Molecular dissection of the ciliary gate

睫状门的分子解剖

• 批准号: 8690500
• 负责人: Jinghua Hu
• 金额: $ 23.85万
• 依托单位: MAYO CLINIC ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-01 至 2018-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8690500
• 关键词: Abbreviations; Affect; Animal Model; Animals; Autosomal Dominant Polycystic Kidney; Autosomal Recessive Polycystic Kidney; Bardet-Biedl Syndrome; Binding; Biological Assay; Biology; Caenorhabditis elegans; Cell surface; Centrioles; Chickens; Cilia; Data; Defect; Development; Devices; Disease; Dissection; Distal; Dyes; Embryonic Development; Environment; Etiology; Eukaryotic Cell; Excision; Exhibits; Eye; Fiber; Functional disorder; Future; Genes; Genetic Models; Hereditary Disease; Homeostasis; Homologous Gene; Human; Human Genetics; Human Pathology; Joubert syndrome; Kidney; Knock-out; Letters; Life; Light; Limb structure; Link; Liver; Location; Maintenance; Membrane; Modeling; Molecular; Mothers; Mutagenesis; Mutate; Names; Nephronophthisis; Neuraxis; Nomenclature; Nuclear Pore; Organ; Organelles; Organism; Pathology; Pathway interactions; Pattern; Personal Satisfaction; Physiological; Play; Polycystic Kidney Diseases; Pore Proteins; Process; Proteins; Research; Role; Seminal; Sensory; Sensory Receptors; Signal Pathway; Signal Transduction; Structure; Study models; Surface; Syndrome; System; TNFRSF6 gene; Testing; appendage; base; ciliopathy; cilium biogenesis; clinically relevant; design; gene function; human disease; insight; kinetosome; neuronal cell body; novel; particle; polypeptide; positional cloning; public health relevance; screening

DESCRIPTION (provided by applicant): Cilia serve as sensory devices on most eukaryotic cells surface and play an essential role in the proper formation of a diversity of organs 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), Joubert syndrome (JBST), Bardet-Biedl syndrome (BBS), nephronophthisis (NPHP), Meckel-Gruber syndrome (MKS), and autosomal recessive polycystic kidney disease (ARPKD), have been characterized molecularly as cilia-related diseases, now known collectively as ciliopathies. The establishment and maintenance of ciliary function are clearly essential for the well-being of an organism. 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. Despite the physiological and clinical relevance of cilia, the core machinery that regulates cilia biogenesis and function as well as the connection between the disease gene function and pathology remain largely elusive. One central question in cilia biology is that how the ciliary gat functionally separates the cilium from the cell body and makes it a discrete sensing organelle. Cilia only form atop mother centrioles (or basal bodies). 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, no molecular information is available regarding the composition as well as the function of TFs. In a forward mutagenesis screening that aimed to identify the determinants of ciliogenesis in C. elegans, we isolated and cloned a novel gene dyf-19, which is the sole homolog of poorly characterized human fbf1 gene. Our preliminary data showed that worm DYF-19 and human FBF1 exhibit specific localization pattern on transition fibers and distal appendages, suggesting a highly conserved TF-related function for DYF- 19/FBF1. Further analyses suggested that DYF-19 regulates the ciliary entry of assembled IFT particles on transition fibers as well as the ciliary entry of several ciliary sensory receptors. Our preliminary studies reveal the first bona fde component of TFs and demonstrate the essential roles of the TFs in cilia formation and function. Additionally, we identified two more genuine TF components, TALPID-3 and HYLS-1. Our preliminary data indicate that DYF-19, TALPID-3, and HYLS-1 functionally interact in the context of TFs. Most interestingly, talpid-3 knockout chicken is confirmed to be a ciliopathy model and human hyls1 gene is one causal locus for the ciliopathy Hydrolethalus syndrome. Due to the essential roles of cilia in mammalian early embryonic development, the study of the connections between cilia and disease are extremely difficult in humans and other mammalian model organisms. Thus, alternative experimental systems are necessary. C. elegans enables the exploration of these questions in living animals. The highly conserved ciliogenic proteins, ciliogenesis pathway, and cilia sensory function make Caenorhabditis elegans a powerful model for characterizing the physiological roles of ciliary genes in their native cellular environments. Our data support the central hypothesis of this proposal that DYF-19 acts as a functional component to define TFs as a "ciliary gate" that governs access of nascent proteins into the cilia. and that disruption of this "gate" compromises cilia formation and function. The proposed studies have great potential to unveil breakthroughs in cilia research in the near future, and would provide seminal information about how cilia biogenesis and sensory function are regulated in their native environment, shed light on the etiologies of ciliopathies.

