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: 8044806
• 负责人: Jan Lammerding
• 金额: $ 2.11万
• 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 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8044806
• 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; Tissues; Work; base; design; env Gene Products; improved; insight; lamin C; 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(申请人提供)：核膜蛋白lamins A和C的突变导致Emery-Dreifuss肌营养不良症、跛带型肌营养不良症和一系列其他统称为椎板病的疾病。潜在的疾病机制尚不清楚，但最近的证据表明，核-细胞骨架连接中断可能导致肌肉表型。层蛋白A和C是正确定位LINC复合体核膜所必需的，该复合体将细胞核连接到细胞骨架，但特定的层蛋白A/C突变的影响尚不清楚。我的长期目标是了解Lamin A/C突变是否可以通过LINC复合体干扰核-细胞骨架的耦合，导致异常的细胞内力传递和机械转导，这可能有助于肌肉板层病的组织特异性表型，类似于Duchenne肌营养不良的病因学。我的具体目标是： 1.验证LINC复合体是力从细胞骨架传递到细胞核所必需的假设。我将测量在应用细胞骨架应变后，选择性地破坏具有显性负突变的LINC复合体或通过RNA干扰使基因沉默后的核变形。 2.验证特定的层粘连蛋白A/C突变可将复合体定位于核膜的假设。我将分析来自椎板病患者、肌肉椎板病小鼠模型和健康/野生型对照细胞中LINC复合体成分的细胞内分布和扩散移动性。 3.为了验证Lamin A/C突变可以改变通过LINC复合体的力传递的假设，我设计了一种新的磁珠附着实验来定量通过LINC复合体传递的最大力，我将把这项技术应用于携带特定lamin A/C突变的细胞。 4.验证破坏LINC复合体改变机械转导的假说。我将在LINC复合体功能被选择性破坏的细胞中测量应变诱导的基因表达。 研究Lamin A/C突变对核-细胞骨架偶联的影响将有助于我们更好地理解核膜的正常和组织特异性功能，并可能导致对诸如Emery-Dreifuss肌营养不良症等肌肉板层疾病的分子机制的新见解，为这些疾病的治疗提供新的靶点。