Role of Troponin T Isoforms in Nemaline Myopathy

肌钙蛋白 T 同工型在线形肌病中的作用

基本信息

批准号：
8531650
负责人：
Jian-Ping Jin
金额：
$ 31.85万
依托单位：
WAYNE STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2002
资助国家：
美国
起止时间：
2002-12-01 至 2017-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8531650
关键词：
3 year old Acidosis Adult Affect Age Alleles Amino Acids Amish Animal Model Animals Apoptosis Appearance Basic Science Binding Biochemical Biological Biology Birth Breast Breast Diseases Cardiac Case Study Cells Cessation of life Characteristics Charge Chickens Child Clinic Clinical Medicine Codon Nucleotides Communities Complement Complex Contracture County Cytosol Deformity Development Disease Disease Progression Dissociation Dominant-Negative Mutation Down-Regulation Embryo Enhancers European Event Exhibits Family Fatigue Fiber Financial compensation Frequencies Functional disorder Future Gene Dosage Gene Expression General Population Genes Genetic Goals Haploidy Health Healthcare Hereditary Disease Heterozygote Immigrant Incidence Individual Infant Inherited Investigation Isoelectric Point Knock-in Mouse Life Link Medicine Messenger RNA Metabolic Methods Microfilaments Molecular Molecular Genetics Muscle Muscle Cells Muscle Contraction Muscle Development Muscle Fibers Muscle Tremors Muscle Weakness Muscle function Muscle hypotonia Muscular Atrophy Mutagenesis Mutant Strains Mice Mutation Myofibrils Myopathy N-terminal Names Nemaline Myopathies Neonatal Neuromuscular Diseases Nonsense Mutation Nuclear Family Nucleotides Ohio Pathogenesis Pathology Patients Pennsylvania Perinatal Phenotype Physiological Play Population Postpartum Period Progress Reports Protein Isoforms Proteins Proteolysis RNA Splicing Rare Diseases Reagent Regulation Reporting Research Research Personnel Research Project Grants Respiratory Failure Respiratory Insufficiency Role Shapes Skeletal Muscle Slow-Twitch Muscle Fibers Stress Striated Muscles Supplementation Symptoms System Termination of pregnancy Terminator Codon Testing Therapeutic Thin Filament Time Tremor Tropomyosin Troponin Troponin T United States National Institutes of Health Up-Regulation Vertebrates Work base bench to bedside career cytotoxicity dosage effective therapy fetal fetal diagnosis genetic regulatory protein in utero insight loss of function meetings mouse model muscle degeneration novel polypeptide postnatal promoter retinal rods skeletal wasting

