Leucyl-tRNA synthetase is required for the myogenic differentiation of C2C12 myoblasts, but not for hypertrophy or metabolic alteration of myotubes
Leucyl-tRNA synthetase is required for the myogenic differentiation of C2C12 myoblasts, but not for hypertrophy or metabolic alteration of myotubes
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DOI：
10.1016/j.yexcr.2018.02.002
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发表时间：
2018-03-15
影响因子：
3.7
通讯作者：
Yoshizawa, Fumiaki
中科院分区：
文献类型：
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作者：
Sato, Yoriko;Sato, Yusuke;Yoshizawa, Fumiaki
哺乳动物靶标雷帕霉素(MTOR)信号通过感知细胞内的能量状态和营养物质来控制骨骼肌细胞的分化、生长和代谢。最近，亮氨酰-tRNA合成酶(LARS)被发现是参与mTOR信号激活的亮氨酸的胞内感受器。然而，目前还没有证据表明LARS激活mTOR信号及其在骨骼肌细胞中的生理作用。在这项研究中，我们通过小干扰(Si)-RNA敲除来确定LARS在mTOR信号的激活、骨骼肌细胞分化、肥大和代谢中的潜在作用。SiRNA介导的LARS基因敲除降低了磷酸化的p70S6激酶，抑制了C2C12小鼠成肌细胞向肌管的分化，表现为融合指数降低，成肌标志物的mRNA和蛋白表达水平降低。重要的是，si-LARS从分化的早期就降低了胰岛素样生长因子2(IGF2)的mRNA表达水平，表明mTOR-IGF2轴与LARS之间可能存在关联。然而，通过测量肌管直径和检测肥大标志物的mRNA和蛋白表达，LARS基因敲除并没有减少分化的肌管的磷酸化mTOR，也没有影响肌管的肥大。同样，细胞外流量分析仪显示，LARS基因敲除并不影响肌管的新陈代谢(糖酵解和线粒体呼吸)。这些结果表明，LARS是通过激活mTOR信号途径实现骨骼肌分化所必需的，但不是肌管肥大或代谢改变所必需的。
Mammalian target of rapamycin (mTOR) signaling controls skeletal muscle cell differentiation, growth, and metabolism by sensing the intracellular energy status and nutrients. Recently, leucyl-tRNA synthetase (Lars) was identified as an intracellular sensor of leucine involved in the activation of mTOR signaling. However, there is still no evidence for the activation of mTOR signaling by Lars and its physiological roles in skeletal muscle cells. In this study, we determined the potential roles of Lars for the activation of mTOR signaling, skeletal muscle cell differentiation, hypertrophy, and metabolism using small interfering (si)-RNA knockdown. siRNA-mediated knockdown of Lars decreased phosphorylated p70 S6 kinase and inhibited the differentiation of C2C12 mouse myoblasts into myotubes, as evidenced by a decreased fusion index and decreased mRNA and protein expression levels of myogenic markers. Importantly, si-Lars decreased the level of Insulin-like growth factor 2 (Igf2) mRNA expression from the early stages of differentiation, indicating the possibility of an association between the mTOR-IGF2 axis and Lars. However, Lars knockdown did not decrease phosphorylated mTOR in differentiated myotubes, nor did it affect the hypertrophy of myotubes as evidenced by measuring their diameters and detecting the mRNA and protein expression of hypertrophy markers. Similarly, an extracellular flux analyzer showed that Lars knockdown did not affect the metabolism (glycolysis and mitochondrial respiration) of myotubes. These results demonstrate that Lars is required for skeletal muscle differentiation through the activation of mTOR signaling, but not for hypertrophy or metabolic alteration of myotubes.