Calcineurin activates interleukin-6 transcription in mouse skeletal muscle in vivo and in C2C12 myotubes in vitro
Calcineurin activates interleukin-6 transcription in mouse skeletal muscle in vivo and in C2C12 myotubes in vitro
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DOI：
10.1152/ajpregu.00325.2009
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发表时间：
2010-01-01
影响因子：
2.8
通讯作者：
Reed, Jason M.
中科院分区：
文献类型：
--
作者：
Allen, David L.;Uyenishi, Jill J.;Reed, Jason M.
艾伦DL，Uyenishi JJ，Cleary AS，Mehan RS，林赛SF，里德JM。钙调神经磷酸酶激活白细胞介素-6在小鼠骨骼肌在体内和C2 C12肌管在体外转录。Am J Physiol Regul Integr Comp Physiol 298：R198-R210，2010年。首次发表于2009年11月11日; doi：10.1152/ajpregu.00325.2009。骨骼肌细胞因子白细胞介素-6（IL-6）的表达在一次耐力运动后会大幅增加，这似乎是由细胞内钙离子的增加介导的。我们研究了耐力运动对体内骨骼肌IL-6 mRNA水平和启动子活性的影响，以及钙激活的钙调神经磷酸酶信号通路对体内和体外肌肉IL-6表达的作用。小鼠胫骨前肌（TA）中的IL-6 mRNA水平通过单次跑步机运动或3天自愿轮跑增加2-10倍。此外，IL-6启动子驱动的荧光素酶转基因激活TA跑步机和车轮运行的运动和注射钙调磷酸酶质粒。运动也增加了钙调磷酸酶调节基因MCIP 1的肌肉mRNA表达，如用钙离子载体A23187治疗C2 C12肌管。C2 C12肌管与组成型活性钙调磷酸酶构建体的共转染显著增加，而与钙调磷酸酶抑制剂CAIN共转染抑制小鼠IL-6启动子-报告基因构建体的活性。与肌细胞增强因子-2（MEF-2）表达构建体共转染增加了基础IL-6启动子活性，增强了钙调神经磷酸酶共转染的效果，而与MEF-2拮抗剂MITR共转染抑制了钙调神经磷酸酶激活的IL-6启动子活性。令人惊讶的是，与另一种钙调神经磷酸酶激活的转录因子，T细胞核因子激活因子（NFAT）的显性阴性形式的共转染，大大增强了C2 C12肌管中的基础和钙调神经磷酸酶刺激的IL-6启动子活性。MEF-2 DNA结合位点的突变减弱，而NFAT DNA结合位点的突变增强基础和钙调神经磷酸酶激活的IL-6启动子活性。最后，CREB和C/EBP是基础IL-6启动子活性所必需的，足以增加IL-6启动子活性，但在钙调神经磷酸酶激活的IL-6启动子活性中的作用最小。总之，这些结果表明，骨骼肌细胞中的IL-6转录可以被NFAT拮抗的钙调神经磷酸酶-MEF-2轴激活。
Allen DL, Uyenishi JJ, Cleary AS, Mehan RS, Lindsay SF, Reed JM. Calcineurin activates interleukin-6 transcription in mouse skeletal muscle in vivo and in C2C12 myotubes in vitro. Am J Physiol Regul Integr Comp Physiol 298: R198-R210, 2010. First published November 11, 2009; doi:10.1152/ajpregu.00325.2009.-Expression of the cytokine interleukin-6 (IL-6) by skeletal muscle is hugely increased in response to a single bout of endurance exercise, and this appears to be mediated by increases in intracellular calcium. We examined the effects of endurance exercise on IL-6 mRNA levels and promoter activity in skeletal muscle in vivo, and the role of the calcium-activated calcineurin signaling pathway on muscle IL-6 expression in vivo and in vitro. IL-6 mRNA levels in the mouse tibialis anterior (TA) were increased 2-10-fold by a single bout of treadmill exercise or by 3 days of voluntary wheel running. Moreover, an IL-6 promoter-driven luciferase transgene was activated in TA by both treadmill and wheel-running exercise and by injection with a calcineurin plasmid. Exercise also increased muscle mRNA expression of the calcineurin regulatory gene MCIP1, as did treatment of C2C12 myotubes with the calcium ionophore A23187. Cotransfection of C2C12 myotubes with a constitutively active calcineurin construct significantly increased while cotransfection with the calcineurin inhibitor CAIN inhibited activity of a mouse IL-6 promoter-reporter construct. Cotransfection with a myocyte enhancer-factor-2 (MEF-2) expression construct increased basal IL-6 promoter activity and augmented the effects of calcineurin cotransfection, while cotransfection with the MEF-2 antagonist MITR repressed calcineurin-activated IL-6 promoter activity in vitro. Surprisingly, cotransfection with a dominant-negative form of another calcineurin-activated transcription factor, nuclear factor activator of T cells (NFAT), greatly potentiated both basal and calcineurin-stimulated IL-6 promoter activity in C2C12 myotubes. Mutation of the MEF-2 DNA binding sites attenuated, while mutation of the NFAT DNA binding sites potentiated basal and calcineurin-activated IL-6 promoter activity. Finally, CREB and C/EBP were necessary for basal IL-6 promoter activity and sufficient to increase IL-6 promoter activity but had minimal roles in calcineurin-activated IL-6 promoter activity. Together, these results suggest that IL-6 transcription in skeletal muscle cells can be activated by a calcineurin-MEF-2 axis which is antagonized by NFAT.