项目摘要

DESCRIPTION (provided by applicant): Amish Nemaline Myopathy (ANM) is an autosomal recessive muscle disorder found among the Old Order Amish in Pennsylvania and Ohio, affecting 1 out of every ~500 births. The ANM allele contains a nonsense mutation in the slow skeletal muscle troponin T (TnT) gene (TNNT1), which results in truncation of the TnT protein at amino acid 179. This truncated slow TnT cannot incorporate into myofibrils and is degraded inside the myocyte. Phenotypically, individuals with ANM suffer from muscle tremors, contractures and hypotonia. The affected infants are clinically normal at birth but rapidly develop the ANM symptoms that usually result in death from respiratory failure during the second year. No effective treatment is available. This research project aims to understanding the pathogenesis of ANM and the pathophysiology of ANM muscle for the optimal goal of developing a cure of the disease. The following three specific aims are proposed: Aim I: To characterize animal models deficient in slow TnT. We shall study the skeletal muscle function in a slow TnT knockdown mouse model in which lowered slow TnT gene expression results in decreased slow TnT protein, muscle atrophy and a switch to fast fiber phenotypes with decreased tolerance to fatigue. We shall also study the phenotype of a mouse model in which the ANM nonsense mutation is knocked in the slow TnT gene to investigate the pathogenesis and disease progression. Aim II: To examine the conditional effects of slow TnT gene haploidy and cytotoxicity of the truncated slow TnT on ANM pathogenesis and pathophysiology. ANM heterozygotes have reported circumstantial muscle symptoms. We shall investigate the potential effect of half dosage of slow TnT gene on muscle function in heterozygote slow TnT knock-in mutant mice. Non-myofilament-associated TnT fragments exhibit cytotoxicity. We shall examine whether and how this potential cytotoxicity of the ANM slow TnT fragment contributes to muscle degeneration in ANM patients. Aim III: To study the regulation and function of cardiac TnT in skeletal muscle in order to develop therapeutic compensation in ANM patients. We have found evidence in a subtype of ANM that the loss of slow skeletal TnT function in ANM may be partially compensated for by the continuing expression of cardiac TnT in skeletal muscles. Functional significance and activation of this compensatory cardiac TnT gene expression will be investigated at protein, muscle cell and tissue, and animal levels toward the development of a specific treatment for ANM. In an alternative approach, suppression of the ANM nonsense stop codon will also be explored using emerging reagents. Based on progresses we have made to date, these studies employ state-of-the-art molecular genetic, biochemical, cell biological and physiological methods and novel experimental systems. The results will significantly further the pursuit of an effective therapy for this devastating disease. PUBLIC HEALTH RELEVANCE: Nemaline myopathies are a group of neuromuscular disorders characterized by muscle weakness and rod-shaped "nemaline" inclusions in skeletal muscle fibers. All known inherited nemaline myopathies are caused by mutations in sarcomeric thin filament proteins. Amish Nemaline Myopathy (ANM) is a lethal inherited nemaline myopathy present at very high incidences (1 out of 500 births) in the Old Order Amish communities in Pennsylvania and Ohio. The genetic basis of ANM is a single nucleotide nonsense mutation in the gene encoding the slow skeletal muscle isoform of troponin T (TnT), a muscle-specific Ca2+- regulatory protein. This nonsense mutation truncates the slow TnT polypeptide at amino acid 179 and renders the TnT protein incapable of incorporating into myofibrils and being rapidly degraded. This loss of function mechanism is consistent with the autosomal recessive inheritance of the disease. Phenotypically, individuals with ANM suffer from muscle tremors, contractures and hypotonia. Symptoms are trivial at birth but rapidly worsen, usually resulting in death from respiratory failure during the second year. No effective treatment is currently available. Although ANM is a rare disease in the general population, its uniformly devastating progression combined with an already elucidated molecular cause merits much further investigation. Two to three ANM babies are born every year in Pennsylvania and Ohio; none will live to be 3 years old. We are currently the only research team in the world working on ANM and our progress thus far has already given the affected families and the general Amish community a hope for effective treatments in the near future. This research project aims to further our progress towards the development of a therapy by investigating TnT isoform function and regulation. We will study slow TnT-deficient animal models for the understanding of ANM pathology and muscle function. We will examine the potential effect of decreased slow TnT gene dosage on muscle function and the cytotoxicity of truncated slow TnT for links to muscle degeneration in ANM. Together with exploring the possibility of suppressing the ANM nonsense stop codon, we will focus on investigating the functional significance and activation of compensatory cardiac TnT expression in slow TnT-deficient skeletal muscle for use as a specific treatment for ANM. ANM is the only known disease caused by a recessive mutation in a TnT gene. By investigating the pathophysiology of ANM, we will also gain important insights into the functional significance of different TnT isoforms and the Ca2+-regulation of striated muscle contraction, a fundamental topic in biology and medicine.

