Figure 4.

Effect of miR-212 on Wnt/β-catenin signaling pathway activity. A, Luciferase reporter gene evaluating β-catenin Transcription factor/Lymphoid enhancer-binding factor (TCF/LEF) promoter activity using TOPflash and FOPflash vectors. The binding site of 3-TCF were localized in the TOPflash reporter vector, and the mutated binding sites of TCF were localized in TOPflash. As indicated, HCCLM3 cells were cotransfected with different expression vectors. MiR-212 treatment reduced β-catenin TCF/LEF promoter activity, whereas anti-miR-212 treatment increased β-catenin TCF/LEF promoter activity. B, Immunofluorescence assay for β-catenin indicated that β-catenin translocated from the nucleus to the cytoplasm upon increased expression of miR-212. When cells were transfected with anti-miR-212, the localization of β-catenin in the cells shifted from the cytoplasm to the nucleus, compared with miR-con. *P < .05. C, Western blot of Axin2, LEF-1, and c-Myc in miR-212-transduced HCCLM3 cells or anti-miR-212-transduced HCCLM3 cells. Evaluation of the expression levels of β-catenin in the cell nuclei of miR-212- and anti-miR-212-transduced HCCLM3 cells. GAPDH, control for cytoplasmic protein; Lamin B, control for nuclear protein. HCC indicates hepatocellular carcinoma.