miR-1 induces G1 phase arrest and inhibits myoblast proliferation by direct targeting Cyclin D1.

<p>(A) Identification of miR-1 regulatory element in the 3′UTR of mouse Cyclin D1. (B) Evaluation of effect of miR-1 mimics and miR-133 mimics on reporters containing three different regions of Cyclin D1–3′UTR in C2C12 myoblasts transfected with miR-1 or miR-133 mimics as indicated. Error bars represent the SD of three independent experiments. (C) Western blot analysis of Cyclin D1 protein expression in C2C12 myoblasts transfected with miR-1 mimics or control oligos. (D) Cell cycle analysis of C2C12 myoblasts transfected with miR-1 mimics or control oligos. Data shown are from a typical experiment performed. (E) Growth curves of C2C12 myoblasts were determined by MTT assay. Cells were transfected with miR-1 mimics or control oligos. Error bars represent the SD of three independent experiments. (F and G) Proliferation of C2C12 myoblasts was evaluated by BrdU incorporation. Cells were transfected with miR-1 mimics. Representative images of cells were taken by fluorescence microscope (F). The percentage of BrdU positive cells was measured (G). Data shown are from a typical experiment performed in triplicate. **p<0.01.