miR-195-5p directly targets Wnt3a
TargetScan, miRanda and miRBase were used to predict the potential targets of miR-195-5p. Among the candidate target genes, Wnt3a had a miR-195-5p-binding site in their 3′UTR (Fig. 4A). After up-regulation or down-regulation of miR-195-5p level in cells, the protein expression of Wnt3a was decreased or increased correspondingly (P < 0.05) (Fig. 4B-C). In order to test whether miR-195-5p directly targets Wnt3a, dual luciferase reporter assay was constructed, which had either the Wnt3a 3′UTR-wt or the Wnt3a 3′UTR-mut (Fig. 4A). The miR-195-5p mimics inhibited the luciferase reporter activity of the Wnt3a 3′UTR-wt by 19% (P < 0.05), however, the activity was markedly increased with the Wnt3a 3′UTR-mut and miR-195-5p mimics co-transfection (P < 0.05) (Fig. 4D).