Nuclear-cytoskeletal coupling in muscular laminopathies

肌肉核纤层蛋白病中的核细胞骨架耦合

• 批准号: 7582243
• 负责人: Jan Lammerding
• 金额: $ 25.88万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 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 2012-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7582243
• 关键词: Adhesions; Affect; Biochemical; Biological Assay; Biomedical Engineering; Biopsy; Cell Nucleus; Cell physiology; Cells; Complex; Couples; Coupling; Cytoskeletal Modeling; Cytoskeleton; Data; Defect; Disease; Dominant-Negative Mutation; Duchenne muscular dystrophy; Dystrophin; Emery-Dreifuss Muscular Dystrophy; Etiology; Fibroblasts; Fracture; Gene Expression; Gene Silencing; Goals; Lamin Type A; Lamins; Lead; Limb-Girdle Muscular Dystrophies; Mammalian Cell; Measures; Mechanical Stimulation; Mechanics; Membrane Proteins; Methods; Modeling; Molecular; Molecular Biology Techniques; Mus; Muscle; Muscular Dystrophies; Mutation; Normal Cell; Normal tissue morphology; Nuclear; Nuclear Envelope; Patients; Phenotype; Positioning Attribute; Protein Family; Protein Isoforms; Proteins; RNA Interference; Research Personnel; Role; Signal Transduction; Striated Muscles; Techniques; Testing; The Sun; Tissues; Work; base; design; env Gene Products; improved; insight; magnetic beads; mouse model; mutant; novel; programs; response; tool; transmission process

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.

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 肌营养不良症、肢带肌营养不良症以及统称为核纤层蛋白病的广泛其他疾病。潜在的疾病机制尚不清楚，但最近的证据表明，核-细胞骨架耦合破坏可能导致肌肉表型。核纤层蛋白 A 和 C 是 LINC 复合物正确核膜定位所必需的，该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-细胞骨架耦合，从而导致异常的细胞内力传递和机械转导，从而可能导致肌肉核纤层蛋白病中的组织特异性表型，类似于疾病的病因学杜氏肌营养不良症。我的具体目标是： 1. 检验以下假设：从细胞骨架到细胞核的力传递需要 LINC 复合体。在使用显性失活突变体选择性破坏 LINC 复合物或通过 RNA 干扰进行基因沉默后，我将测量细胞骨架应变对细胞骨架应变的响应。 2. 检验特定核纤层蛋白 A/C 突变可以将 LINC 复合物定位到核膜的假设。我将分析来自核纤层蛋白病患者、肌肉核纤层蛋白病小鼠模型和健康/野生型对照细胞中 LINC 复合物成分的细胞内分布和扩散迁移率。 3. 为了检验核纤层蛋白 A/C 突变可以改变通过 LINC 复合体的力传递的假设，设计了一种新型磁珠粘附测定法来量化可以通过 LINC 复合体传递的最大力，我将将此技术应用于细胞携带特定核纤层蛋白 A/C 突变。 4. 检验 LINC 复合体的破坏会改变机械转导的假设。我将测量 LINC 复合体功能被选择性破坏的细胞中应变诱导的基因表达。 研究核纤层蛋白 A/C 突变对核-细胞骨架耦合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，并有助于对导致 Emery-Dreifuss 等肌肉核纤层蛋白病的分子机制有新的认识。肌营养不良症，可能为这些疾病的治疗提供新的靶点。