Role of phosphorylation in cardiac muscle myosin

磷酸化在心肌肌球蛋白中的作用

• 批准号: 8746718
• 负责人: James Sellers
• 金额: $ 23.63万
• 依托单位: NATIONAL HEART, LUNG, AND BLOOD INSTITUTE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8746718
• 关键词: Address; Biological Assay; Cardiac; Cardiac Myosins; Goals; Heart; Heart Diseases; Human; In Vitro; Isometric Exercise; Left; Light; Measurement; Mechanics; Muscle; Myocardial; Myocardium; Myosin Light Chain Kinase; Myosin Regulatory Light Chains; Output; Performance; Phosphorylation; Physiological; Play; Property; Protein Kinase; Rattus; Rattus norvegicus; Recombinants; Role; Sampling; Series; Speed; Up-Regulation; Ventricular; Work; cell motility; non-muscle myosin; optical traps

Understanding how cardiac myosin regulatory light chain (RLC) phosphorylation alters cardiac muscle mechanics is important because it is often altered in cardiac disease. The effect this protein phosphorylation has on muscle mechanics during a physiological range of shortening velocities, during which the heart generates power and performs work, has not been addressed. We have expressed and phosphorylated recombinant Rattus norvegicus left ventricular RLC. In vitro we have phosphorylated these recombinant species with cardiac myosin light chain kinase and zipper-interacting protein kinase. We compare rat permeabilized cardiac trabeculae, which have undergone exchange with differently phosphorylated RLC species. We were able to enrich trabecular RLC phosphorylation by 40% compared with controls and, in a separate series, lower RLC phosphorylation to 60% of control values. Compared with the trabeculae with a low level of RLC phosphorylation, RLC phosphorylation enrichment increased isometric force by more than 3-fold and peak power output by more than 7-fold and approximately doubled both maximum shortening speed and the shortening velocity that generated peak power. We augmented these measurements by observing increased RLC phosphorylation of human and rat HF samples from endocardial left ventricular homogenate. These results demonstrate the importance of increased RLC phosphorylation in the up-regulation of myocardial performance and suggest that reduced RLC phosphorylation is a key aspect of impaired contractile function in the diseased myocardium. We are currently using optical trapping and in vitro motility assays to explore the effect of phosphorylation on purified cardiac muscle myosin.

了解心肌肌球蛋白调节轻链(RLC)磷酸化如何改变心肌力学很重要，因为它经常在心脏病中改变。在心脏产生能量和做功的生理速度范围内，这种蛋白磷酸化对肌肉力学的影响尚未得到解决。我们表达并磷酸化了重组褐家鼠左心室RLC。在体外，我们用心肌肌球蛋白轻链激酶和拉链相互作用蛋白激酶对这些重组物种进行了磷酸化。我们比较了大鼠的通透性心脏小梁，它们与不同磷酸化的RLC物种进行了交换。与对照组相比，我们能够使小梁RLC的磷酸化增加40%，而在单独的系列中，RLC的磷酸化降低到控制值的60%。与低水平RLC磷酸化的骨小梁相比，RLC磷酸化浓缩使等长力增加了3倍以上，峰值功率输出增加了7倍以上，最大缩短速度和产生峰值功率的缩短速度都增加了近一倍。我们通过观察来自心内膜左心室匀浆的人和大鼠心内膜心内膜匀浆中的人和大鼠HF样本的RLC磷酸化增加来扩大这些测量。这些结果证明了RLC磷酸化增加在上调心肌功能中的重要性，并提示RLC磷酸化减少是病变心肌收缩功能受损的一个关键方面。 我们目前正在使用光学捕获和体外运动分析来探索磷酸化对纯化的心肌肌球蛋白的影响。