Molecular Regulation of Titin Elasticity by Post-Translational Modification

翻译后修饰对肌联蛋白弹性的分子调控

基本信息

批准号：
9121728
负责人：
Edward Charles Eckels
金额：
$ 4.32万
依托单位：
COLUMBIA UNIVERSITY HEALTH SCIENCES
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-06-01 至 2019-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9121728
关键词：
Affect Aging Architecture Atherosclerosis Attention Cardiac Cardiac Myocytes Collagen Cysteine Cytoskeleton Diabetes Mellitus Disease Disulfides Elasticity Electron Microscopy Extracellular Structure Filament Fluorescence Microscopy Genetic Goals Heart Heart Diseases Hypertension Impairment Individual Ischemia Lead Length Link Location Magnetism Measures Mechanics Microscopy Modification Molecular Mus Muscle Mutation Myocardial Myocardial Infarction Myocardium Myofibrils Nitrogen Oxidation-Reduction Oxidative Stress Oxygen Performance Peroxonitrite Phenotype Physiological Post-Translational Protein Processing Proline Property Quantum Dots Reaction Reactive Nitrogen Species Reactive Oxygen Species Regulation Reperfusion Therapy Resistance Resolution Role S-Nitrosothiols Sarcomeres Signal Pathway Skeletal Muscle Stretching Structure Sulfhydryl Compounds Thick Filament Thinking Tissues Translating Tyrosine Ursidae Family Ventricular Remodeling Work connectin experience familial dilated cardiomyopathy insight mouse model mutant nanoGold nanometer nitration novel oxidation particle prevent public health relevance research study sensor single molecule

项目摘要

DESCRIPTION (provided by applicant): During the filling of the heart or during the contraction of opposing muscle groups, the sarcomere experiences passive stretch that increases its overall length by several hundred nanometers. In the past, it was believed that extracellular structures, mainly collagen, were responsible for the integrity of the sarcomere and provided resistance to prevent over-stretching of the sarcomere. Within the past thirty years, it has become clear that titin, the third filament of the sarcomere, bears the majority of the force during passive stretch f muscle tissue, and is responsible for setting the optimal working length of the sarcomere. Only in 2012, after genetic sequencing of hundreds of individuals, was it shown that mutations in titin are the leading cause of inherited dilated cardiomyopathy. Now that there exists a clear link between titin mutations and disease, attention has turned towards identifying the normal physiological role of titin. Besides organizing the thick filament, the I-band segment of titin deforms to accommodate stretching of the sarcomere. Unstructured regions of titin rich in proline extend like molecular springs. Structured Ig domains, on the other hand, unfold at forces of several piconewtons to reveal cryptic residues that can undergo post-translational modification. The I-band of titin is unusually rich in cryptic cysteine residues, which can react with both oxidative and nitrosylative species when exposed by force. Hence, titin is thought to be an important redox sensor in skeletal and cardiac muscle. I propose to study the effects of post-translational modifications on titin elasticity and folding. These studies have important implications for how myocardial mechanics change after myocardial infarction, or in the setting of diseases such as diabetes, hypertension, and atherosclerosis. Aim 1 will study how reactive oxygen species alter the stability of titin Ig domain by blocking folding or inducing disulfide formation. Aim 2 is to determine if reactive nitrogen species react with residues in titin Ig domains alter titin mechanics and how mechanical stability depends on the location of the modification within the Ig fold. Aim 3 seeks to utilize a novel mouse model containing a genetically encoded tag in titin to measure the extent of titin Ig unfolding in muscle tissue using super-resolution and electron microscopy. This unique combination of single molecule and single myofibril experiments will demonstrate how changes at the molecular level translate into a "mechanical phenotype." These studies provide insights into how oxidative insults, such as those present in ischemia/reperfusion tend to affect the cytoskeleton of muscle, altering myocardial mechanics, and initiating signaling pathways that lead to remodeling and further impairment of cardiac function and diastolic performance.

描述（由适用提供）：在填充心脏或相反的肌肉群体的收缩期间，肌节经历被动延伸，将其整体长度增加数百纳米。过去，人们认为细胞外结构（主要是胶原蛋白）是肌膜的完整性的原因，并提供了防止过度伸展肌膜的抵抗力。在过去的三十年中，很明显，肌动蛋白的第三个细丝，在被动拉伸F肌肉组织中具有大部分力，并且负责设定肉瘤的最佳工作长度。仅在2012年，在数百个个体的遗传测序之后，才表明钛中的突变是遗传性扩张性心肌病的主要原因。既然已经存在钛突变与疾病之间的明确联系，那么注意力已转向确定钛的正常生理作用。除了组织厚的细丝外，Titin的I带段变形以适应肌膜的伸展。富含脯氨酸的钛的非结构化区域像分子弹簧一样延伸。另一方面，结构化的IG结构域在几个piconnewtons的力下展开，以揭示可以进行翻译后修饰的密码残差。 Titin的I带异常富含加密半胱氨酸残留物，当通过力暴露时，它们可以与氧化物和亚硝基化物质反应。因此，被认为是骨骼肌和心脏肌肉中的重要氧化还原传感器。我建议研究翻译后修饰对钛弹性和折叠的影响。这些研究对心肌梗死后或糖尿病，高血压和动脉粥样硬化等疾病的情况有重要意义。 AIM 1将通过阻断折叠或诱导的二硫键形成来研究活性氧如何改变钛Ig结构域的稳定性。 AIM 2是确定反应性氮种是否与钛Ig结构域中的残留物反应改变TITIN机械，以及机械稳定性如何取决于IG折叠中修饰的位置。 AIM 3试图利用一种新型的小鼠模型，该模型包含Titin中的一般编码标签，以使用肌肉组织中的TITIN Ig的程度超分辨率和电子显微镜。这种独特的单分子和单肌原纤维实验的组合将证明分子水平的变化如何转化为“机械表型”。这些研究提供了有关氧化物损伤（例如缺血/再灌注中存在的氧化物损伤）倾向于影响肌肉的细胞骨架，改变心肌力学以及启动信号通路，从而导致重塑并进一步损害心脏功能和舒张性表现的信号通路。