Fig. 5. TWIST1 ceRNAs regulate TWIST1 expression in LUAD cell lines.

a, b Double-label staining showed that TWIST1 expression was significantly increased after transfection with SLC12A5-3′UTR (a)/ZFHX4-3′UTR (b), and decreased after transfection with sh-SLC12A5/ZFHX4 in H1299 cells. c Western blot for TWIST1 protein levels (upper) and quantitation of TWIST1 protein and mRNA levels (lower) in H1299 cells transfected with miR-194-3p or miR-514a-3p inhibitors as well as sh-TWIST1/sh-SLC12A5/sh-ZFHX4. d Western blot for TWIST1 protein levels (upper) and quantitation of TWIST1 protein and mRNA levels (lower) in A549 cells transfected with miR-194-3p or miR-514a-3p mimics as well as TWIST1-3′UTR/ SLC12A5-3′UTR / ZFHX4-3′UTR regions. e Luciferase activity in H1299 cells co-transfected with shRNA against TWIST1 ceRNAs as well as miR-194-3p or miR-514a-3p inhibitors and a luciferase-TWIST1 3′UTR reporter construct. f, g Luciferase activity in H1299 cells co-transfected with 3′UTR regions of SLC12A5 (f) and ZFHX4 (g) as well as miR-194-3p or miR-514a-3p mimics and a luciferase-TWIST1 3′UTR reporter construct. All data are mean ± s.d. The data statistical significances were assessed by Student’s t-test compared to the NC group. *P < 0.05, **P < 0.001