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. 2021 Oct 7;11:710525. doi: 10.3389/fonc.2021.710525

Figure 4.

Figure 4

miR-183-5p mediates ZNF304-regulated FOXO4 expression in ccRCC. (A) Potential microRNAs targeted 3′UTR of FOXO4 were predicted in three online target-prediction programs, and showed with venn diagram. (B) Biotin pull-down assay was used to enrich miRNAs, and RT-qPCR was used to detect these candidate miRNAs. (C) 786-0 cells were transfected with oeZNF304 or shZNF304 or control vectors. Candidate miRNAs were detected by using RT-qPCR. U6 served as a negative control. (D) 786-0 cells were co-transfected with the FOXO4 3′UTR and miR-183-5p or miR-150-5p mimics. Luciferase reporter assays showed that miR-183-5p reduced FOXO4 3′UTR luciferase activity. (E) The prediction of miR-183-5p binding site in the FOXO4 3′UTR. (F, G) 786-0 and SW839 cells were transfected with the indicated miRNAs, and then FOXO4 protein expression was measured by western blotting. (H) 786-0 and SW839 cells were co-transfected with the indicated miRNAs and vectors, and cell viability was measured by MTT. All data are from three independent experiments and expressed as the mean ± SEM. *P < 0.05, **P < 0.01 vs. the corresponding controls.