Myosin phosphorylation in skeletal muscle

描述(由申请人提供)： 肌球蛋白轻链激酶(MLCK)催化钙/钙调素依赖性磷酸化 在马达蛋白的调节轻链(RLC)，肌球蛋白II。有两个基因 MLCKs一种是在包括骨骼肌纤维在内的所有细胞中表达平滑肌MLCKs， 而另一种仅在成人骨骼肌纤维中表达明显的骨骼肌MLCK。平滑肌MLCK可磷酸化平滑和非肌肉RLC，但不能磷酸化骨骼肌RLC。然而，骨骼肌MLCK很容易使所有的RLC磷酸化。在骨骼肌中，钙离子激活收缩是通过肌钙蛋白-原肌球蛋白这一细丝调节系统来调节的。最近的报道表明，钙/钙调蛋白的形成是低的。 相对于细胞内总钙调素，可能限制了多个靶点的激活 蛋白质。在完整肌肉中的实验表明，RLC磷酸化和 强直后快速抽动骨骼肌等长力幅值或Treppe的增强。 RLC磷酸化-作用力关系的相对重要性尚不清楚，因为 结皮纤维中力量发展的速度和程度的增加以及其他因素 影响完整肌肉的收缩性能，包括长度依赖效应 肌浆网钙离子释放、肌丝间距和钙调蛋白 钙释放通道RyR1的调节。本申请中描述的研究将 测定(1)游离钙浓度与钙/钙调素的关系 在C2C12肌管和酶分散的小鼠骨骼肌纤维中形成 生物传感器分子，(2)结合钙/钙调素的时空分布 和骨骼肌纤维中独特的生物传感器骨骼肌和平滑肌MLCKs的激活 来自转基因动物，以及(3)如果RLC磷酸化和增加的力幅度 消融骨骼肌对小鼠快速抽动骨骼肌纤维的抑制作用 MLCK基因。这些研究的结果将提供对钙依赖的洞察 在运动中激发的增强肌肉性能的机制。

DESCRIPTION (provided by applicant): Myosin light chain kinase (MLCK) catalyzes the Ca2+/calmodulin-dependent phosphorylation of the regulatory light chain (RLC) of the motor protein, myosin II. There are two genes for MLCKs. One expresses smooth muscle MLCKs in all cells, including skeletal muscle fibers, whereas the other expresses a distinct skeletal muscle MLCK only in adult skeletal muscle fibers. Smooth muscle MLCK phosphorylates smooth and nonmuscle RLC but not skeletal muscle RLC. However, skeletal muscle MLCK readily phosphorylates all RLCs. In skeletal muscle Ca2+ activation of contraction is mediated through a thin filament regulatory system, troponin-tropomyosin. Recent reports indicate that Ca2+/calmodulin formation is low relative to the total cellular calmodulin and may be limiting for activation of multiple target proteins. Experiments in intact muscles show a correlation between RLC phosphorylation and post-tetanic potentiation of isometric force amplitude or treppe in fast-twitch skeletal muscles. The relative importance of RLC phosphorylation-force relationship is unclear due to modest increases in the rate and extent of force development in skinned fibers and to other factors affecting contractile performance in intact muscles including length-dependent effects on Ca2+ release from the sarcoplasmic reticulum, inter-myofilament spacing and calmodulin regulation of Ryr1, the calcium release channel. The research described in this application will determine (1) the relationship between the free Ca2+ concentration and Ca2+/ calmodulin formation in C2C12 myotubes and enzymatically dispersed mouse skeletal muscle fibers with biosensor molecules, (2) the temporal and spatial distributions of Ca2+/calmodulin binding to and activation of unique biosensor skeletal and smooth muscle MLCKs in skeletal muscle fibers from transgenic animals, and (3) if RLC phosphorylation and increased force amplitude are inhibited in fast-twitch skeletal muscle fibers from mice with ablation of the skeletal muscle MLCK gene. The results from these investigations will provide insights into Ca2+-dependent mechanisms recruited during exercise that enhance muscle performance.