Stac3 Is Required for Myotube Formation and Myogenic Differentiation in Vertebrate Skeletal Muscle
Stac3 Is Required for Myotube Formation and Myogenic Differentiation in Vertebrate Skeletal Muscle
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DOI：
10.1074/jbc.m112.361311
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发表时间：
2012-12-21
影响因子：
4.8
通讯作者：
Johnston, Ian A.
中科院分区：
文献类型：
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作者：
Bower, Neil I.;de la Serrana, Daniel Garcia;Johnston, Ian A.
从大西洋鲑鱼消减杂交文库中筛选到一个在骨骼肌中表达量最高的营养调控基因Stac 3。鲑鱼Stac 3 mRNA与鲑鱼原代生肌培养物分化过程中的肌细胞生成素和myoD 1a表达高度相关，并受氨基酸可用性的调节。在斑马鱼胚胎中，stac 3最初在肌节近轴细胞和分割后的快肌纤维中表达。吗啉敲低导致肌原纤维蛋白组装缺陷，特别是在慢肌纤维中，并降低了刺猬受体补丁的水平。使用哺乳动物C2 C12肌源性细胞系在体外进一步表征Stac 3的功能。在C2 C12肌源性细胞系的分化过程中，Stac 3 mRNA表达增加。通过RNAi敲低Stac 3抑制肌管形成，并且微阵列分析显示参与细胞周期、粘着斑、细胞骨架以及促肌生成因子Igfbp-5和Igf 2的转录物下调。RNAi处理的细胞抑制Akt信号传导，外源性胰岛素样生长因子（Igf）2不能挽救表型，然而，Igf/Akt信号传导没有被阻断。Stac 3的过表达导致Igfbp-5 mRNA水平的增加，但并不导致分化的增加。在同步化细胞中，Stac 3 mRNA在细胞周期的G（1）期最丰富。与对照组相比，RNAi处理的细胞更小，具有更高的增殖率和减少的G（1）期细胞比例，表明在G（1）期检查点中的作用。这些结果确定Stac 3作为一个新的基因所需的成肌分化和肌原纤维蛋白组装在脊椎动物。
Stac3 was identified as a nutritionally regulated gene from an Atlantic salmon subtractive hybridization library with highest expression in skeletal muscle. Salmon Stac3 mRNA was highly correlated with myogenin and myoD1a expression during differentiation of a salmon primary myogenic culture and was regulated by amino acid availability. In zebrafish embryos, stac3 was initially expressed in myotomal adaxial cells and in fast muscle fibers post-segmentation. Morpholino knockdown resulted in defects in myofibrillar protein assembly, particularly in slow muscle fibers, and decreased levels of the hedgehog receptor patched. The function of Stac3 was further characterized in vitro using the mammalian C2C12 myogenic cell line. Stac3 mRNA expression increased during the differentiation of the C2C12 myogenic cell line. Knock-down of Stac3 by RNAi inhibited myotube formation, and microarray analysis revealed that transcripts involved in cell cycle, focal adhesion, cytoskeleton, and the pro-myogenic factors Igfbp-5 and Igf2 were down-regulated. RNAi-treated cells had suppressed Akt signaling and exogenous insulin-like growth factor (Igf) 2 was unable to rescue the phenotype, however, Igf/Akt signaling was not blocked. Overexpression of Stac3, which results in increased levels of Igfbp-5 mRNA, did not lead to increased differentiation. In synchronized cells, Stac3 mRNA was most abundant during the G(1) phase of the cell cycle. RNAi-treated cells were smaller, had higher proliferation rates and a decreased proportion of cells in G(1) phase when compared with controls, suggesting a role in the G(1) phase checkpoint. These results identify Stac3 as a new gene required for myogenic differentiation and myofibrillar protein assembly in vertebrates.