Figure 2.

LINC01535 represses the miR‐214/EZH2 regulatory axis. A, Dual luciferase reporter assay in HeLa cells co‐transfected with luciferase reporter containing EZH2 3’UTR or nothing, LINC01535 overexpression plasmid (pcDNA‐LINC01535) or empty plasmid (pcDNA), and miR‐214 mimics or miR‐NC. Results are displayed as the relative ratio of Firefly luciferase activity to Renilla luciferase activity. B, Dual luciferase reporter assay in HeLa cells co‐transfected with luciferase reporter containing EZH2 3’UTR or nothing and LINC01535 specific shRNA (sh‐LINC01535) or negative control shRNA (sh‐NC). Results are displayed as the relative ratio of Firefly luciferase activity to Renilla luciferase activity. C, EZH2 protein levels in HeLa cells after co‐transfection of pcDNA‐LINC01535 or pcDNA and miR‐214 mimics or miR‐NC. D, EZH2 protein levels in HeLa cells after transfection of sh‐LINC01535 or sh‐NC. E, ChIP assays were performed in HeLa cells using EZH2 specific antibody or non‐specific IgG. The enriched DNA was detected by qRT‐PCR with primers specific for the promoter of miR‐214. F, After transfection of EZH2 overexpression plasmid or empty plasmid into HeLa cells, EZH2 protein expression level and miR‐214 expression level was detected by Western blot and qRT‐PCR, respectively. G, After transfection of EZH2 specific shRNA or negative control shRNA into HeLa cells, EZH2 protein expression level and miR‐214 expression level was detected by Western blot and qRT‐PCR, respectively. H, After co‐transfection of pcDNA‐LINC01535 or pcDNA and sh‐EZH2 or sh‐NC into HeLa cells, miR‐214 expression level was detected by qRT‐PCR. I, After transfection of sh‐LINC01535 or sh‐NC into HeLa cells, miR‐214 expression level was detected by qRT‐PCR. Results are shown as mean ± SD of three independent experiments. **P < 0.01, ***P < 0.001, ns, not significant, by Student's t test (B, E, F, G, I) or one‐way ANOVA followed by Dunnett's multiple comparison test (A, H)