IRF-5 is a target of miR-302a.
A, predicted miR-302a–binding sites in the 3′-UTR of the IRF-5 mRNA. Perfect matches in seed regions are indicated by lines. Mutations (underlined) were generated in the binding sites in the 3′-UTR for the reporter gene assay. B, A549 cells were transfected with the WT or the indicated mutants of the IRF-5 3′-UTR reporter plasmid and miR-302a for 48 h before luciferase assays. C, experiments were performed as described in B, except that cells were transfected with the miR-302a inhibitor or inhibitor controls. D, THP-1 cells were transfected with the miR-302a mimic or mimic controls for 24 h before real-time RT-PCR (top) and Western blot (bottom) analyses. E, experiments were performed as described in D, except that a nonspecific inhibitor control or miR-302a inhibitor was used. In the real-time RT-PCR experiments, the control was designated as 1. Bar graphs present means ± S.D. (error bars), n = 3 (**, p < 0.01; *, p < 0.05; n.s., not significant).