Figure 5.

FZD3 Is a Target of miR-182-5p

(A) Schematic of the candidate genes of miR-182-5p using four prediction tools. (B) GO and KEGG pathway analysis of candidate targets of miR-182-5p. (C) Schematic of the common candidate genes involved in regulating the Wnt/β-catenin signaling pathway. (D) mRNA expression of PPP3R1, FZD3, and PPP3CA in OS cells transfected with miR-NC or a miR-182-5p mimic. (E) Predicted miR-182-5p target sequences in the 3′ UTRs of FZD3 genes. (F) Relative FZD3 reporter activities in OS cells co-transfected with miR-182-5p and luciferase reporters. (G) Immunoprecipitation of the Ago2/RISC (RNA-induced silencing complex) using the Pan-Ago2 antibody in OS cells overexpressing miR-NC or miR-182-5p. IgG was used as a negative control and β-actin was used as an internal control. (H) Quantitative real-time PCR analysis of miR-182-5p incorporated into RISC in OS cells overexpressing miR-182-5p compared with the levels in the control. (I) Quantitative real-time PCR of FZD3 incorporated into RISC in OS cells overexpressing miR-182-5p. (J) FZD3 expression levels in OS cells transfected with a miR-182-5p mimic or anti-miR-182-5p were quantified by western blot. In all experiments, bars represent means ± SD from three independent experiments. ∗p < 0.05, ∗∗p < 0.01.