Lehtinen S K, Rahkila P, Helenius M, Korhonen P, Salminen A
Department of Cell Biology, University of Jyväskylä, Finland.
Biochem Biophys Res Commun. 1996 Dec 4;229(1):36-43. doi: 10.1006/bbrc.1996.1754.
Terminal differentiation of myocytes involves withdrawal from the cell cycle, induction of myogenin expression, and finally formation of myotubes. To study the factors that regulate the initial phase of muscle differentiation, we analyzed the binding activities of transcription factors AP-1, Sp-1, and NF-kappa B in L6, C2C12, and rhabdomyosarcoma BA-Han-1C cells. Temporal changes in transcription factor binding activities were compared to the activation of myogenin promoter-driven CAT reporter gene and the expression level of myogenin, a master gene of myogenic differentiation. We observed a prominent decrease in the nuclear binding activities of AP-1, Sp-1, and NF-kappa B already 12 to 24 h after the transfer of cells to differentiation medium. The response was very similar in L6 and C2C12 myocytes and in BA-Han-1C rhabdomyosarcoma cells. The down-regulation clearly preceded the activation of myogenin promoter and the induction of myogenin and retinoblastoma expression, as well as the initiation of myocyte fusion. Cholera toxin and okadaic acid, established inhibitors of myogenin expression and muscle differentiation, strongly up-regulated the binding activities of AP-1, Sp-1, and NF-kappa B in differentiation medium. Myogenin expression and myocyte fusion were also inhibited. Levels of nuclear c-Fos and c-Jun proteins, components of the AP-1 complex, showed a prominent decrease already after 12 h in differentiation medium. These results show that the down-regulation of the proliferation-promoting transcription factors is a prerequisite to the initiation of myocyte differentiation.
肌细胞的终末分化涉及退出细胞周期、诱导肌细胞生成素表达，最终形成肌管。为了研究调节肌肉分化初始阶段的因素，我们分析了L6、C2C12和横纹肌肉瘤BA-Han-1C细胞中转录因子AP-1、Sp-1和NF-κB的结合活性。将转录因子结合活性的时间变化与肌细胞生成素启动子驱动的CAT报告基因的激活以及肌细胞生成素（肌源性分化的主导基因）的表达水平进行了比较。我们观察到，在将细胞转移到分化培养基后12至24小时，AP-1、Sp-1和NF-κB的核结合活性就显著降低。在L6和C2C12肌细胞以及BA-Han-1C横纹肌肉瘤细胞中，反应非常相似。这种下调明显先于肌细胞生成素启动子的激活、肌细胞生成素和成视网膜细胞瘤表达的诱导以及肌细胞融合的启动。霍乱毒素和冈田酸是已确定的肌细胞生成素表达和肌肉分化抑制剂，它们在分化培养基中强烈上调AP-1、Sp-1和NF-κB的结合活性。肌细胞生成素的表达和肌细胞融合也受到抑制。AP-1复合物的组成成分核c-Fos和c-Jun蛋白水平在分化培养基中培养12小时后就显著下降。这些结果表明，促进增殖的转录因子的下调是肌细胞分化启动的前提条件。
