miR-550a downregulates <i>CPEB4</i> expression by directly targeting its 3′UTR.

<p>(<b>A</b>) Scheme for the identification of candidate genes combining microarray assays and a target prediction algorithm. (<b>B</b>) The putative binding sequences for miR-550a within the human (Has), chimpanzee (Ptr), rhesus (MMl) and mouse (Mmu) <i>CPEB4</i> 3′UTR. Seed sequences are indicated as emphasized with shadow. (<b>C</b>) The sequences of the putative miR-550a-binding site in the wild-type (emphasized with shadow) and mutant (underlined) <i>CPEB4</i> 3′UTR. (<b>D</b>) The relative luciferase activity of luciferase reporters with wild-type or mutant <i>CPEB4</i> 3′UTRs was determined in HEK 293T cells, which were co-transfected with the miR-550a mimic or negative control (nc). Renilla luciferase activity was analyzed as an internal control. There is statistical significance between the wild-type group and the mutant group with miR-550a transfection. Representative experiments are shown with the mean ± SD. (<b>E</b>) The protein levels of <i>CPEB4</i> were determined by western blot assays in Huh-7 cells infected with miR-550a or mock vector and Huh-7 cells transfected with the miR-550a inhibitor or negative control (nc).