描述（由申请人提供）：纤毛作为大多数真核细胞表面的感觉装置，在发育过程中多种器官的正确形成中发挥重要作用。纤毛通过鞭毛内运输（IFT）和感觉转导能力在所有纤毛生物中都是高度保守的。近十年来，随着人类疾病基因定位克隆技术的快速发展，许多疾病，如常染色体显性多囊肾病（ADPKD）、Joubert综合征（JBST）、Bardet-Biedl综合征（BBS）、肾病（NPHP）、Meckel-Gruber综合征（MKS）和常染色体隐性多囊肾病（ARPKD），在分子上被定性为与纤毛相关的疾病，现在统称为纤毛病。纤毛功能的建立和维持显然对生物体的健康至关重要。与普遍存在的纤毛一致，许多纤毛病以综合征性疾病的形式出现，影响多个器官，包括肾、肝、肢体、眼睛和中枢神经系统。尽管纤毛具有生理和临床意义，但调控纤毛生物发生和功能的核心机制以及疾病基因功能与病理之间的联系在很大程度上仍是难以捉摸的。纤毛生物学的一个核心问题是纤毛门如何将纤毛从细胞体中分离出来，并使其成为一个独立的传感细胞器。纤毛仅在母体中心粒（或基生体）上形成。在纤毛发生过程中，母中心粒的远端附属物转变为过渡纤维（tf），形成一个连接基体和纤毛基膜的9叶螺旋桨结构。其独特的亚细胞位置使其成为纤毛门的一个很好的候选者。然而，没有关于tf的组成和功能的分子信息。在一项旨在确定秀丽隐杆线虫纤毛发生决定因素的前向诱变筛选中，我们分离并克隆了一个新的基因dyf-19，这是人类fbf1基因的唯一同源基因。我们的初步数据显示，蠕虫DYF-19和人类FBF1在过渡纤维和远端附属物上表现出特定的定位模式，表明DYF-19 /FBF1具有高度保守的tf相关功能。进一步分析表明，DYF-19调节了过渡纤维上组装的IFT颗粒的纤毛进入，以及几种纤毛感觉受体的纤毛进入。我们的初步研究揭示了TFs的第一个真正成分，并证明了TFs在纤毛形成和功能中的重要作用。此外，我们还鉴定了另外两种正品TF成分，TALPID-3和HYLS-1。我们的初步数据表明，DYF-19、TALPID-3和HYLS-1在tf的背景下具有功能相互作用。最有趣的是，talid -3基因敲除鸡被证实是纤毛病的模型，而人类hyls1基因是纤毛病lethalus综合征的一个致病位点。由于纤毛在哺乳动物早期胚胎发育中的重要作用，在人类和其他哺乳动物模式生物中研究纤毛与疾病之间的关系是非常困难的。因此，替代实验系统是必要的。秀丽隐杆线虫使得在活体动物中探索这些问题成为可能。高度保守的纤毛蛋白、纤毛发生途径和纤毛感觉功能使秀丽隐杆线虫成为表征纤毛基因在其原生细胞环境中生理作用的有力模型。我们的数据支持该提案的中心假设，即DYF-19作为一个功能性成分，将tf定义为控制新生蛋白进入纤毛的“纤毛门”。这个“门”的破坏会损害纤毛的形成和功能。这些研究有可能在不久的将来揭示纤毛研究的突破，并为纤毛在自然环境下的生物发生和感觉功能的调节提供开创性的信息，揭示纤毛病的病因。