Dystroglycan in Epithelial and Neural Development

上皮和神经发育中的肌营养不良聚糖

• 批准号: 7666018
• 负责人: JAMES M KRAMER
• 金额: $ 26.43万
• 依托单位: NORTHWESTERN UNIVERSITY AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-09-01 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7666018
• 关键词: Ablation; Abnormal Cell; Alleles; Animals; Basement membrane; Binding; Binding Proteins; Binding Sites; Biological; Biological Process; Brain; Caenorhabditis elegans; Cell Adhesion; Cell Surface Proteins; Cell Surface Receptors; Cells; Complex; Cytoplasmic Tail; Defect; Development; Dystroglycan; Dystrophin; Elements; Enhancers; Epithelial; Epithelial Cells; Epithelium; Extracellular Protein; Genetic; Genetic Screening; Goals; Homologous Gene; Human Pathology; In Vitro; Laminin; Lesion; Ligands; Link; Maintenance; Mediating; Muscle; Muscle Cells; Muscular Dystrophies; Mutagenesis; Mutation; Neurons; Organism; Orthologous Gene; Pathway interactions; Phenotype; Physiological; Process; Proteins; RNA Interference; Reagent; Receptor Cell; Research Design; Research Personnel; Role; Signal Pathway; Signal Transduction; Structure; Synaptic Transmission; Technology; Tertiary Protein Structure; Tissues; Transgenic Animals; Transgenic Organisms; Variant; Yeasts; axon guidance; cell type; extracellular; gain of function; genetic analysis; in vivo; insight; laminin-1; loss of function; migration; mutant; neurodevelopment; novel; programs; receptor; relating to nervous system; tool; yeast two hybrid system

DESCRIPTION (provided by applicant): Dystroglycan (DG) is a conserved cell surface receptor that serves to link extracellular basement membrane and intracellular cytoskeletal and signaling components. DG has been extensively studied in muscle cells where it links laminin to dystrophin, as part of the complex that is disrupted in muscular dystrophies. Dystrophies associated with brain abnormalities, such as Fukuyama muscular dystrophy, result from defects of DG processing and reveal its importance in neural and epithelial cells. The specific functions of DG in non-muscle cells, however, are poorly understood. We have shown that the C. elegans DG ortholog, DGN-1, has critical functions in many neural/epithelial cells, but is not required in muscle. DGN-1 mutant defects include abnormal cell adhesion and migration, aberrant axon guidance and defective synaptic transmission. Thus, in vivo studies using the powerful genetic tools available in C. elegans may clarify how DG functions in neurons and epithelia. DG functions are highly cell type dependent, and we hypothesize that specific extracellular and intracellular factors are critical in determining the physiological roles of DG in different contexts. Using transgenic analysis of altered DGN-1 constructs expressed in DGN-1 null animals, we will determine which domains of DGN-1 are involved in the particular neural/epithelial functions we have described. The importance of particular extracellular ligands will be analyzed by transgenic expression of dominantly interfering ligand domains and/or complete replacement of ligand molecules with specifically altered versions. Intracellular ligands will be identified using yeast two-hybrid technology and RNAi knockdown of candidates. Genetic screens will be pursued to generate novel DGN-1 mutations that will define important aspects of DG structure, and extragenic enhancers or suppressors of DGN-1 function, that can identify interacting molecules. Also, we have shown that transgenic expression of altered DGN-1 can dominantly induce neural/epithelial phenotypes, and genetic screens to suppress these effects will be used to identify upstream and/or downstream effectors of DGN-1 function. This combination of targeted and random genetic approaches will provide in vivo evidence for particular extracellular and intracellular ligand interactions underlying specific DG functions in specific cells.

描述（由申请人提供）：肌营养不良聚糖（DG）是一种保守的细胞表面受体，用于连接细胞外基底膜和细胞内细胞骨架和信号传导组分。DG已在肌细胞中被广泛研究，其中它将层粘连蛋白连接到肌营养不良蛋白，作为在肌营养不良症中被破坏的复合物的一部分。与脑异常相关的营养不良，如福山肌营养不良，是由DG加工缺陷引起的，并揭示了其在神经和上皮细胞中的重要性。然而，DG在非肌肉细胞中的具体功能知之甚少。我们证明了C.秀丽线虫DG直系同源物，DGN-1，在许多神经/上皮细胞中具有关键功能，但在肌肉中不需要。DGN-1突变缺陷包括异常的细胞粘附和迁移、异常的轴突导向和有缺陷的突触传递。因此，在体内研究中使用强大的遗传工具，可在C。elegans可能阐明DG在神经元和上皮中的功能。DG的功能是高度依赖于细胞类型，我们假设，特定的细胞外和细胞内的因素是至关重要的，在确定DG的生理作用，在不同的情况下。使用在DGN-1缺失动物中表达的改变的DGN-1构建体的转基因分析，我们将确定DGN-1的哪些结构域参与我们所描述的特定神经/上皮功能。将通过显性干扰配体结构域的转基因表达和/或用特异性改变的形式完全替换配体分子来分析特定细胞外配体的重要性。将使用酵母双杂交技术和候选物的RNAi敲低来鉴定细胞内配体。将进行遗传筛选，以产生新的DGN-1突变，这些突变将定义DG结构的重要方面，以及DGN-1功能的基因外增强子或抑制子，这些突变可以识别相互作用的分子。此外，我们已经表明，改变的DGN-1的转基因表达可以显性诱导神经/上皮表型，和基因筛选，以抑制这些影响将被用于确定上游和/或下游效应的DGN-1功能。这种靶向和随机遗传方法的组合将为特定细胞中特定DG功能的特定细胞外和细胞内配体相互作用提供体内证据。