Regulation of myosin II dynamics by phosphorylation and dephosphorylation of its light chain in epithelial cells
Regulation of myosin II dynamics by phosphorylation and dephosphorylation of its light chain in epithelial cells
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DOI：
10.1091/mbc.e06-07-0590
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发表时间：
2007-02-01
影响因子：
3.3
通讯作者：
Yonemura, Shigenobu
中科院分区：
文献类型：
--
作者：
Watanabe, Toshiyuki;Hosoya, Hiroshi;Yonemura, Shigenobu
非肌肉肌球蛋白II是一种以肌动蛋白为基础的运动蛋白，在肌动蛋白的细胞骨架组织和细胞运动中起重要作用。虽然已知其调节轻链(MRLC)的磷酸化在体外参与肌球蛋白II细丝的组装和运动活动，但目前尚不清楚MRLC磷酸化如何在体内调节肌球蛋白II的动力学。我们建立了表达MRLC增强绿色荧光蛋白或其突变体的Madin Darby犬肾脏II上皮细胞克隆。时间推移成像显示，肌球蛋白II的正常动力学需要磷酸化和去磷酸化。影响肌球蛋白磷酸化和MRLC突变体的抑制剂表明，MRLC的单磷酸化是必需的，并且足以维持应力纤维。二磷酸化的MRLC稳定了肌球蛋白II的细丝，并局部分布在发生收缩的应力纤维区域，表明二磷酸化参与了肌球蛋白II的组装和收缩的空间调节。我们进一步发现，肌球蛋白磷酸酶或拉链相互作用蛋白激酶根据肌球蛋白II ATPase的活性定位于应激纤维。
Nonmuscle myosin II, an actin-based motor protein, plays an essential role in actin cytoskeleton organization and cellular motility. Although phosphorylation of its regulatory light chain (MRLC) is known to be involved in myosin II filament assembly and motor activity in vitro, it remains unclear exactly how MRLC phosphorylation regulates myosin II dynamics in vivo. We established clones of Madin Darby canine kidney II epithelial cells expressing MRLC-enhanced green fluorescent protein or its mutants. Time-lapse imaging revealed that both phosphorylation and dephosphorylation are required for proper dynamics of myosin II. Inhibitors affecting myosin phosphorylation and MRLC mutants indicated that monophosphorylation of MRLC is required and sufficient for maintenance of stress fibers. Diphosphorylated MRLC stabilized myosin II filaments and was distributed locally in regions of stress fibers where contraction occurs, suggesting that diphosphorylation is involved in the spatial regulation of myosin II assembly and contraction. We further found that myosin phosphatase or Zipper-interacting protein kinase localizes to stress fibers depending on the activity of myosin II ATPase.