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. 2015 Mar 30;6(13):10964–10977. doi: 10.18632/oncotarget.3696

Figure 5. miR-942 directly targets sFRP4, GSK3β, and TLE1.

Figure 5

(A) Indicated cells transfected with TOP flash/FOP flash and Renilla pRL-TK plasmids were subjected to dual-luciferase assays 48 hours after transfection. Reporter activity detected was normalized by Renilla luciferase activity. (B) Predicted miR-942 target sequences in the 3′-UTRs of sFRP4, GSK3β and TLE1 and mutant containing three altered nucleotides in the seed sequence of miR-942 (miR-942-mu). (C) Western blotting analysis of sFRP4, GSK3β, TLE1 and nuclear β-catenin in the indicated cells. α-Tubulin served as the loading control. (D) Luciferase activity of targets 3′UTR in the indicated cells. (E) miRNP immunoprecipitation assay showed associations of miR-942 with sFRP4, GSK3β and TLE1. GAPDH and 5s rRNA served as a negative control. (F) Quantification of indicated tumoursphere cells with specific siRNA transfection. (G) Luciferase activity of TOP flash/FOP flash in the indicated cells. Each bar represents the mean ± SD of three independent experiments. * P < 0.05.