Figure 2

miR-130a is a direct target gene of YAP-TEAD. (A) YAP induces miR-130a expression in an S94-dependent manner. pri-miR-130a expression levels in indicated MCF10A stable cells were determined by quantitative RT-PCR. Experiments were performed in triplicates. (B) Knockdown of TEADs reduces miR-130a level. Cells were infected with the shRNA targeting TEAD1/3/4. The levels of mature miR-130a were determined. Experiments were performed in triplicates. (C) YAP represses miR-130a sensor activity in an S94-dependent manner. HepG2 cells were transfected with miR-130a sensor and YAP WT or mutants for luciferase assay. Experiments were performed in duplicates. (D) miR-130a genomic structure. miR-130a promoter is in blue, pri-miR-130a encoding region is in green, and pre-miR-130a encoding region is in red. Regions amplified by ChIP-PCR primers are labeled as P1 to P11. Regions amplified by primers used in RT-PCR confirmation of TSS are labeled as a-e. The proximal promoter cloned for luciferase reporter is indicated. (E, F) YAP and TEAD bind to the proximal promoter of miR-130a. HepG2 cells expressing Flag-YAP-5SA (E) or native HepG2 cells (F) were processed for ChIP with control IgG or antibodies against TEAD1 and YAP followed by quantitative PCR. Experiments were performed in duplicates. (G) YAP activates miR-130a promoter reporter in a TEAD-dependent manner. WT or mutant miR-130a reporters were transfected with or without YAP and TEAD2 into HEK293T cells for luciferase assay. Experiments were performed in duplicates.