Ait-Mou Younss, Hsu Karen, Farman Gerrie P, Kumar Mohit, Greaser Marion L, Irving Thomas C, de Tombe Pieter P
Department of Cell and Molecular Physiology, Loyola University Chicago, Stritch School of Medicine, Maywood, IL 60153;
Department of Cell and Molecular Physiology, Loyola University Chicago, Stritch School of Medicine, Maywood, IL 60153; Department of Biological and Chemical Sciences, Illinois Institute of Technology, Chicago, IL 60616;
Proc Natl Acad Sci U S A. 2016 Feb 23;113(8):2306-11. doi: 10.1073/pnas.1516732113. Epub 2016 Feb 8.
The Frank-Starling mechanism of the heart is due, in part, to modulation of myofilament Ca(2+) sensitivity by sarcomere length (SL) [length-dependent activation (LDA)]. The molecular mechanism(s) that underlie LDA are unknown. Recent evidence has implicated the giant protein titin in this cellular process, possibly by positioning the myosin head closer to actin. To clarify the role of titin strain in LDA, we isolated myocardium from either WT or homozygous mutant (HM) rats that express a giant splice isoform of titin, and subjected the muscles to stretch from 2.0 to 2.4 μm of SL. Upon stretch, HM compared with WT muscles displayed reduced passive force, twitch force, and myofilament LDA. Time-resolved small-angle X-ray diffraction measurements of WT twitching muscles during diastole revealed stretch-induced increases in the intensity of myosin (M2 and M6) and troponin (Tn3) reflections, as well as a reduction in cross-bridge radial spacing. Independent fluorescent probe analyses in relaxed permeabilized myocytes corroborated these findings. X-ray electron density reconstruction revealed increased mass/ordering in both thick and thin filaments. The SL-dependent changes in structure observed in WT myocardium were absent in HM myocardium. Overall, our results reveal a correlation between titin strain and the Frank-Starling mechanism. The molecular basis underlying this phenomenon appears not to involve interfilament spacing or movement of myosin toward actin but, rather, sarcomere stretch-induced simultaneous structural rearrangements within both thin and thick filaments that correlate with titin strain and myofilament LDA.
心脏的Frank-Starling机制部分归因于肌节长度（SL）对肌丝Ca(2+)敏感性的调节[长度依赖性激活（LDA）]。LDA背后的分子机制尚不清楚。最近的证据表明巨蛋白肌联蛋白参与了这一细胞过程，可能是通过使肌球蛋白头部更靠近肌动蛋白来实现的。为了阐明肌联蛋白应变在LDA中的作用，我们从表达肌联蛋白巨大剪接异构体的野生型（WT）或纯合突变型（HM）大鼠中分离出心肌，并将肌肉拉伸至2.0至2.4μm的肌节长度。拉伸后，与WT肌肉相比，HM肌肉的被动力、收缩力和肌丝LDA降低。对舒张期WT抽搐肌肉进行时间分辨小角X射线衍射测量，结果显示拉伸诱导肌球蛋白（M2和M6）和肌钙蛋白（Tn3）反射强度增加，以及横桥径向间距减小。在松弛的透化心肌细胞中进行的独立荧光探针分析证实了这些发现。X射线电子密度重建显示粗肌丝和细肌丝的质量/有序性均增加。在HM心肌中未观察到WT心肌中所观察到的与肌节长度相关的结构变化。总体而言，我们的结果揭示了肌联蛋白应变与Frank-Starling机制之间的相关性。这一现象背后的分子基础似乎不涉及丝间间距或肌球蛋白向肌动蛋白的移动，而是肌节拉伸诱导的细肌丝和粗肌丝内同时发生的结构重排，这些重排与肌联蛋白应变和肌丝LDA相关。
