Nuclear-cytoskeletal coupling in muscular laminopathies

描述（由申请人提供）：核膜蛋白核纤层蛋白A和C的突变导致Emery-Dreifuss型肌营养不良、肢带型肌营养不良和统称为核纤层蛋白病的广谱其他疾病。潜在的疾病机制尚不清楚，但最近的证据表明，破坏核细胞骨架耦合可能有助于肌肉表型。核纤层蛋白A和C是LINC复合物正确核膜定位所必需的，LINC复合物将细胞核连接到细胞骨架，但特定核纤层蛋白A/C突变的影响仍不清楚。我的长期目标是了解核纤层蛋白A/C突变是否可以通过LINC复合物干扰核-细胞骨架偶联，导致异常的细胞内力传递和机械转导，这可能有助于肌层蛋白病的组织特异性表型，类似于Duchenne肌营养不良症的疾病病因。我的具体目标是： 1.为了验证LINC复合物是从细胞骨架到细胞核的力传递所必需的假设。我将测量在用显性负突变体选择性破坏LINC复合物或通过RNA干扰进行基因沉默后，细胞骨架应变对核变形的反应。 2.为了验证特定核纤层蛋白A/C突变可以将LINC复合物定位到核膜的假设。我将分析细胞内的分布和扩散流动性的LINC复合物成分的细胞从laminopathy患者，小鼠模型的肌肉laminopathies，和健康/野生型对照。 3.为了检验核纤层蛋白A/C突变可以改变通过LINC复合物的力传递的假设，设计了一种新的磁珠粘附试验来量化可以通过LINC复合物传递的最大力，我将把这种技术应用于携带特定核纤层蛋白A/C突变的细胞。 4.为了检验LINC复合物的破坏改变机械转导的假设。我将在LINC复合体功能被选择性破坏的细胞中测量菌株诱导的基因表达。 研究核纤层蛋白A/C突变对核-细胞骨架偶联的影响将有助于提高我们对核膜正常和组织特异性功能的理解，并可能导致对肌层蛋白病（如Emery-Dreifuss肌营养不良症）的分子机制的新见解，可能为治疗这些疾病提供新的靶点。

DESCRIPTION (provided by applicant): Mutations in the nuclear envelope proteins lamins A and C cause Emery-Dreifuss muscular dystrophy, limbgirdle muscular dystrophy, and a broad spectrum of other diseases collectively called laminopathies. The underlying disease mechanism is unclear, but recent evidence suggests that disrupted nuclear-cytoskeletal coupling could contribute to the muscular phenotypes. Lamins A and C are required for the correct nuclear envelope localization of the LINC complex, which connects the nucleus to the cytoskeleton, but the effects of specific lamin A/C mutations remain unclear. My long term goal is to understand if lamin A/C mutations can disturb nuclear-cytoskeletal coupling through the LINC complex, resulting in abnormal intracellular force transmission and mechanotransduction that could contribute to the tissue specific phenotypes in muscular laminopathies, similar to the disease etiology of Duchenne muscular dystrophy. My specific aims are: 1. To test the hypothesis that the LINC complex is required for force transmission from the cytoskeleton to the nucleus. I will measure nuclear deformation in response to applied cytoskeletal strain after selective disruption of the LINC complex with dominant negative mutants or gene silencing by RNA interference. 2. To test the hypothesis that specific lamin A/C mutations can LINC complex localization to the nuclear envelope. I will analyze intracellular distribution and diffusional mobility of LINC complex components in cells from laminopathy patients, mouse models of muscular laminopathies, and healthy/wild-type controls. 3. To test the hypothesis that lamin A/C mutations can alter force transmission through the LINC complex, have designed a novel magnetic bead adhesion assay to quantify the maximal force that can be transmitted through the LINC complex and I will apply this technique to cells carrying specific lamin A/C mutations. 4. To test the hypothesis that disruption of the LINC complex alters mechanotransduction. I will measure strain-induced gene expression in cells in which LINC complex function has been selectively disrupted. Studying the effect of lamin A/C mutations on nuclear-cytoskeletal coupling will help to improve our understanding of normal and tissue-specific functions of the nuclear envelope and can lead to new insights into the molecular mechanisms responsible for muscular laminopathies such as Emery-Dreifuss muscular dystrophy, potentially providing new targets for the treatment of these diseases.