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Molecular and Cellular Pathogenesis of Emery-Dreifuss Muscular Dystrophy

Emery-Dreifuss 肌营养不良症的分子和细胞发病机制

基本信息

批准号：
8215808
负责人：
Jason Cheol Choi
金额：
$ 5.57万
依托单位：
COLUMBIA UNIVERSITY HEALTH SCIENCES
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-02-01 至 2013-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8215808
关键词：
Actins Affect Amino Acids Binding Binding Proteins Biochemical Biological Assay Blood Circulation Cardiac Cardiac Myocytes Cardiomyopathies Cell Nucleus Cells Clinical Co-Immunoprecipitations Complex Cytoskeleton Data Defect Development Digitonin Disease Emery-Dreifuss Muscular Dystrophy Energy Transfer Ensure Fluorescence Fluorescence Microscopy Fluorescence Recovery After Photobleaching Genes Goals Growth Factor Heart Hereditary Disease Human In Vitro Inflammatory Integral Membrane Protein Intermediate Filaments Knock-in Mouse Knowledge Lamin Type A Lead Life Link MAPK1 gene MAPK8 gene Maintenance Measures Mediating Membrane Proteins Methods Mitogen-Activated Protein Kinases Molecular Mutation Myocardium N-terminal Nuclear Nuclear Import Nuclear Inner Membrane Nuclear Lamina Nuclear Structure Nuclear Translocation Pathogenesis Pathway interactions Patients Photobleaching Point Mutation Protein Kinase Interaction Proteins Scapuloilioperoneal Atrophy with Cardiopathy Signal Pathway Skeletal Muscle Small Interfering RNA Stimulus Stress Surface Symptoms Techniques Testing Therapeutic Intervention Time Tissues Yeasts base biological adaptation to stress cytokine effective therapy extracellular mouse model muscular dystrophy mouse model mutant overexpression prevent reconstitution research study response treatment strategy yeast two hybrid system

项目摘要

DESCRIPTION (provided by applicant): The overall goal of this application is to define the molecular pathogenesis of cardiomyopathy associated with Emery-Dreifuss Muscular Dystrophy (EDMD). Autosomal dominant EDMD arises from mutations in a gene encoding A-type lamins, which are intermediate filaments involved in the maintenance of nuclear structure. Further, A-type lamins (along with suns and nesprins) are a component of the LINC complex, which establish a physical connection between the nucleus and the actin cytoskeleton. In EDMD mouse models of EDMD, mitogen activated protein (MAP) kinases ERK and JNK are activated in the heart tissue prior to the development of cardiomyopathy. As MAP kinase activation is well established to be linked to cardiomyopathy, determining how A-type lamin mutations cause activation of MAP kinases will provide key information on the molecular pathogenesis of cardiomyopathy. We hypothesize that EDMD causing mutations lamin A facilitate MAP kinase activation via two distinct mechanisms: 1) mutant lamin A may disrupt the UNC complex, compromising the integrity of the nucleo- cytoskeleton and generating a stress response that activates the MAP kinase pathway and 2) functional disruption of MAP kinase anchors by direct and indirect mechanisms. Lamin A mutations may directly inhibit the function of lamin A as a putative MAP kinase sequestering anchor to facilitate nuclear translocation. Further, mutations in lamin A may disrupt the LINC complex and hence, the function of cytoplasmic anchors that require intact cytoskeleton. To test our hypothesis, we propose two specific aims. Aim 1 will determine whether disrupting the LINC complex in primary cardiomyocytes will activate MAP kinases. We will disrupt the LINC complex by expressing mutant forms of lamin A, suns, and nesprins that prevent interactions necessary to establish the LINC complex and assess for MAP kinase activation by standard biochemical and fluorescence microscopy techniques. Aim 2 will characterize lamin A and MAP kinase interactions in intact cells by various methods and the effect of lamin A mutations on this interaction. We will assess the functional consequence of lamin A mutation on MAP kinase anchors by measuring the mobility of MAP kinases by FRAP and FLIP analysis. Heart muscle damage is the most serious and life-threatening symptom of EDMD. Understanding how defective proteins of the cell nucleus lead to EDMD-associated heart muscle damage will provide the knowledge necessary to devise an effective treatment strategy. This knowledge may also be applicable to similar heart muscle damage resulting from disorders other than EDMD.

描述（由申请人提供）：本申请的总体目标是确定与Emery-Dreifuss肌营养不良（EDMD）相关的心肌病的分子发病机制。常染色体显性EDMD是由编码A型核纤层蛋白的基因突变引起的，A型核纤层蛋白是参与维持核结构的中间丝。此外，A型核纤层蛋白（沿着太阳和nesprins）是LINC复合物的组分，其在细胞核和肌动蛋白细胞骨架之间建立物理连接。在EDMD的EDMD小鼠模型中，有丝分裂原活化蛋白（MAP）激酶ERK和JNK在心肌病发展之前在心脏组织中被激活。由于MAP激酶激活已被证实与心肌病有关，因此确定A型核纤层蛋白突变如何引起MAP激酶激活将为心肌病的分子发病机制提供关键信息。我们假设引起EDMD的突变核纤层蛋白A通过两种不同的机制促进MAP激酶活化：1）突变核纤层蛋白A可能破坏MAP复合物，损害核细胞骨架的完整性并产生激活MAP激酶途径的应激反应，和2）通过直接和间接机制功能性破坏MAP激酶锚。核纤层蛋白A突变可能直接抑制核纤层蛋白A作为一个假定的MAP激酶螯合锚，以促进核转位的功能。此外，核纤层蛋白A中的突变可能破坏LINC复合物，并因此破坏需要完整细胞骨架的细胞质锚的功能。为了验证我们的假设，我们提出了两个具体目标。目的1将确定在原代心肌细胞中破坏LINC复合物是否会激活MAP激酶。我们将通过表达核纤层蛋白A、suns和nesprins的突变形式破坏LINC复合物，这些突变形式阻止建立LINC复合物所必需的相互作用，并通过标准生化和荧光显微镜技术评估MAP激酶活化。目的2将通过各种方法表征完整细胞中核纤层蛋白A和MAP激酶的相互作用以及核纤层蛋白A突变对这种相互作用的影响。我们将通过FRAP和FLIP分析测量MAP激酶的迁移率来评估核纤层蛋白A突变对MAP激酶锚的功能后果。心肌损害是EDMD最严重和危及生命的症状。了解细胞核中有缺陷的蛋白质如何导致EDMD相关的心肌损伤，将为设计有效的治疗策略提供必要的知识。这一知识也可能适用于EDMD以外的疾病引起的类似心肌损伤。