Role of the cardiac cytoskeleton in mRNA localization and hypertrophy

心脏细胞骨架在 mRNA 定位和肥大中的作用

• 批准号: 10582513
• 负责人: Emily A Scarborough
• 金额: $ 6.72万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-06-01 至 2023-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10582513
• 关键词: Actins; Active Biological Transport; Adult; Affect; Biological; Blood; Calcium; Cardiac; Cardiac Myocytes; Cell physiology; Cells; Cessation of life; Chronic; Complex; Coupled; Cytoskeleton; Data; Dependence; Deposition; Diffusion; Disease; Elements; Exercise; Genetic; Goals; Growth; Health; Health Care Costs; Heart; Heart Diseases; Heart Hypertrophy; Heart failure; Human; Hypertrophy; In Vitro; Individual; Kinetics; Knowledge; Location; Maintenance; Mechanics; Messenger RNA; Microscopy; Microtubule-Associated Proteins; Microtubules; Modeling; Molecular; Motor; Muscle Cells; Myosin ATPase; Neonatal; Oxygen; Pathologic; Pharmacology; Physiological; Play; Post-Translational Protein Processing; Pregnancy; Process; Proteins; Pump; Rattus; Reagent; Regulation; Research; Research Proposals; Resolution; Ribosomal Proteins; Ribosomes; Role; Sarcomeres; Site; Stress; Structure; System; Testing; Therapeutic; Transcript; Translating; Translations; United States; Visualization; Work; base; cell growth; cell type; cost; heart function; hemodynamics; improved; in vivo; in vivo Model; mouse model; new growth; protein degradation; protein expression; protein function; response; spatiotemporal; tool; trafficking

PROJECT SUMMARY In the adult heart, remodeling in response to changing hemodynamic demands occurs primarily through hypertrophy, the addition of new sarcomeres to individual cardiomyocytes. Physiological hypertrophy, triggered by pregnancy or exercise, maintains normal organization of cardiac structure and can improve cardiac function. Pathological hypertrophy, however, often precedes heart failure. This can induce drastic changes to the cardiomyocyte cytoskeleton. However, with the exception of cellular mechanics, the role of the cardiomyocyte cytoskeleton in both health and disease has remained understudied. Recent data suggest that both the actinomyosin and microtubule network may play a role in mRNA and ribosomal localization in cardiomyocytes, though the mechanism remains unknown. In non-muscle cell types, actin-based and microtubule-based directed mRNA transport is well characterized. Additionally, myosin and other sarcomeric mRNAs, ribosomes, and protein degradation machinery appear to localize to the sarcomere in the cardiomyocyte, supporting a model of local translation for sarcomere maintenance and/or de novo formation. But how these new sarcomeres are formed remains particularly unclear, and the dependence of hypertrophy on proper mRNA transport and localization is unknown. The goal of my research proposal is to determine the relationship between mRNA localization and cardiac hypertrophy both in health and disease. My preliminary data establishes that the microtubule network is essential for mRNA localization in the rat cardiomyocyte in vivo and in vitro. To determine the specific mechanism of mRNA transport, I will utilize pharmacological reagents to destabilize the actin network and specifically inhibit motor proteins in isolated adult rat cardiomyocytes to test if mRNA localization changes. I will also use the MS2-MCP system to live track single mRNA transcripts to unambiguously define the mode of mRNA transport. I will leverage molecular biological tools to mislocalize specific sarcomeric transcripts and to visualize sites of new sarcomere deposition in growth-responsive neonatal rat cardiomyocytes to test if mRNA sublocalization is required for cardiac hypertrophy. Finally, I will evaluate if hypertrophy disrupts proper mRNA localization, and if so, dissect the relative contribution of pathophysiological microtubule network changes to this mislocalization using both in vitro and in vivo models of hypertrophy. Taken together, the proposed research will definitively establish the mechanism of mRNA transport and localization in the cardiomyocyte and its function in cardiac hypertrophy.

项目总结 在成人心脏中，对不断变化的血流动力学需求作出反应的重塑主要通过 肥大，单个心肌细胞增加新的肌节。触发的生理性肥厚 通过怀孕或锻炼，维持心脏结构的正常组织，并能改善心脏功能。 然而，病理性肥厚往往先于心力衰竭。这可能会导致对 心肌细胞骨架。然而，除了细胞力学，心肌细胞的作用 细胞骨架在健康和疾病中的作用一直没有得到充分的研究。 最近的数据表明，放线肌球蛋白和微管网络都可能在mRNA和 核糖体在心肌细胞中的定位，尽管其机制尚不清楚。在非肌肉细胞类型中， 基于肌动蛋白和基于微管的信使核糖核酸的定向转运被很好地描述。此外，肌球蛋白和 其他肌节的mRNAs、核糖体和蛋白质降解机制似乎定位于肌节。 在心肌细胞中，支持肌节维持和/或从头开始的局部翻译模型 队形。但是这些新的肌节是如何形成的仍然特别不清楚，以及对 肥大对mRNA的正确转运和定位尚不清楚。我的研究提案的目标是 确定mRNA定位与健康和疾病心肌肥厚的关系。我的 初步数据表明，微管网络对于大鼠的mRNA定位是必不可少的 体内和体外培养的心肌细胞。为了确定mrna转运的具体机制，我将利用 破坏肌动蛋白网络稳定并特异性抑制分离的运动蛋白的药物试剂 检测成年大鼠心肌细胞的mRNA定位是否发生变化。我还将使用MS2-MCP系统进行现场跟踪 单一的信使核糖核酸转录本明确定义信使核糖核酸的运输方式。我会利用分子 错位特定肌节转录本和显示新肌节沉积部位的生物工具 在生长反应敏感的新生大鼠心肌细胞中检测心肌细胞是否需要mRNA亚阻断化 肥大。最后，我将评估肥大是否扰乱了适当的mrna定位，如果是，就解剖 病理生理微管网络改变对这一错误定位的相对贡献 体外和体内肥大模型。综上所述，拟议的研究将最终确立 心肌细胞内信使核糖核酸转运和定位机制及其在心肌肥厚中的作用。