Improving cell membrane repair to treat muscular dystrophy

改善细胞膜修复以治疗肌营养不良症

基本信息

批准号：
10674686
负责人：
Miguel Aaron Lopez Perez
金额：
$ 4.31万
依托单位：
OHIO STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-08-01 至 2024-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10674686
关键词：
Address Age Amino Acid Motifs Binding Biochemical Biological Assay Birth C-terminal Cell Death Cell membrane Cessation of life Childhood Coupled Data Development Disease Dot Immunoblotting Duchenne muscular dystrophy Dystrophin Engineering Eukaryotic Cell Event Fellowship Fluorescence Future Genes Genetic Diseases Goals Heart Injuries Homeostasis Human In Vitro Incidence Injury Knockout Mice Laboratories Laser injury Lead Length Life Link Lipids Longevity Mammalian Cell Measurement Measures Mechanical Stress Mediating Membrane Modeling Molecular Mus Muscle Muscle Cells Muscle Development Muscle Fibers Muscle Weakness Muscular Atrophy Muscular Dystrophies Mutation Myocardial Infarction Myocardium Myopathy Necrosis Neurodegenerative Disorders Neuromuscular Diseases Pathology Patients Phenotype Phosphatidylserines Play Predisposition Protein Binding Domain Protein Engineering Protein Family Proteins Quality of life Recombinant Proteins Recombinants Regenerative capacity Reporting Research Design Research Personnel Role Rotation Sarcolemma Site Skeletal Muscle Structural Protein Subgroup TRIM Family TRIM Gene TRIM Motif Tertiary Protein Structure Testing Therapeutic Therapeutic Effect Training Treatment Efficacy Tryptophan Utrophin Wasting Syndrome X-Linked Genetic Diseases career disease-causing mutation experience falls improved in vivo innovation male mdx mouse member molecular targeted therapies mouse model muscle degeneration muscle strength mutant new therapeutic target novel novel therapeutics overexpression prevent protein expression repaired response severe injury skeletal skill acquisition therapeutic development therapeutic protein training opportunity translational potential ubiquitin-protein ligase

项目摘要

Project Summary Duchenne Muscular Dystrophy (DMD) is a fatal degenerative muscle disease caused by mutations in the X- linked dystrophin gene. DMD is the most prevalent form of muscular dystrophy with an incidence of 1:5,000 live male births. Dystrophin mutations lead to the loss of protein expression and compromised sarcolemmal membrane integrity. Injury from mechanical stress often leads to muscle fiber death that eventually overwhelms the regenerative capacity of the muscle. Repeated injury severely reduces skeletal muscle strength contributing to early death by the second to fourth decade of life. Current treatment options can prolong survival and improve quality of life; however, they are not curative. Therefore, there remains a need for novel therapies that target the molecular mechanisms underlying disease pathology. Previous studies determined that the tripartite motif protein 72/mitsugumin 53 (TRIM72/MG53) is essential for proper cell membrane repair in skeletal and cardiac muscle through its binding of phosphatidylserine (PS). Loss of MG53 function has been associated with the development of muscular dystrophy and increased susceptibility to cardiac injury. MG53 is a muscle-enriched TRIM family E3 ubiquitin ligase protein that our laboratory previously reported can increase membrane repair by overexpression or by exogenous application of recombinant MG53 (rhMG53) to ameliorate disease pathology in multiple models of muscular dystrophy. This fellowship project aims to interrogate the role of rhMG53 protein domains in this observed effect on membrane repair. My preliminary data suggest that the E3 ubiquitin ligase activity is not required for the enhanced membrane repair phenotype. This observation supports the hypothesis that the canonical TRIM E3 ubiquitin ligase activity of MG53 is not required for therapeutic membrane repair capacity, therefore therapeutic effects may be achieved by compact versions of the MG53 protein that can bind PS. In this fellowship application I propose to test this hypothesis using three specific and independent aims. Aim 1 screens rhMG53 mutant protein constructs to identify rhMG53 protein domains that are essential for membrane repair. Aim 2 defines the mechanistic role of phosphatidylserine binding to rhMG53 by identifying protein domain binding partners. Aim 3 evaluates the therapeutic potential of identified rhMG53 mutant protein constructs when exogenously delivered to intact muscle fibers ex vivo. These studies will provide an excellent training experience by elucidating previously uncharacterized MG53 protein interactions that will enable the development of novel protein therapeutics for DMD. These experimental efforts will be coupled with additional skill development activities to produce an integrated training experience to allow my continued development toward a career as an independent investigator in the neuromuscular disease field.

项目摘要 Duchenne肌肉营养不良（DMD）是由X-突变引起的致命退化性肌肉疾病连接的肌营养不良基因。 DMD是肌肉营养不良的最普遍的形式，发生率为1：5,000 活着的男性出生。肌营养不良蛋白突变导致蛋白质表达的丧失并损害了肌膜膜完整性。机械压力受伤通常会导致肌肉纤维死亡，最终导致压倒了肌肉的再生能力。重复损伤严重减少骨骼肌力量在生命的第二到第四十年中导致早期死亡。当前的治疗选择可以延长生存并改善生活质量；但是，它们没有治愈。因此，仍然需要针对疾病病理基础的分子机制的新型疗法。先前的研究确定三方基序72/mitsugumin 53（TRIM72/mg53）对于适当的细胞至关重要通过磷脂酰丝氨酸（PS）的结合，骨骼肌和心肌的膜修复。损失MG53 功能与肌肉营养不良的发展有关，并增加了对心脏损伤。 MG53是一种富含肌肉的装饰家族E3泛素连接酶蛋白，我们的实验室先前报道的可以通过过表达或外源应用增加膜修复重组MG53（RHMG53）在多种肌肉营养不良模型中改善疾病病理学。这奖学金项目旨在询问RHMG53蛋白质域在这种观察到的对膜的影响中的作用维修。我的初步数据表明，增强的E3泛素连接酶活性不需要膜修复表型。该观察结果支持了规范修剪E3泛素的假设 MG53的连接酶活性对于治疗膜修复能力不需要，因此治疗效果可以通过可结合PS的MG53蛋白的紧凑型版本来实现。在此奖学金申请中我建议使用三个特定和独立的目标检验这一假设。 AIM 1屏幕RHMG53突变体蛋白质构建体以鉴定RHMG53蛋白质结构域，这对于膜修复至关重要。 AIM 2定义通过鉴定蛋白质结构结合伴侣，磷脂酰甲酯与RHMG53结合的机械作用。 AIM 3评估鉴定出的RHMG53突变蛋白构建体的治疗潜力输送到完整的肌肉纤维后。这些研究将通过阐明先前未表征的MG53蛋白质相互作用，这将使新型的发展 DMD的蛋白质疗法。这些实验努力将与其他技能开发相结合活动以产生综合培训经验，以使我继续发展的职业发展神经肌肉疾病领域的独立研究者。