Cilia Assembly and Transport in the Vertebrate Retina

脊椎动物视网膜中纤毛的组装和运输

• 批准号: 8187542
• 负责人: Brian D Perkins
• 金额: $ 27.43万
• 依托单位: TEXAS A&M UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-08-01 至 2012-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8187542
• 关键词: Address; Adult; Affect; Alleles; Anatomy; Animal Model; Apical; Architecture; Bardet-Biedl Syndrome; Behavioral; Binding; Biological Models; Blindness; Body Regions; Candidate Disease Gene; Cell Polarity; Cell Survival; Cells; Cilia; Clinical; Complex; Defect; Development; Disease; Docking; Dominant-Negative Mutation; Dsh protein; Dynein ATPase; Electron Microscopy; Electrons; Embryo; Functional disorder; Gene Components; Gene Expression; Gene Mutation; Genes; Genetic; Goals; Golgi Apparatus; Human; Individual; Joubert syndrome; Kidney Diseases; LIS1 protein; Lead; Link; Maintenance; Mental Retardation; Microscopic; Microtubules; Modeling; Molecular; Monomeric GTP-Binding Proteins; Motor; Movement; Mutation; Organelles; Pathway interactions; Pattern; Phenotype; Photoreceptors; Play; Polydactyly; Population; Positioning Attribute; Process; Proteins; Reagent; Regulation; Retina; Retinal; Retinal Degeneration; Retinal Diseases; Retinal Dystrophy; Role; Side; Signal Pathway; Signal Transduction; Site; Situs Inversus; Stereotyping; Structure; Surface; System; TNFRSF5 gene; Techniques; Testing; Tissues; Transgenic Organisms; Travel; Vertebrate Photoreceptors; Vertebrates; Vision; Zebrafish; base; body position; cell motility; ciliopathy; cilium biogenesis; dynactin; hereditary blindness; kinetosome; light microscopy; loss of function; mutant; novel; null mutation; particle; photoreceptor degeneration; prevent; protein transport; retinal damage; therapy development; tool; trafficking

DESCRIPTION (provided by applicant): The long-term goal of this project is to understand the molecular basis of cilia formation and maintenance in vertebrate photoreceptor cells and how mutations in cilia genes cause retinal degeneration. In vertebrates, the assembly and maintenance of photoreceptor outer segments begins with the formation of a connecting cilium. The connecting cilium contains a microtubule-based axoneme that is anchored to the apical inner segment by a basal body. Cilia formation begins with the docking of basal bodies at the apical surface of the inner segment. Extension and maintenance of the ciliary axoneme requires the bi-directional motility process of Intraflagellar Transport (IFT) to carry proteins from the photoreceptor inner segment and outer segment. Genetic mutations disrupting the assembly, structure, or function of basal bodies and/or cilia result in a spectrum of diseases known as ciliopathies. These multisyndromic disorders often present with retinal degeneration, kidney disease, mental retardation, and polydactyly. In the current application, we will utilize loss-of-function strategies in zebrafish to investigate the mechanisms controlling basal body localization and determine how mutations in cep290 and arl13b, which are causative for Joubert Syndrome and other ciliopathies, lead to retinal degeneration. In Specific Aim 1, we will examine zebrafish carrying null mutations in cep290 and arl13b for retinal phenotypes. We will also test cep290 and arl13b for functional interactions with Bardet-Biedl Syndrome (BBS) genes, IFT genes, and components of the Planar Cell Polarity (PCP) pathway. These interactions will identify potential second-site modifiers that enhance expression of photoreceptor phenotypes. In Specific Aim 2, we will test the hypothesis that the dynactin complex regulates cytoplasmic dynein motors during basal body docking and ciliogenesis by examining zebrafish mutants in the p150 and p50 subunits of dynactin. In Specific Aim 3, we will directly test the hypothesis that the PCP pathway functions in photoreceptors to control the polarized positioning of basal bodies and that defects in PCP signaling can contribute to photoreceptor degeneration. Our preliminary evidence indicates that basal bodies indeed show a highly polarized arrangement within the adult zebrafish retina. We will determine if this patterning exists during development. We will then express dominant-negative forms the core PCP protein Disheveled (Dvl) in photoreceptors and determine if this leads to retinal degeneration. The results of these studies will allow us to study genes that play critical roles during both cilia assembly and maintenance and will identify novel gene candidates for hereditary blindness so that therapies can be developed to prevent vision loss. PUBLIC HEALTH RELEVANCE: In the vertebrate retina, photoreceptor survival depends on the proper formation and maintenance of the connecting cilium and outer segment. The cilium is a complex organelle that is of great clinical importance because dysfunction in cilia assembly or function can lead to retinal degeneration, kidney disorders, mental retardation, situs inversus, polydactyly, and other conditions. An understanding of the mechanisms that control the formation, positioning, and structural integrity of cilia will lead to the development of treatments for ciliary diseases.

描述(申请人提供)：该项目的长期目标是了解脊椎动物感光细胞纤毛形成和维持的分子基础，以及纤毛基因突变如何导致视网膜退化。在脊椎动物中，光感受器外节的组装和维持始于连接纤毛的形成。连接纤毛含有微管为基础的轴丝，该轴丝由一个基准体固定在顶端内段。纤毛的形成始于基体在内节顶面的对接。纤毛轴丝的伸展和维持需要鞭毛内运输(IFT)的双向运动过程，以携带来自感光细胞内段和外段的蛋白质。基因突变破坏了基本体和/或纤毛的组装、结构或功能，导致了一系列称为纤毛病的疾病。这些多综合征疾病通常表现为视网膜变性、肾脏疾病、智力低下和多指畸形。在目前的应用中，我们将利用斑马鱼的功能丧失策略来研究控制基底体定位的机制，并确定导致Joubert综合征和其他纤毛疾病的CEP290和Arl13b突变如何导致视网膜退化。在特定的目标1中，我们将检查携带CEP290和Arl13b零突变的斑马鱼的视网膜表型。我们还将测试CEP290和Arl13b与Bardet-Biedl综合征(BBS)基因、IFT基因和平面细胞极性(PCP)途径组件的功能相互作用。这些相互作用将确定潜在的第二位点修饰，以增强光感受器表型的表达。在特定的目标2中，我们将通过研究斑马鱼dynactin的p150和p50亚基突变来验证这一假设，即dynactin复合体在基体对接和纤毛发生过程中调节细胞质dynein马达。在特定的目标3中，我们将直接测试PCP通路在光感受器中起作用以控制基底体的极化位置以及PCP信号缺陷可能导致光感受器退化的假设。我们的初步证据表明，在成年斑马鱼的视网膜中，基底确实显示出高度极化的排列。我们将确定在开发过程中是否存在这种模式。然后，我们将在光感受器中表达显性-阴性形式的核心PCP蛋白脱发(DVL)，并确定这是否会导致视网膜退化。这些研究的结果将使我们能够研究在纤毛组装和维持过程中发挥关键作用的基因，并将识别遗传性失明的新候选基因，以便开发预防视力丧失的治疗方法。 公共卫生相关性：在脊椎动物的视网膜中，光感受器的生存依赖于连接纤毛和外段的适当形成和维持。纤毛是一个复杂的细胞器，具有重要的临床意义，因为纤毛组装或功能障碍可导致视网膜变性、肾脏疾病、智力低下、内翻、多指和其他情况。对纤毛的形成、定位和结构完整性的控制机制的了解将有助于睫状体疾病治疗的发展。