Dystroglycan in Epithelial and Neural Development

描述(申请人提供)：营养不良多糖(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成分。DG在肌肉细胞中得到了广泛的研究，在那里它将层粘连蛋白与肌营养不良蛋白联系起来，作为肌肉营养不良症中被破坏的复合体的一部分。与脑异常相关的营养不良，如福山肌营养不良，是由于DG加工缺陷引起的，并揭示了DG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却知之甚少。我们已经证明，线虫DG同源基因DGN-1在许多神经/上皮细胞中具有关键功能，但在肌肉中不是必需的。DGN-1突变缺陷包括异常的细胞黏附和迁移、异常的轴突引导和突触传递缺陷。因此，使用线虫强大的遗传工具进行的活体研究可能会阐明DG在神经元和上皮中的功能。DG的功能高度依赖于细胞类型，我们假设特定的细胞外和细胞内因子在不同的环境中决定DG的生理作用是至关重要的。通过对在DGN-1缺失动物中表达的改变的DGN-1结构进行转基因分析，我们将确定DGN-1的哪些结构域参与了我们所描述的特定的神经/上皮功能。特定细胞外配体的重要性将通过转基因表达主要干扰的配体结构域和/或用特定改变的版本完全取代配体分子来分析。将使用酵母双杂交技术和RNAi敲除候选基因来鉴定细胞内配体。基因筛选将被用来产生新的DGN-1突变，它将定义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，可以识别相互作用的分子。此外，我们已经证明，转基因表达改变的DGN-1可以主要诱导神经/上皮表型，抑制这些影响的遗传筛选将被用于识别DGN-1功能的上游和/或下游效应。这种靶向和随机遗传方法的结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。

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.