Taniguchi M, Ishikawa H
J Cell Biol. 1982 Feb;92(2):324-32. doi: 10.1083/jcb.92.2.324.
We studied the in situ reconstitution of myosin filaments within the myosin-extracted myofibrils in cultured chick embryo skeletal muscle cells using the electron microscope and polarization microscope. Myosin was first extracted from the myofibrils in glycerinated muscle cells with a high-salt solution containing 0.6 M KCl. When rabbit skeletal muscle myosin was added to the myosin-extracted cells in the high-salt solution, thin filaments in the ghost myofibrils were bound with myosin to form arrowhead complexes. Subsequent dilution of KCl in the myosin solution to 0.1 M resulted in the formation of thick myosin filaments within the myofibrils, increasing the birefringence of the myofibrils. When Mg-ATP was added such myosin-reassembled myofibrils were induced either to form supercontraction bands or to restore the sarcomeric arrangement of thick and thin filaments. Under the polarization microscope, vibrational movement of the myofibrils was seen transiently upon addition of Mg-ATP, often resulting in a regular arrangement of myofibrils in register. These myofibrils, with reconstituted myosin filaments, structurally and functionally resembled the native myofibrils. The findings are discussed with special reference to the myofibril formation in developing muscle cells.
我们利用电子显微镜和偏振显微镜，研究了培养的鸡胚骨骼肌细胞中肌球蛋白提取的肌原纤维内肌球蛋白丝的原位重组。首先用含有0.6M KCl的高盐溶液从甘油化肌肉细胞的肌原纤维中提取肌球蛋白。当将兔骨骼肌肌球蛋白添加到高盐溶液中已提取肌球蛋白的细胞时，空壳肌原纤维中的细肌丝与肌球蛋白结合形成箭头复合物。随后将肌球蛋白溶液中的KCl稀释至0.1M，导致肌原纤维内形成粗肌球蛋白丝，增加了肌原纤维的双折射。当添加Mg-ATP时，这种肌球蛋白重组的肌原纤维被诱导形成超收缩带或恢复粗细肌丝的肌节排列。在偏振显微镜下，添加Mg-ATP后可短暂观察到肌原纤维的振动运动，这通常会导致肌原纤维规则排列对齐。这些具有重组肌球蛋白丝的肌原纤维在结构和功能上与天然肌原纤维相似。我们结合发育中肌肉细胞的肌原纤维形成对这些发现进行了讨论。
