A new post-translational modification, citrullination, changes in heart failure

一种新的翻译后修饰，瓜氨酸化，心力衰竭的变化

• 批准号: 8256288
• 负责人: Jennifer E Van Eyk
• 金额: $ 24.3万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-03-01 至 2014-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8256288
• 关键词: Actins; Affect; Amino Acids; Area; Arginine; Arginine deiminase; Autoimmune Process; Binding; Cardiac; Cardiac Myocytes; Cardiomyopathies; Chronic; Comb animal structure; Contractile Proteins; Data; Dilated Cardiomyopathy; Enzymes; Functional disorder; Glutamine; Grant; Heart; Heart failure; Human; Immune System Diseases; Inflammatory; Lead; Light; Mass Spectrum Analysis; Messenger RNA; Methods; Microfilaments; Modeling; Modification; Molecular Biology; Mus; Muscle; Muscle Cells; Myocardial; Myocardial Contraction; Myocardial dysfunction; Myocarditis; Myocardium; Myosin ATPase; Pathogenesis; Patients; Physiological; Play; Post-Translational Protein Processing; Posttranslational Amino Acid Modification; Protein Isoforms; Protein-arginine deiminase; Proteins; Regulation; Role; System; Testing; Thin Filament; Time; Tropomyosin; Troponin; Viral; Work; base; constriction; innovation; interest; mimetics; mouse model; mutant; myosin-binding protein C; novel; pressure; programs

DESCRIPTION (provided by applicant): Protein citrullination (deimination), a post-translational modification (PTM) altering the amino acid residue arginine (Arg), has recently become an area of interest due its emerging role in human auto-immune diseases. We are the first group to establish that this PTM is present in the human heart. Specifically, our preliminary data shows citrullination occurs in myofilament proteins such as tropomyosin (Tm), myosin (heavy and light chain) and myosin binding protein C. Furthermore, we find an increase in the extent of deiminated proteins in myocardium obtained from patients with heart failure (HF) compared to control hearts. Based on these results, we hypothesize that citrullination plays a role in the pathogenesis of HF, particularly affecting myocardial contraction. Furthermore, we hypothesize that citrullination is exacerbated in HF especially when there is an autoimmune component. This exploratory grant is aimed to establish the relation between protein citrullination, peptidyl arginine deiminase (PAD), the enzyme responsible for this PTM, and cardiac and myocardial dysfunction in the failing heart. In Specific Aim 1, PAD isoform expression and citrullinated myocardial proteins are explored in three mouse models of HF, chronic aortic constriction and two inflammatory dilated cardiomyopathies. Using a recently developed mass spectrometry-based method, citrullinated proteins and the modified amino acid residues will be identified and quantified. In addition, using a combination of molecular biology and a second innovative mass spectroscopy approach, we will unambiguously define and quantify the PAD isoforms in cardiac myocytes. Specific aim 2 focuses on the assessment of the functional role of PAD and protein citrullination in cardiac myocytes. Combing molecular biology and pharmacological methods to manipulate PAD isoform activity, the extent of modified proteins is correlated to myocardial contractile function. Tm mutants with Ala (non modifiable) or Gln (a potential mimetic of Arg-Cit) replacing the modifiable citrullinatable Arg residues, will be tested for functional alterations contraction. This program is highly novel as protein citrullination and PAD system has never been explored in the heart, yet our new evidence supports its potential to modulate muscle contractility. This work will lead to understanding of an entirely new mechanism for post-translational protein regulation in the heart. PUBLIC HEALTH RELEVANCE: For the first time, we show that protein citrullination (deimination), a post-translational modification (altering the amino acid residue arginine (Arg) occurs in the heart, targeting a number of contractile proteins, and that it increases with heart failure. This exploratory R21 proposal aims to establish the relation between protein citrullination, peptidyl arginine deiminase (PAD), the enzyme responsible for this PTM, and myocardial contractile dysfunction in three different models of HF. This highly novel program will lead to understanding of an entirely new mechanism for post-translational protein regulation in the heart.

描述（由申请人提供）：瓜氨酸蛋白（脱亚胺化）是一种改变氨基酸残基精氨酸（Arg）的翻译后修饰（PTM），由于其在人类自身免疫疾病中的新兴作用，最近已成为关注的领域。我们是第一个确定这种PTM存在于人类心脏中的小组。具体而言，我们的初步数据显示瓜氨酸存在于肌丝蛋白中，如原肌球蛋白（Tm）、肌球蛋白（重链和轻链）和肌球蛋白结合蛋白C。此外，我们发现与对照心脏相比，从心力衰竭（HF）患者获得的心肌中脱亚胺蛋白的程度增加。基于这些结果，我们假设瓜氨酸在HF的发病机制中起作用，特别是影响心肌收缩。此外，我们假设瓜氨酸在HF中加重，特别是当存在自身免疫成分时。这项探索性资助旨在确定瓜氨酸蛋白、肽基精氨酸脱亚胺酶（PAD）（导致PTM的酶）与衰竭心脏的心脏和心肌功能障碍之间的关系。在具体目标1中，在HF、慢性主动脉缩窄和两种炎性扩张性心肌病的三种小鼠模型中探索了PAD同种型表达和瓜氨酸化心肌蛋白。使用最近开发的基于质谱的方法，瓜氨酸化蛋白和修饰的氨基酸残基将被识别和定量。此外，使用分子生物学和第二个创新的质谱方法相结合，我们将明确定义和量化心肌细胞中的PAD亚型。具体目标2侧重于评估PAD和瓜氨酸蛋白在心肌细胞中的功能作用。结合分子生物学和药理学方法来操纵PAD异构体活性，修饰蛋白的程度与心肌收缩功能相关。用Ala（不可修饰）或Gln（Arg-Cit的潜在模拟物）替代可修饰的瓜氨酸化Arg残基的Tm突变体将测试功能改变收缩。该计划非常新颖，因为瓜氨酸蛋白和PAD系统从未在心脏中进行过探索，但我们的新证据支持其调节肌肉收缩力的潜力。这项工作将导致对心脏中翻译后蛋白质调节的全新机制的理解。 公共卫生相关性：我们首次发现，瓜氨酸蛋白（脱亚胺化）是一种翻译后修饰（改变精氨酸（Arg）的氨基酸残基），发生在心脏中，靶向许多收缩蛋白，并且随着心力衰竭而增加。这项探索性R21提案旨在确定蛋白质瓜氨酸、肽基精氨酸脱亚胺酶（PAD）（导致PTM的酶）与三种不同HF模型中心肌收缩功能障碍之间的关系。这个高度新颖的程序将导致对心脏中翻译后蛋白质调节的全新机制的理解。