorders that affect multiple organs, including the kidney, liver, limb, eye, and central nervous system. One central question in cilia biology is that how the ciliary gate functionally separates the cilium from the cell body and makes it a discrete sensing organelle. During ciliogenesis, the distal appendages of the mother centriole transform to transition fibers (TFs), which form a 9-bladed propeller structure connecting the basal body to the ciliary base membrane. The distinct subcellular location of TFs makes it a good candidate for the ciliary gate. Nonetheless, the paramount challenges being that molecular insights about the establishment, either structural or functional, of TFs as the ciliary gate remain poorly defined. Due to the essential roles of cilia in mammalian embryonic development, the study of the connections between cilia and disease are extremely difficult in mammalian models. Thus, alternative experimental systems are necessary. Caenorhabditis elegans has been established as an effective model for characterizing the physiological roles of ciliary proteins in their native cellular environments that is relevant for understanding mammalian biology due to the highly conserved cilia composition and signaling. We pioneered the application of C. elegans as a model to study the biological importance of TFs. Our preliminary studies show that DYF-19 physically associates with different players to regulate distinct cilia gating: with the DYF-19-TALPID-3-ANK-26 functional module in regulating IFT import, whereas DYF-19-CCDC-85 module in regulating gating for membrane proteins. On the other hand, HYLS1 coordinate with GAS8 to regulate the establishment of the ciliary gate. We also retrieved novel worm mutants with likely disrupted TF integrity in a forward genetic screening. Furthermore, our initial studies suggested that the key discoveries made in C. elegans are highly conserved in mammalian cells. In this proposal, we will determine the full components and activities of underlying pathways so that the fundamental roles of the ciliary gate in the context of cilia and ciliopathies are better understood. We plan to achieve this goal by pursuing three specific questions: i), how cilia gating is achieved? ii) how the ciliary gate is established? and iii) if the core pathways for the ciliary gate are conserved in mammalian cells? By combining C. elegans with mammalian systems, we are confident to provide seminal information about the molecular identity and the core conserved pathways of the ciliary gate, and substantially extend our understanding of cilia biology as well as of the pathogenesis of human ciliopathies.