Cellular miR-181b directly targets MEV NS1 mRNA coding region.

<p>(<b>A</b>) Schematic layout of the MEV genome DNA, NS1 mRNA and presumptive target of miR-181b via the 9 sequential complementary nucleotides as indicated. The target and tetranucleotide mutant target segment was cloned into pGL3-control reporter vector. (<b>B</b>) Luciferase activity of lysates of F81 cells after 36 h co-transfection with pGL3-181t, pRL-TK and miR-181b or miR-181c mimics or inhibitors. NC mimics and inhibitors were used as controls. (<b>C</b>) As in <b>B)</b>, except that mut pGL3-181t, mut-1 miR-181b mimics and inhibitors were used for transfection. (<b>D</b>) Luciferase activity of lysates of F81 cells infected with MEV at an MOI of 0.1 or transfected with pcDNA-NS1 (0.1 µg/well), and co-transfected with reporter constructs (0.1 µg/well) and miRNA mimics (10 pmol/well) in 24-well plates. Data are from 3 independent experiments (mean ± SD). Statistical significance was analyzed by Student’s <i>t</i> test; *<i>P</i><0.05; **<i>P</i><0.01; ***<i>P</i><0.001; ns, not significant.