Figure 4

MicroRNA specific activities. (A) MiR-335 negatively regulates luciferase expression in a plasmid coupling its coding sequence with FOXP3 3′UTR. Renilla luciferase reporter assays with constructs holding FOXP3 3′-UTR sequences from the indicated genes were co-transfected into HEK293T cells along with a firefly luciferase transfection control plasmid either alone or together with miR-335. (B) MiR-9 and miR-155 negatively regulate luciferase expression in a plasmid coupling its coding sequence with CTLA-4 3′UTR. Renilla luciferase reporter assays with constructs holding CTLA-4 3′-UTR sequences, wild type or miR-site deleted, were co-transfected into HEK293T cells along with a firefly luciferase transfection control plasmid either alone or together with miR-9, −155. MiR-24 (C) and miR-335 (D) specifically targets GARP 3′UTR and negatively regulate luciferase reporter expression. Renilla and firefly luciferase reporter assays with constructs holding GARP 3′-UTR sequences, wild type or miR-site deleted, were co-transfected into HEK293T cells along with miR-24, −335. Relative luciferase values normalized to transfections without miRNA are shown. Data represent mean ± SD (error bars) of three independent experiments, each performed in triplicate. (*p < 0.05; **, p < 0.01, Student's t test).