Golgi-associated NO-cGMP Signaling Defect in Muscular Dystrophy

肌营养不良症中高尔基体相关的 NO-cGMP 信号缺陷

基本信息

批准号：
8303029
负责人：
Joseph A Beavo
金额：
$ 58.91万
依托单位：
UNIVERSITY OF WASHINGTON
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-09-25 至 2014-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8303029
关键词：
Actins Address Adrenergic Agents Amino Acids Antibodies Applications Grants Attention Attenuated Automobile Driving Bathing Biochemical Biological Assay Biotin Blood Vessels Brain Caliber Cardiac Cell Line Cell Nucleus Cells Characteristics Clinical Complement Complex Contracts Coupling Cyclic GMP Cyclic GMP-Dependent Protein Kinases Cyclic Nucleotides Cytoplasm Cytoskeleton Data Defect Diffuse Diffusion Duchenne muscular dystrophy Dystrophin Electric Stimulation Enzymes Excision Exhibits Exons Extracellular Matrix Fatigue Female Fiber G-substrate Generations Genetic Golgi Apparatus Health Heart Immunoblotting Injury Ion Channel Ischemia Knockout Mice Knowledge Laboratories Lead Maintenance Mass Spectrum Analysis Measures Mediating Membrane Methodology Methods Modification Molecular Mus Muscle Muscle Contraction Muscle Development Muscle Fibers Muscle Proteins Muscle function Muscular Dystrophies Myoblasts Myocardium Myopathy NOS1 gene Nerve Neurons Nitric Oxide Nitric Oxide Pathway Nitric Oxide Synthase Nitric Oxide Synthase Type I Organelles Pathway interactions Patients Pattern Phenotype Phosphodiesterase Inhibitors Phosphorylation Phosphotransferases Physiological Post-Translational Protein Processing Predisposition Preparation Production Property Protein Binding Protein Isoforms Proteins Proteomics Publishing RNA Splicing Recovery Regulation Reporting Research Residual state Resistance Role Ryanodine Receptor Calcium Release Channel SKIL gene Sarcolemma Scaffolding Protein Severities Signal Pathway Signal Transduction Signal Transduction Pathway Signaling Protein Site Skeletal Muscle Sodium Dodecyl Sulfate-PAGE Soleus Muscle Soluble Guanylate Cyclase Surface Testing Transgenic Mice Transgenic Organisms Two-Dimensional Gel Electrophoresis Variant Vasoconstrictor Agents Vasodilation adrenergic aged attenuation base boys drinking water glucose transport improved inhibitor/antagonist male mdx mouse muscle degeneration muscle form mutant new therapeutic target novel overexpression palmitoylation phosphoric diester hydrolase protein transport public health relevance response scaffold sex sildenafil syntrophin tadalafil therapeutic target tibialis anterior muscle tool vasoconstriction water channel

项目摘要

DESCRIPTION (provided by applicant): The mechanism by which the loss of dystrophin leads to Duchenne Muscular Dystrophy is generally attributed to membrane fragility due to disruption of the linkage between the extracellular matrix and the cortical actin cytoskeleton. This structural role for dystrophin and its associated proteins is supported by substantial evidence. In addition, the dystrophin complex serves as a sarcolemmal scaffold for signaling proteins, including neuronal nitric oxide synthase (nNOS5, the muscle-specific isoform), several kinases, water and ion channels and other proteins. The demonstration that transgenic expression of nNOS in mdx mouse muscle ameliorates the dystrophic phenotype has focused attention on this signaling protein. Recently, we have found that a second isoform of this enzyme, nNOS2, is localized on the Golgi, and that its expression at this site is reduced in mdx muscle, even before the first phenotypic signs of dystrophy are evident. Furthermore, we find that soluble guanylate cyclase (sGC), which is activated by NO, and cGMP-activated protein kinase G (PKG) are colocalized with nNOS2 on the Golgi. Mice lacking both nNOS5 and nNOS2 (KN2) have numerous deficiencies, including a severe myopathic defect. Based on these preliminary results, we will characterize muscle from KN2 mice at the structural, biochemical and functional levels. In addition, we will test the hypothesis that transgenic expression of nNOS2 in skeletal muscle will rescue/ameliorate the KN2 and mdx phenotypes. The role of nNOS2 in blunting 1-adrenergic vasoconstriction in contracting muscle will provide information on the importance of this enzyme in alleviating functional ischemia, a component of muscular dystrophy. Finally, we will determine the Golgi substrates of the NO/cGMP/PKG pathway and of direct protein nitrosylation. Finally, we will identify the phosphodiesterase(s) (PDEs) that regulate(s) the NO/cGMP/PKG pathway. A complete understanding of this signaling pathway will enable a more rationale approach to selecting therapeutic targets, including PDE inhibitors, which slow the progression of muscle degeneration and ameliorate the dystrophic phenotype. PUBLIC HEALTH RELEVANCE: The research proposed here will provide new information about the mechanisms that lead to muscle degeneration in muscular dystrophies. The focus on a novel signaling mechanism at the Golgi, a cellular organelle that regulates protein modification and trafficking to the surface membrane, may reveal new therapeutic targets for slowing the progress of muscle degeneration. In general, the knowledge generated by this project will be relevant to many types of muscle diseases.

描述（由申请人提供）：肌营养不良蛋白缺失导致杜氏肌营养不良症的机制通常归因于细胞外基质和皮质肌动蛋白细胞骨架之间的连接破坏导致的膜脆性。抗肌萎缩蛋白及其相关蛋白的这种结构作用得到了大量证据的支持。此外，抗肌营养不良蛋白复合物还可作为信号蛋白的肌膜支架，包括神经元一氧化氮合酶（nNOS5，肌肉特异性亚型）、几种激酶、水和离子通道以及其他蛋白质。 mdx 小鼠肌肉中 nNOS 转基因表达可改善营养不良表型的证明已将注意力集中在这种信号蛋白上。最近，我们发现这种酶的第二种亚型 nNOS2 位于高尔基体上，并且甚至在营养不良的第一个表型迹象明显之前，它在 mdx 肌肉中的表达就减少了。此外，我们发现由 NO 激活的可溶性鸟苷酸环化酶 (sGC) 和 cGMP 激活的蛋白激酶 G (PKG) 与高尔基体上的 nNOS2 共定位。同时缺乏 nNOS5 和 nNOS2 (KN2) 的小鼠存在许多缺陷，包括严重的肌病缺陷。基于这些初步结果，我们将从结构、生化和功能水平上表征 KN2 小鼠的肌肉。此外，我们将测试骨骼肌中 nNOS2 的转基因表达将挽救/改善 KN2 和 mdx 表型的假设。 nNOS2 在减弱肌肉收缩过程中 1-肾上腺素能血管收缩方面的作用将提供有关该酶在缓解功能性缺血（肌营养不良症的一个组成部分）中的重要性的信息。最后，我们将确定 NO/cGMP/PKG 途径和直接蛋白质亚硝基化的高尔基体底物。最后，我们将确定调节 NO/cGMP/PKG 途径的磷酸二酯酶 (PDE)。对该信号通路的全面了解将使我们能够采取更合理的方法来选择治疗靶点，包括 PDE 抑制剂，它可以减缓肌肉退化的进展并改善营养不良表型。公共健康相关性：此处提出的研究将提供有关导致肌营养不良症中肌肉退化的机制的新信息。对高尔基体（一种调节蛋白质修饰和运输到表面膜的细胞器）的新型信号传导机制的关注可能会揭示减缓肌肉退化进程的新治疗靶点。一般来说，该项目产生的知识将与多种类型的肌肉疾病相关。