Figure 5

AP-1 directly regulates miR-199a-2 transcription. (a) Quantitative PCR analysis of primary (pri-) and pre-mature (pre-) miR-199a-1 and miR-199a-2 levels in DCA- and TG-treated hepatocytes after 24 h. The mRNA levels were normalized to GAPDH in each sample. (b) Predicted AP-1-binding sites in the miR-199A2 promoter are shown. Reporter vectors containing wild-type or deleted or mutant promoters were constructed, and luciferase activities were detected in hepatocytes after TG, DCA, or DMSO treatment. (c) ChIP analysis showed high AP-1 (c-Jun) enrichment at the MIR199A2 promoter in HL-7702/L02 cells. Enriched DNA was assayed with qRT-PCR and relative enrichment is normalized to control IgG. The positions of the PCR amplicon are labeled according to the predicted AP-1 binding sites. **P<0.01, n=3. (d) Quantitative PCR analysis of pri- or pre-miR-199a2, and mature miR-199a-5p levels in AP-1 inh SP600125-treated hepatocytes after DCA or TG treatment for 24 h. The mRNA levels were normalized to GAPDH in each sample. (e) The relative luciferase activities of promoter reporters in AP-1 inh SP600125-treated hepatocytes after DCA or TG treatment for 24 h. Data are shown as means±S.D. of three independent experiments. **P<0.01