Contraction-stimulated glucose uptake in skeletal muscle requires Rac1, but the molecular mechanism of its activation is not fully understood. Treadmill running was applied to induce C57BL/6 mouse hind limb skeletal muscle contraction in vivo and electrical pulse stimulation contracted C2C12 myotube cultures in vitro. The protein levels or activities of AMPK or the Rac1-specific GEF, Tiam1, were manipulated by activators, inhibitors, siRNA-mediated knockdown, and adenovirus-mediated expression. Activated Rac1 was detected by a pull-down assay and immunoblotting. Glucose uptake was measured using the 2-NBD-glucose fluorescent analog. Electrical pulse stimulated contraction or treadmill exercise upregulated the expression of Tiam1 in skeletal muscle in an AMPK-dependent manner. Axin1 siRNA-mediated knockdown diminished AMPK activation and upregulation of Tiam1 protein expression by contraction. Tiam1 siRNA-mediated knockdown diminished contraction-induced Rac1 activation, GLUT4 translocation, and glucose uptake. Contraction increased Tiam1 gene expression and serine phosphorylation of Tiam1 protein via AMPK. These findings suggest Tiam1 is part of an AMPK-Tiam1-Rac1 signaling pathway that mediates contraction-stimulated glucose uptake in skeletal muscle cells and tissue.

中文翻译：

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Tiam1 介导骨骼肌细胞中 Rac1 激活和收缩诱导的葡萄糖摄取

骨骼肌中收缩刺激的葡萄糖摄取需要 Rac1，但其激活的分子机制尚不完全清楚。应用跑步机跑步诱导体内 C57BL/6 小鼠后肢骨骼肌收缩，电脉冲刺激体外收缩 C2C12 肌管培养物。AMPK 或特定于 Rac1 的 GEF Tiam1 的蛋白质水平或活性受激活剂、抑制剂、siRNA 介导的敲低和腺病毒介导的表达的控制。通过下拉测定和免疫印迹检测激活的 Rac1。使用 2-NBD-葡萄糖荧光类似物测量葡萄糖摄取。电脉冲刺激收缩或跑步机运动以 AMPK 依赖性方式上调骨骼肌中 Tiam1 的表达。Axin1 siRNA 介导的敲低通过收缩减少了 AMPK 激活和 Tiam1 蛋白表达的上调。Tiam1 siRNA 介导的组合式减少了收缩诱导的 Rac1 激活、GLUT4 易位和葡萄糖摄取。收缩通过 AMPK 增加 Tiam1 基因表达和 Tiam1 蛋白的丝氨酸磷酸化。这些发现表明 Tiam1 是 AMPK-Tiam1-Rac1 信号通路的一部分，该通路介导骨骼肌细胞和组织中收缩刺激的葡萄糖摄取。