Allele-specific Effects-Single Amino Acid Exchanges/cTnT

等位基因特异性效应-单氨基酸交换/cTnT

• 批准号: 7216515
• 负责人: Jil C Tardiff
• 金额: $ 3.42万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-12-01 至 2007-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7216515
• 关键词: alleles; aminoacid metabolism; cardiovascular disorder epidemiology; cardiovascular function; cardiovascular stress test; confocal scanning microscopy; congenital heart disorder; electron microscopy; gene mutation; gene targeting; genetic disorder; genetic markers; genetically modified animals; heart contraction; heart dimension /size; hypertrophic myocardiopathy; laboratory mouse; microfilaments; myocardium; myofibrils; protein protein interaction; protein structure function; sarcomeres; tropomyosin; troponin

DESCRIPTION (provided by applicant): The regulatory function of the cardiac sarcomere resides in the thin filament. It is a complex macromolecular structure comprised of multiple protein subunits, which interact and modulate contractile function in response to both chronic and acute physiologic stress. The binding of Ca2+ to Troponin C initiates a cascade of allosteric changes in the interactions of the proteins within the troponin (cTnC, cTnl and cTnT) and tropomyosin-actin complexes, facilitating the formation of the actinomyosin complex and the power stroke of muscle contraction. While normal hearts retain the ability to alter these complex interactions, for example, via contractile protein isoform shifts and post-translational modifications, many naturally-occurring thin filament mutations are poorly tolerated. In fact, mutations in cardiac Troponin T (cTnT) result in a particularly severe form of Familial Hypertrophic Cardiomyopathy (FHC) characterized by a high frequency of early sudden cardiac death in the absence of overt ventricular hypertrophy. The direct link between the gene mutation and the complex clinical phenotype remains unknown. Troponin T is a highly elongated protein that interacts with all other components of the thin filament and has been described as the "glue" of the regulatory system. The full ternary structure is unknown and thus the details of these complex protein-protein interactions remain unclear. The hypothesis for this application is that individual cTnT mutations disrupt discrete aspects of these protein-protein interactions causing specific changes in thin filament function and leading to distinct cardiovascular phenotypes. To address this hypothesis we have developed three independent transgenic mouse models based solely on amino acid substitutions at Codon 92 (Arg92GIn, Arg92Trp and Arg92Leu), a known mutational hotspot in human cTnT. Initial characterization has revealed allele-specific changes in cardiac mass, contractility and hypertrophic signaling pathways. In these initial studies we have selected specific aspects of the observed phenotypes to serve as physiologic "indicators" of the differences in thin filament function caused by each mutation. Establishing the mechanistic link between the specific cTnT mutant allele and it's cellular phenotype will both provide new insights into the fundamental biology of thin filament function and further our understanding of the pathogenic pathways involved in thin filament related cardiomyopathies.

描述（申请人提供）：心肌肌节的调控功能存在于细丝中。它是一个复杂的大分子结构，由多个蛋白质亚基组成，在慢性和急性生理应激下相互作用和调节收缩功能。Ca2+与肌钙蛋白C的结合启动了肌钙蛋白（cTnC， cTnl和cTnT）和原肌球蛋白-肌动蛋白复合物内蛋白质相互作用的级联变构变化，促进了放线肌球蛋白复合物的形成和肌肉收缩的力量中风。虽然正常心脏保留了改变这些复杂相互作用的能力，例如，通过收缩蛋白异构体移动和翻译后修饰，但许多自然发生的细丝突变耐受性很差。事实上，心肌肌钙蛋白T （cTnT）突变可导致一种特别严重的家族性肥厚性心肌病（FHC），其特征是在没有明显心室肥厚的情况下，早期心源性猝死的频率很高。基因突变与复杂临床表型之间的直接联系尚不清楚。肌钙蛋白T是一种高度细长的蛋白质，与细丝的所有其他成分相互作用，被描述为调节系统的“胶水”。完整的三元结构是未知的，因此这些复杂的蛋白质-蛋白质相互作用的细节仍然不清楚。该应用的假设是，单个cTnT突变破坏了这些蛋白质-蛋白质相互作用的离散方面，导致细丝功能的特定变化，并导致不同的心血管表型。为了解决这一假设，我们开发了三种独立的转基因小鼠模型，仅基于密码子92 （Arg92GIn， Arg92Trp和Arg92Leu）的氨基酸替换，这是人类cTnT中已知的突变热点。初步表征揭示了心脏质量、收缩性和肥厚信号通路的等位基因特异性变化。在这些初步研究中，我们选择了观察到的表型的特定方面作为每个突变引起的细丝功能差异的生理“指标”。建立特定cTnT突变等位基因与其细胞表型之间的机制联系，将为细丝功能的基础生物学提供新的见解，并进一步了解细丝相关心肌病的致病途径。