Department of Molecular Physiology and Biophysics, Cardiovascular Research Institute, University of Vermont, Burlington, VT.
Division of Cell Biology and Imaging, Department of Radiology, University of Massachusetts Medical School, Worcester, MA.
J Gen Physiol. 2021 Mar 1;153(3). doi: 10.1085/jgp.202012726.
Myosin and actin filaments are highly organized within muscle sarcomeres. Myosin-binding protein C (MyBP-C) is a flexible, rod-like protein located within the C-zone of the sarcomere. The C-terminal domain of MyBP-C is tethered to the myosin filament backbone, and the N-terminal domains are postulated to interact with actin and/or the myosin head to modulate filament sliding. To define where the N-terminal domains of MyBP-C are localized in the sarcomere of active and relaxed mouse myocardium, the relative positions of the N terminus of MyBP-C and actin were imaged in fixed muscle samples using super-resolution fluorescence microscopy. The resolution of the imaging was enhanced by particle averaging. The images demonstrate that the position of the N terminus of MyBP-C is biased toward the actin filaments in both active and relaxed muscle preparations. Comparison of the experimental images with images generated in silico, accounting for known binding partner interactions, suggests that the N-terminal domains of MyBP-C may bind to actin and possibly the myosin head but only when the myosin head is in the proximity of an actin filament. These physiologically relevant images help define the molecular mechanism by which the N-terminal domains of MyBP-C may search for, and capture, molecular binding partners to tune cardiac contractility.
肌球蛋白和肌动蛋白丝在肌肉肌节内高度组织化。肌球蛋白结合蛋白 C（MyBP-C）是一种位于肌节 C 带的灵活的杆状蛋白。MyBP-C 的 C 端结构域与肌球蛋白丝主干相连，而 N 端结构域则被假设与肌动蛋白和/或肌球蛋白头部相互作用，以调节丝滑动。为了确定 MyBP-C 的 N 端结构域在活跃和松弛的小鼠心肌肌节中的定位，使用超分辨率荧光显微镜在固定的肌肉样本中对 MyBP-C 的 N 端和肌动蛋白的相对位置进行成像。通过粒子平均提高了成像的分辨率。这些图像表明，在活跃和松弛的肌肉标本中，MyBP-C 的 N 端位置偏向肌动蛋白丝。将实验图像与考虑已知结合伴侣相互作用的计算图像进行比较表明，MyBP-C 的 N 端结构域可能与肌动蛋白结合，并且可能与肌球蛋白头部结合，但只有当肌球蛋白头部靠近肌动蛋白丝时才会结合。这些与生理相关的图像有助于定义 MyBP-C 的 N 端结构域可能寻找和捕获分子结合伴侣以调节心脏收缩性的分子机制。
