Regulatory light chain phosphorylation augments length-dependent contraction in PTU-treated rats
Regulatory light chain phosphorylation augments length-dependent contraction in PTU-treated rats
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DOI：
10.1085/jgp.201812158
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发表时间：
2019-01-01
影响因子：
3.8
通讯作者：
Tanner, Bertrand C. W.
中科院分区：
文献类型：
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作者：
Breithaupt, Jason J.;Pulcastro, Hannah C.;Tanner, Bertrand C. W.
肌动蛋白-肌球蛋白跨桥的力量产生受整个心跳中的细丝蛋白和肌节长度(SL)的调节。先前的工作表明，肌球蛋白调节紧链(RLC)结合在肌球蛋白重链的颈部，当磷酸化时，可以增加心脏的收缩能力。我们最近使用主要由较快的α-心肌肌球蛋白重链亚型组成的皮肤大鼠心肌条显示，跨桥动力学随着SLS的增加而保持，并且RLC的磷酸化放大了这一效应。在本研究中，为了评估RLC磷酸化如何随着肌球蛋白运动速度的变化而影响长度依赖的肌球蛋白功能，我们使用丙基硫氧嘧啶(PTU)饮食诱导大鼠心室中较慢的β-肌球蛋白重链亚型表达>；95%。通过随机长度扰动分析，我们测量了RLC磷酸化对1.9和2.2微米长皮肤大鼠乳头肌条上钙激活的等长收缩和肌球蛋白跨桥动力学的影响。最大张力和钙敏感性随着SL的增加而增加，而RLC的磷酸化在2.2µm SL时增强了这种反应。在2.2µm SL时，RLC的磷酸化可使粘弹性心肌硬度略有增加，但在1.91µm SL时则不明显，因此提示RLC的磷酸化增加了较长SLS时的β-肌球蛋白重链结合或僵硬。随着SL的增加，跨桥分离速度减慢，这为延长跨桥附着提供了一种潜在的机制，以增强较长SLS下依赖长度的收缩激活。RLC磷酸化不影响β-肌球蛋白重链脱落率的长度依赖性减慢。结合我们之前的研究，这些数据表明，随着SL的增加，α和β-肌球蛋白重链亚型都显示出长度依赖的激活反应和肌球蛋白附着的延长，而RLC磷酸化在较长的SLS增加了长度依赖的激活。然而，在比较心脏亚型时，我们发现β-肌球蛋白重链始终表现出比α-肌球蛋白重链更大的长度依赖性敏感性。我们的工作表明，在两种心肌肌球蛋白重链亚型中，RLC磷酸化是调节心肌收缩能力的重要因素。
Force production by actin-myosin cross-bridges in cardiac muscle is regulated by thin-filament proteins and sarcomere length (SL) throughout the heartbeat. Prior work has shown that myosin regulatory tight chain (RLC), which binds to the neck of myosin heavy chain, increases cardiac contractility when phosphorylated. We recently showed that cross-bridge kinetics stow with increasing SLs, and that RLC phosphorylation amplifies this effect, using skinned rat myocardial strips predominantly composed of the faster alpha-cardiac myosin heavy chain isoform. In the present study, to assess how RLC phosphorylation influences length-dependent myosin function as myosin motor speed varies, we used a propylthiouracil (PTU) diet to induce >95% expression of the slower beta-myosin heavy chain isoform in rat cardiac ventricles. We measured the effect of RLC phosphorylation on Ca2+-activated isometric contraction and myosin cross-bridge kinetics (via stochastic length perturbation analysis) in skinned rat papillary muscle strips at 1.9- and 2.2-mu m SL. Maximum tension and Ca2+ sensitivity increased with SL, and RLC phosphorylation augmented this response at 2.2-mu m SL. Subtle increases in viscoelastic myocardial stiffness occurred with RLC phosphorylation at 2.2-mu m SL, but not at 1.91 mu m SL, thereby suggesting that RLC phosphorylation increases beta-myosin heavy chain binding or stiffness at longer SLs. The cross-bridge detachment rate slowed as SL increased, providing a potential mechanism for prolonged cross-bridge attachment to augment length-dependent activation of contraction at longer SLs. Length-dependent slowing of beta-myosin heavy chain detachment rate was not affected by RLC phosphorylation. Together with our previous studies, these data suggest that both alpha- and beta-myosin heavy chain isoforms show a length-dependent activation response and prolonged myosin attachment as SL increases in rat myocardial strips, and that RLC phosphorylation augments length-dependent activation at longer SLs. In comparing cardiac isoforms, however, we found that beta-myosin heavy chain consistently showed greater length-dependent sensitivity than alpha-myosin heavy chain. Our work suggests that RLC phosphorylation is a vital contributor to the regulation of myocardial contractility in both cardiac myosin heavy chain isoforms.