Molecular Mechanisms of Disease in a Novel Feline Model of Familial Hypertrophic

新型家族性肥厚型猫科动物模型中疾病的分子机制

• 批准号: 8111960
• 负责人: Samantha P Harris
• 金额: $ 19.04万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-15 至 2014-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8111960
• 关键词: Adolescent; Affect; Alanine; Alleles; Amino Acid Substitution; Amino Acids; Animal Model; Antibodies; Arrhythmia; Basic Science; Biopsy; Breeding; Cardiac; Cardiac Myocytes; Cardiac Myosins; Cardiomyopathies; Cell Fractionation; Cell physiology; Cells; Codon Nucleotides; Data; Defect; Detergents; Development; Disease; Dominant-Negative Mutation; Engineering; Epitopes; Family Felidae; Felis catus; Functional disorder; Gene Dosage; Genetic; Goals; Growth; Heart; Homeostasis; Human; Hypertrophic Cardiomyopathy; Hypertrophy; Impairment; Inherited; Knock-out; Knockout Mice; Lead; Life; Link; Lysosomes; Maine; Measurement; Measures; Messenger RNA; Microfilaments; Missense Mutation; Modeling; Molecular; Mus; Muscle; Muscle Cells; Mutation; Myocardial; Myocardium; Myofibrils; Neonatal; Nonsense Codon; Output; Pathway interactions; Point Mutation; Poison; Preparation; Process; Proline; Property; Proteins; Rattus; Reporting; Research; Resources; Sampling; Sarcomeres; Signal Transduction; Skin; Sudden Death; System; Techniques; Testing; Therapeutic; Tissues; Ubiquitin; Western Blotting; citrate carrier; design; disease diagnosis; disease phenotype; disease-causing mutation; functional loss; human disease; indexing; insight; mouse model; multicatalytic endopeptidase complex; mutant; myosin-binding protein C; novel; outcome forecast; polypeptide; protein aggregate; protein folding; public health relevance; research study; response; sudden cardiac death; trafficking; treatment strategy; young adult

DESCRIPTION (provided by applicant): The long-range goal of the proposed research is to understand the molecular mechanisms by which mutations in Myosin Binding Protein-C (MyBP-C) cause hypertrophic cardiomyopathy (HCM), an autosomal dominant disorder that affects 1 in 500 people and is the most common cause of sudden cardiac death in adolescents and young adults. Mutations in cardiac (c) MyBP-C are among the most frequent causes of HCM with >149 distinct mutations described so far. However, despite progress in identifying genetic causes of HCM, the molecular mechanism(s) by which any single cMyBP-C mutation causes disease are still unknown. Reduced amounts of cMyBP-C were recently reported in affected human myocardium, suggesting that loss of functional cMyBP-C from an affected allele (i.e., haploinsufficiency) is a common factor contributing to cardiac dysfunction. However, alternative possibilities that affected proteins impair contractile function or that processing and degradation of improperly folded proteins cause aberrant cell function have not been eliminated. These distinctions are critical for designing effective therapeutic strategies to overcome HCM, yet definitive evidence in support of the different possibilities has not been obtained in part because of the limited availability of human biopsies and in part because engineered mouse models do not fully recapitulate either the human disease phenotype or the proximal cell processes that lead to disease. Experiments proposed here will overcome these limitations by utilizing the only naturally occurring large animal model of HCM that both closely resembles the human disease phenotype and that has a known genetic cause. The mutation is a spontaneous missense mutation in cMyBP-C in domestic Maine Coon cats that results in a proline for alanine substitution at codon 31 (A31P). The mutation results in a single amino acid substitution, but causes an anomalous decrease in total amounts of cMyBP-C protein. Because similar decreases in cMyBP-C have been reported for missense cMyBP-C mutations in human myocardium, the feline A31P model offers a unique opportunity to distinguish between three primary factors proposed as causative in human disease i.e., dominant negative effects of a single amino acid point mutation, gene dosage effects, and cellular protein folding/trafficking defects. The proposed experiments will test the hypothesis that reduced amounts of cMyBP-C in sarcomeres (i.e., haploinsufficiency) of affected cats leads to contractile deficits that ultimately cause cardiac dysfunction. Specific aims of the project are to determine 1) the expression level and subcellular localization of total cMyBP-C protein in cats carrying the A31P mutation, 2) effects of the A31P mutation on myocyte contractile properties, and 3) effects of the A31P mutation on the myocardial ubiquitin-proteasome (UPS) system. Collectively, results from these studies will provide critical insights into molecular mechanisms by which mutations in cMyBP-C cause disease and will provide a significant and lasting impact on HCM research by developing a unique animal model that will be a resource for basic research and that will aid in the design and testing of therapeutic strategies for the treatment of cardiomyopathies linked to cMyBP-C. PUBLIC HEALTH RELEVANCE: The proposed studies will investigate basic pathogenic mechanisms by which the A31P mutation in the cardiac myosin binding protein-C (cMyBP-C) causes inherited hypertrophic cardiomyopathy (HCM) in Maine coon cats, a breed of domestic cat. In humans, HCM affects an estimated 1 in 500 people and is the leading cause of sudden death in adolescent and young adults. Because mutations affecting cMyBP-C are a leading cause of HCM in humans, it is anticipated that the proposed studies in Maine Coon cats will provide insights into human disease and will ultimately contribute to advances in disease diagnosis, prognosis, and treatment of HCM linked to cMyBP-C.

