miRNAs repressed by PML-RAR control crucial cancer genes. (A) Luciferase assay performed in 293T cells transfected with the empty renilla luciferase reporter gene psiCHECK2 (dark histograms) or with the reporter fused to the uPAR 3′ UTR (psiCHECK2–3′ UTR uPAR; gray histograms). The cells were also transfected with the pcDNA3 plasmid or the PML-RARA–expressing vector and treated or not with ATRA (1 μM) for 16 hours as indicated. Results are expressed as RLUs, 1 representing the value obtained with the empty psiCHECK2 plasmid in absence of PML-RARA and treatment. (B) RT-qPCR analyses directed against the RARB mRNA and the miR-195 in 293T cells transfected and treated as in panel A. (C) Luciferase assay performed in 293T cells transfected with the empty renilla luciferase reporter gene psiCHECK2 (dark histograms) or with the reporter fused to the EBF-3 3′ UTR (psiCHECK2-3′ UTR EBF-3; gray histograms). Cells were treated and results analyzed as in panel A. (D) LUC assay performed in 293T cells transfected with the psiCHECK2 or psiCHECK2-3′ UTR uPAR together with specific LNA miRNA inhibitors as indicated. LNA anti–miR-32 was used as a negative control. The results are expressed as RLU, 1 being the value obtained with the empty psiCHECK2 plasmid for each LNA treatment (control).