Identification of genes responsible for sarcolemmal integrity in C. elegans

描述（由申请人提供）：后生动物物种中的许多类型的细胞在机械应力条件下运行，并进化为科普质膜损伤。如在几种肌营养不良症中所观察到的，质膜损伤的预防或修复失败可导致疾病，或可影响疾病进展。膜修复的更详细的机制的理解，需要找到各种疾病的治疗干预膜损伤的病理生理基础。详细了解修复机制的第一步是全面鉴定组分分子。我们开发了一种方法，使用显微线虫C。用于鉴定介导受损肌膜（肌肉质膜）修复的分子和细胞组分。利用C. elegans体，允许在体内检测荧光蛋白，我们开发了一种新的，简单的测定，可以很容易地评估肌膜损伤的程度。通过该试验，我们发现C.秀丽线虫肌营养不良蛋白突变体（杜氏肌营养不良的模型）表现出肌膜渗漏和损伤，尽管很弱。基于这一发现，我们进行了遗传筛选，以分离突变体，有缺陷的肌膜修复，因此，加剧肌营养不良蛋白突变体的肌膜损伤。在这个探索性的建议，我们试图建立屏幕作为一个有价值的工具，确定基因负责肌膜修复。有了几个突出的突变体在手，我们特别建议克隆致病基因的缺陷，在肌膜修复的基因定位和全基因组测序相结合。一旦我们确定了负责基因，我们将使用成熟的C。elegans基因技术与此同时，我们将确定这些已鉴定基因之间的关系，以了解它们如何共同发挥作用以保护和修复肌膜。该项目的成功完成将导致更好地了解肌膜修复的分子机制，更重要的是，将揭示潜在的药物靶点，用于阻断受膜修复影响的肌营养不良症的进展。 公共卫生相关性：许多形式的肌营养不良症，包括杜氏肌营养不良症，导致肌肉膜完整性的破坏。因此，了解肌肉膜的完整性是如何维持和修复具有治疗意义。建议的C. elegans基因研究将确定和表征负责肌肉膜修复的基因。

DESCRIPTION (provided by applicant): Many types of cells in metazoan species operate under conditions of mechanical stress, and are evolved to cope with plasma membrane damage. A failure in either the prevention or repair of plasma membrane damage can cause disease, or can influence disease progression, as observed in several muscular dystrophies. A more detailed mechanistic understanding of membrane repair is required to find therapeutic interventions for a variety of diseases where membrane damage underlies the pathophysiology. The first step towards the detailed understanding of the repair mechanism is a comprehensive identification of the component molecules. We developed a method that uses the microscopic nematode C. elegans for identifying molecular and cellular components that mediate the repair of damaged sarcolemma (muscle plasma membrane). By taking advantage of the transparency of the C. elegans body that allows detection of fluorescence proteins in vivo, we developed a novel, simple assay that can easily evaluate the degree of sarcolemmal damage. With this assay, we found that C. elegans dystrophin mutants (a model of Duchenne muscular dystrophy) exhibit sarcolemmal leakage and damage, albeit weak. Based on this finding, we performed a genetic screen to isolate mutants that have defects in sarcolemmal repair and, as a result, aggravate sarcolemmal damage of dystrophin mutants. In this exploratory proposal, we seek to establish the screen as a valuable tool for identifying genes responsible for sarcolemmal repair. With several prominent mutants in hands, we specifically propose to clone causal genes for defects in sarcolemmal repair by a combination of genetic mapping and whole genome sequencing. Once we identify the responsible genes, we will characterize the cloned genes using well-established C. elegans genetic techniques. In parallel, we will determine the relationship between these identified genes to understand how they function together to protect and repair the sarcolemma. The successful completion of this project will lead to a better understanding of the molecular mechanism of sarcolemmal repair and, more importantly, will reveal potential druggable targets for blocking the progression of muscular dystrophies that are influenced by membrane repair. PUBLIC HEALTH RELEVANCE: Many forms of muscular dystrophy, including Duchenne muscular dystrophy, cause a disruption of the integrity of the muscle membrane. Hence, understanding how muscle membrane integrity is maintained and repaired has a therapeutic implication. The proposed C. elegans genetic study will identify and characterize genes responsible for muscle membrane repair.