描述（由申请人提供）：拟议研究的长期目标是了解肌球蛋白结合蛋白-C（MyBP-C）突变导致肥厚性心肌病（HCM）的分子机制，HCM是一种常染色体显性遗传疾病，影响1/500人，是青少年和年轻人心脏性猝死的最常见原因。心脏（c）MyBP-C突变是HCM最常见的原因之一，迄今为止已描述了超过149种不同的突变。然而，尽管在确定HCM的遗传原因方面取得了进展，但任何单个cMyBP-C突变导致疾病的分子机制仍然未知。最近报道了在受影响的人心肌中cMyBP-C的量减少，这表明来自受影响等位基因的功能性cMyBP-C的丧失（即，单倍不足）是导致心脏功能障碍的常见因素。然而，受影响的蛋白质损害收缩功能或不正确折叠的蛋白质的加工和降解导致细胞功能异常的替代可能性尚未消除。这些区别对于设计有效的治疗策略来克服HCM是至关重要的，但尚未获得支持不同可能性的明确证据，部分原因是人类活检的可用性有限，部分原因是工程小鼠模型不能完全重现人类疾病表型或导致疾病的近端细胞过程。这里提出的实验将克服这些限制，利用唯一的自然发生的大型动物模型HCM，既密切类似于人类疾病的表型，并具有已知的遗传原因。该突变是家养缅因州浣熊猫cMyBP-C中的自发错义突变，导致密码子31处的脯氨酸替换为丙氨酸（A31 P）。该突变导致单个氨基酸取代，但导致cMyBP-C蛋白总量异常减少。因为已经报道了人类心肌中错义cMyBP-C突变的cMyBP-C的类似降低，所以猫A31 P模型提供了一个独特的机会来区分被认为是人类疾病病因的三个主要因素，即，单一氨基酸点突变的显性负效应、基因剂量效应和细胞蛋白质折叠/运输缺陷。所提出的实验将检验以下假设：肌节中cMyBP-C的量减少（即，单倍不足）导致收缩缺陷，最终导致心脏功能障碍。本项目的具体目的是确定1）携带A31 P突变的猫中总cMyBP-C蛋白的表达水平和亚细胞定位，2）A31 P突变对心肌细胞收缩特性的影响，以及3）A31 P突变对心肌泛素-蛋白酶体（UPS）系统的影响。总的来说，这些研究的结果将为cMyBP-C突变导致疾病的分子机制提供重要的见解，并将通过开发一种独特的动物模型对HCM研究产生重大而持久的影响，该模型将成为基础研究的资源，并将有助于设计和测试治疗与cMyBP-C相关的心肌病的治疗策略。 公共卫生相关性：拟议的研究将调查心脏肌球蛋白结合蛋白-C（cMyBP-C）中的A31 P突变导致缅因州浣熊（一种家猫）遗传性肥厚型心肌病（HCM）的基本致病机制。在人类中，HCM影响约1/500人，是青少年和年轻人猝死的主要原因。由于影响cMyBP-C的突变是人类HCM的主要原因，因此预计在缅因州浣熊中进行的拟议研究将提供对人类疾病的见解，并最终有助于疾病诊断、预后和与cMyBP-C相关的HCM治疗的进展。