Pulcastro Hannah C, Awinda Peter O, Methawasin Mei, Granzier Henk, Dong Wenji, Tanner Bertrand C W
Department of Integrative Physiology and Neuroscience, Washington State University Pullman, WA, USA.
Department of Cellular and Molecular Medicine, University of Arizona Tucson, AZ, USA.
Front Physiol. 2016 Jul 29;7:322. doi: 10.3389/fphys.2016.00322. eCollection 2016.
Titin is a giant protein spanning from the Z-disk to the M-band of the cardiac sarcomere. In the I-band titin acts as a molecular spring, contributing to passive mechanical characteristics of the myocardium throughout a heartbeat. RNA Binding Motif Protein 20 (RBM20) is required for normal titin splicing, and its absence or altered function leads to greater expression of a very large, more compliant N2BA titin isoform in Rbm20 homozygous mice (Rbm20 (ΔRRM) ) compared to wild-type mice (WT) that almost exclusively express the stiffer N2B titin isoform. Prior studies using Rbm20 (ΔRRM) animals have shown that increased titin compliance compromises muscle ultrastructure and attenuates the Frank-Starling relationship. Although previous computational simulations of muscle contraction suggested that increasing compliance of the sarcomere slows the rate of tension development and prolongs cross-bridge attachment, none of the reported effects of Rbm20 (ΔRRM) on myocardial function have been attributed to changes in cross-bridge cycling kinetics. To test the relationship between increased sarcomere compliance and cross-bridge kinetics, we used stochastic length-perturbation analysis in Ca(2+)-activated, skinned papillary muscle strips from Rbm20 (ΔRRM) and WT mice. We found increasing titin compliance depressed maximal tension, decreased Ca(2+)-sensitivity of the tension-pCa relationship, and slowed myosin detachment rate in myocardium from Rbm20 (ΔRRM) vs. WT mice. As sarcomere length increased from 1.9 to 2.2 μm, length-dependent activation of contraction was eliminated in the Rbm20 (ΔRRM) myocardium, even though myosin MgADP release rate decreased ~20% to prolong strong cross-bridge binding at longer sarcomere length. These data suggest that increasing N2BA expression may alter cardiac performance in a length-dependent manner, showing greater deficits in tension production and slower cross-bridge kinetics at longer sarcomere length. This study also supports the idea that passive mechanical characteristics of the myocardium influence ensemble cross-bridge behavior and maintenance of tension generation throughout the sarcomere.
肌联蛋白是一种巨大的蛋白质，横跨心脏肌节的Z盘到M带。在I带中，肌联蛋白充当分子弹簧，在整个心跳过程中对心肌的被动机械特性起作用。正常的肌联蛋白剪接需要RNA结合基序蛋白20（RBM20），与几乎只表达更硬的N2B肌联蛋白异构体的野生型小鼠（WT）相比，Rbm20纯合小鼠（Rbm20（ΔRRM））中其缺失或功能改变会导致非常大且更具柔韧性的N2BA肌联蛋白异构体表达增加。先前使用Rbm20（ΔRRM）动物的研究表明，肌联蛋白柔韧性增加会损害肌肉超微结构并减弱Frank-Starling关系。尽管先前对肌肉收缩的计算模拟表明，肌节柔韧性增加会减慢张力发展速率并延长横桥附着时间，但Rbm20（ΔRRM）对心肌功能的所有已报道影响均未归因于横桥循环动力学的变化。为了测试肌节柔韧性增加与横桥动力学之间的关系，我们在来自Rbm20（ΔRRM）和WT小鼠的Ca（2+）激活的、去表皮乳头肌条中使用了随机长度扰动分析。我们发现，与WT小鼠相比，Rbm20（ΔRRM）小鼠心肌中肌联蛋白柔韧性增加会降低最大张力、降低张力-pCa关系的Ca（2+）敏感性，并减慢肌球蛋白解离速率。当肌节长度从1.9μm增加到2.2μm时，Rbm20（ΔRRM）心肌中长度依赖性的收缩激活被消除，尽管肌球蛋白MgADP释放速率降低了约20%，以在更长的肌节长度下延长强横桥结合。这些数据表明，增加N2BA表达可能以长度依赖性方式改变心脏性能，在更长的肌节长度下，张力产生的缺陷更大，横桥动力学更慢。这项研究还支持这样一种观点，即心肌的被动机械特性会影响整体横桥行为以及整个肌节中张力产生的维持。
