Pennsylvania State University College of Medicine, Hershey, PA.
University of Kentucky, Lexington, KY.
J Gen Physiol. 2021 Jul 5;153(7). doi: 10.1085/jgp.202012811.
Mutations in the cardiac myosin regulatory light chain (RLC, MYL2 gene) are known to cause inherited cardiomyopathies with variable phenotypes. In this study, we investigated the impact of a mutation in the RLC (K104E) that is associated with hypertrophic cardiomyopathy (HCM). Previously in a mouse model of K104E, older animals were found to develop cardiac hypertrophy, fibrosis, and diastolic dysfunction, suggesting a slow development of HCM. However, variable penetrance of the mutation in human populations suggests that the impact of K104E may be subtle. Therefore, we generated human cardiac myosin subfragment-1 (M2β-S1) and exchanged on either the wild type (WT) or K104E human ventricular RLC in order to assess the impact of the mutation on the mechanochemical properties of cardiac myosin. The maximum actin-activated ATPase activity and actin sliding velocities in the in vitro motility assay were similar in M2β-S1 WT and K104E, as were the detachment kinetic parameters, including the rate of ATP-induced dissociation and the ADP release rate constant. We also examined the mechanical performance of α-cardiac myosin extracted from transgenic (Tg) mice expressing human wild type RLC (Tg WT) or mutant RLC (Tg K104E). We found that α-cardiac myosin from Tg K104E animals demonstrated enhanced actin sliding velocities in the motility assay compared with its Tg WT counterpart. Furthermore, the degree of incorporation of the mutant RLC into α-cardiac myosin in the transgenic animals was significantly reduced compared with wild type. Therefore, we conclude that the impact of the K104E mutation depends on either the length or the isoform of the myosin heavy chain backbone and that the mutation may disrupt RLC interactions with the myosin lever arm domain.
心脏肌球蛋白调节轻链（RLC，MYL2 基因）中的突变已知会导致具有不同表型的遗传性心肌病。在这项研究中，我们研究了与肥厚型心肌病（HCM）相关的 RLC（K104E）突变的影响。以前在 K104E 的小鼠模型中，发现老年动物会发生心肌肥厚、纤维化和舒张功能障碍，表明 HCM 的发展缓慢。然而，该突变在人类群体中的可变外显率表明 K104E 的影响可能很细微。因此，我们生成了人心肌肌球蛋白亚片段-1（M2β-S1），并在野生型（WT）或 K104E 人心室 RLC 上进行了交换，以评估该突变对心肌肌球蛋白的机械化学性质的影响。在体外运动分析中，M2β-S1 WT 和 K104E 的最大肌动蛋白激活 ATP 酶活性和肌动蛋白滑动速度相似，脱离动力学参数也相似，包括 ATP 诱导的解离速率和 ADP 释放速率常数。我们还检查了从表达人野生型 RLC（Tg WT）或突变型 RLC（Tg K104E）的转基因（Tg）小鼠中提取的α-心肌肌球蛋白的机械性能。我们发现，与 Tg WT 相比，Tg K104E 动物的α-心肌肌球蛋白在运动分析中具有更高的肌动蛋白滑动速度。此外，与野生型相比，突变型 RLC 整合到转基因动物中的α-心肌肌球蛋白的程度显著降低。因此，我们得出结论，K104E 突变的影响取决于肌球蛋白重链骨干的长度或同工型，并且该突变可能破坏 RLC 与肌球蛋白杠杆臂结构域的相互作用。
