Faenza Irene, Bavelloni Alberto, Fiume Roberta, Lattanzi Giovanna, Maraldi Nadir M, Gilmour R Stewart, Martelli Alberto M, Suh Pann-Ghill, Billi Anna Maria, Cocco Lucio
Cellular Signalling Laboratory, Department of Anatomical Science, University of Bologna, Italy.
J Cell Physiol. 2003 Jun;195(3):446-52. doi: 10.1002/jcp.10264.
Phospholipase C beta(1) (PLCbeta(1)) signaling in both cell proliferation and differentiation has been largely investigated, but its role in myoblast differentiation is still unclear. The C2C12 myogenic cell line has been used in this study in order to find out the role of the two subtypes of PLCbeta(1), i.e., a and b in this process. C2C12 myoblast proliferate in response to mitogens and upon mitogen withdrawal differentiates into multinucleated myotubes. We found that differentiation of C2C12 skeletal muscle cells is characterized by a marked increase in the amount of nuclear PLCbeta(1)a and PLCbeta(1)b. Indeed, treatment with insulin induces a dramatic rise of both PLCbeta(1) subtypes expression and activity, as determined by immunochemical and enzymatic assays. Immunofluorescence experiments with anti-PLCbeta(1) specific monoclonal antibody showed a low level of cytoplasmatic and nuclear staining during the initial 12 h of differentiation whilst a massive nuclear staining is appreciable in differentiating cells. The time course of PLCbeta(1) expression versus Troponin T expression clearly indicates that the increase in the amount of PLCbeta(1) takes place 24 h earlier than that of Troponin T. Moreover, the overexpression of the PLCbeta(1)M2b mutant, lacking the nuclear localization signal and entirely located in the cytoplasm, represses the formation of mature multinucleated myotube. Taken together these results suggest that nuclear PLCbeta(1) is a key player in myoblast differentiation, functioning as a positive regulator of this process.
磷脂酶Cβ1（PLCβ1）在细胞增殖和分化中的信号传导已得到广泛研究，但其在成肌细胞分化中的作用仍不清楚。本研究使用C2C12成肌细胞系，以探究PLCβ1的两种亚型，即α和β在此过程中的作用。C2C12成肌细胞在有丝分裂原的刺激下增殖，去除有丝分裂原后分化为多核肌管。我们发现，C2C12骨骼肌细胞的分化特征是核内PLCβ1α和PLCβ1β的量显著增加。事实上，胰岛素处理可诱导两种PLCβ1亚型的表达和活性急剧上升，这通过免疫化学和酶学分析得以确定。用抗PLCβ1特异性单克隆抗体进行的免疫荧光实验表明，在分化开始的12小时内，细胞质和细胞核染色水平较低，而在分化细胞中可观察到大量的细胞核染色。PLCβ1表达与肌钙蛋白T表达的时间进程清楚地表明，PLCβ1量的增加比肌钙蛋白T早24小时发生。此外，缺乏核定位信号且完全位于细胞质中的PLCβ1 M2b突变体的过表达抑制了成熟多核肌管的形成。综上所述，这些结果表明核内PLCβ1是成肌细胞分化的关键参与者，在此过程中起正调节作用。