描述（由申请人提供）：阿米什nemaline肌病（ANM）是一种常染色体隐性肌肉障碍，在宾夕法尼亚州和俄亥俄州的旧秩序阿米什人中发现，每500个出生中有1个。 ANM等位基因在慢速骨骼肌肌钙蛋白T（TNT）基因（TNNT1）中包含一个胡说八道的突变，这会导致氨基酸179处TNT蛋白的截断。这种截断的慢速TNT不能掺入肌原纤维中，并且在肌细胞内降解。从表型上讲，患有ANM的个体患有肌肉震颤，染色和性低下。受影响的婴儿在出生时在临床上是正常的，但迅速发展了ANM症状，通常导致第二年导致呼吸衰竭死亡。没有有效的治疗方法。该研究项目旨在理解ANM的发病机理和ANM肌肉的病理生理学，以开发治愈该疾病的最佳目标。提出了以下三个特定目标：目标I：表征慢速TNT缺乏的动物模型。我们将研究慢慢的TNT敲低小鼠模型中的骨骼肌功能，其中降低的慢速TNT基因表达会导致慢速TNT蛋白，肌肉萎缩和转向快速纤维表型，并且对疲劳的耐受性降低。我们还将研究小鼠模型的表型，其中慢速TNT基因中敲打ANM胡说八道突变以研究发病机理和疾病进展。 AIM II：检查慢速TNT基因单倍体的条件作用和截短的慢速TNT对ANM发病机理和病理生理学的细胞毒性。 ANM杂合子报告了环境肌肉症状。我们将研究一半剂量的慢速TNT基因对杂合子慢速TNT敲击突变小鼠的肌肉功能的潜在影响。非肉眼相关的TNT片段表现出细胞毒性。我们将检查ANM慢TNT片段的这种潜在细胞毒性是否有助于ANM患者的肌肉变性。 AIM III：研究心脏TNT在骨骼肌中的调节和功能，以在ANM患者中发展治疗补偿。我们已经在ANM的亚型中发现了证据表明，ANM缓慢骨骼TNT功能的丧失可能会因骨骼肌中心脏TNT的持续表达而部分补偿。该补偿性心脏TNT基因表达的功能显着性和激活将在蛋白质，肌肉细胞和组织以及动物水平上研究ANM的特定治疗方法。在另一种方法中，还将使用新兴试剂探索对ANM废话停止密码子的抑制。根据我们迄今为止所取得的进展，这些研究采用了最先进的分子遗传，生化，细胞生物学和生理方法以及新型实验系统。结果将大大进一步追求这种毁灭性疾病的有效疗法。公共卫生相关性：黑甲肌病是一组神经肌肉疾病，其特征是骨骼肌纤维中的肌肉无力和棒状的“ nemaline”夹杂物。所有已知的遗传性黑叶肌病都是由肌肉细细丝蛋白突变引起的。阿米什（Amish Nemaline）肌病（ANM）是一种致命的遗传性nemaline肌病，在宾夕法尼亚州和俄亥俄州的旧秩序阿米什（Amish）社区中以非常高的事件（500个出生中的1个）出现。 ANM的遗传基础是编码肌蛋白T（TNT）的慢速骨骼肌同工型的基因中的单个核苷酸胡说八道突变，这是一种肌肉特异性Ca2+ - 调节蛋白。这种废话的突变会截断氨基酸179处的慢速TNT多肽，并使TNT蛋白无法掺入肌纤维并迅速降解。这种功能机制的丧失与疾病的常染色体隐性继承一致。从表型上讲，患有ANM的个体患有肌肉震颤，染色和性低下。症状在出生时是微不足道的，但迅速恶化，通常导致第二年因呼吸衰竭而死亡。目前没有有效的治疗方法。尽管ANM在普通人群中是一种罕见的疾病，但其统一的毁灭性进展与已经阐明的分子原因相结合，值得进一步研究。每年在宾夕法尼亚州和俄亥俄州出生两到三个ANM婴儿。没有人会活到3岁。目前，我们是世界上唯一从事ANM工作的研究团队，到目前为止，我们的进步已经为受影响的家庭和普通社区提供了希望在不久的将来对有效治疗的希望。该研究项目旨在通过研究TNT同工型功能和调节来进步，以进一步发展治疗。我们将研究缓慢的TNT缺乏动物模型，以了解ANM病理和肌肉功能。我们将检查慢速TNT基因剂量减少对肌肉功能的潜在影响以及与ANM中肌肉变性的链接的截短慢速TNT的细胞毒性。加上探索抑制ANM胡说八道密码子的可能性，我们将集中精力研究缓慢的TNT缺陷骨骼肌中代偿性心脏TNT表达的功能意义和激活，以用作ANM的特定治疗方法。 ANM是TNT基因中隐性突变引起的唯一已知疾病。通过研究ANM的病理生理学，我们还将对不同TNT同工型的功能意义以及横纹肌肉收缩的CA2+调节的功能意义，这是生物学和医学中的基本话题。