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. 2015 Mar 8;6(10):7866–7879. doi: 10.18632/oncotarget.3492

Fig.5. LRP6, TRIM29 and Pygo2 are direct targets of miR-432.

Fig.5

(A). Indicated cells transfected with TOPflash or FOPflash 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). Altered nuclear translocation of β-catenin in response to deregulated miR-432 expression. Nuclear fractions of indicated cells were analyzed by western blotting; p84 was used as the loading control. (C). Predicted miR-432 target sequences in the 3′UTRs of LRP6, TRIM29 and Pygo2. The miR-432 mutant (miR-432-mut) contained three altered nucleotides in the seed sequence. (D). Western blotting analysis of LRP6, TRIM29 and Pygo2 expression in the indicated HCC cells. α-Tubulin served as loading control. (E). Luciferase assay of pGL3- LRP6-3′UTR, pGL3- TRIM29-3′UTR and pGL3- Pygo2-3′UTR reporter cotransfected with miR-432, miR-432-inhibitor and miR-432-mut oligonucleotides in HCC cells. (F). Luciferase assay of TCF/LEF transcriptional activity in indicated cells transfected with specific siRNA. Error bars represent the mean ± SD from of three independent experiments. *P < 0.05.