Rui Cai, Naren Qimuge, Meilin Ma, Yingqian Wang, Guorong Tang, Que Zhang, Yunmei Sun, Xiaochang Chen, Taiyong Yu, Wuzi Dong, Gongshe Yang, Weijun Pang
The Journal of biological chemistry 2018 Dec 14MicroRNAs (miRNAs) are noncoding RNAs that regulate gene expression at the post-transcriptional level and are involved in the regulation of the formation, maintenance, and function of skeletal muscle. Using miRNA sequencing and bioinformatics analysis, we previously found that the miRNA miR-664-5p is significantly differentially expressed in longissimus dorsi muscles of Rongchang pigs. However, the molecular mechanism by which miR-664-5p regulates myogenesis remains unclear. In this study, using flow cytometry, 5-ethynyl-2'-deoxyuridine staining, and cell count and immunofluorescent assays, we found that cell-transfected miR-664-5p mimics greatly promoted proliferation of C2C12 mouse myoblasts by increasing the proportion of cells in the S- and G2-phases and up-regulating the expression of cell cycle genes. Moreover, miR-664-5p inhibited myoblast differentiation by down-regulating myogenic gene expression. In contrast, miR-664-5p inhibitor repressed myoblast proliferation and promoted myoblast differentiation. Mechanistically, using dual-luciferase reporter gene experiments, we demonstrated that miR-664-5p directly targets the 3'-UTR of serum response factor (SRF) and Wnt1 mRNAs. We also observed that miR-664-5p inhibits both mRNA and protein levels of SRF and Wnt1 during myoblast proliferation and myogenic differentiation, respectively. Furthermore, the activating effect of miR-664-5p on myoblast proliferation was attenuated by SRF overexpression, and miR-664-5p repressed myogenic differentiation by diminishing the accumulation of nuclear β-catenin. Of note, miR-664-5p's inhibitory effect on myogenic differentiation was abrogated by treatment with Wnt1 protein, the key activator of the Wnt/β-catenin signaling pathway. Collectively, our findings suggest that miR-664-5p controls SRF and canonical Wnt/β-catenin signaling pathways in myogenesis. © 2018 Cai et al.
Rui Cai, Naren Qimuge, Meilin Ma, Yingqian Wang, Guorong Tang, Que Zhang, Yunmei Sun, Xiaochang Chen, Taiyong Yu, Wuzi Dong, Gongshe Yang, Weijun Pang. MicroRNA-664-5p promotes myoblast proliferation and inhibits myoblast differentiation by targeting serum response factor and Wnt1. The Journal of biological chemistry. 2018 Dec 14;293(50):19177-19190
PMID: 30323063
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