Fig. 6

The YAP/TEAD complex directly binds the promoter of snail resulting in upregulating Snail expression. a A549 cells were transfection with snail or si-snail, respectively. The expressions of Snail, E-cadherin and Vimentin were analyzed by RT-PCR, western blot, qPCR assays. b Consensus binding motif of TEAD1 and the binding sites of TEAD1 on the promotors of snail was predicted using JASPAR database. c, d The activities of different fragments of snail promoter were measured by luciferase reporter gene assays in A549 cells. e The activities of pGL3–200 (− 600~ − 400) of snail were measured by luciferase reporter gene assays in A549 cells with transfection of the indicated concentration Myc-YAP and the times. f Luciferase activities were detected in A549 or H1299 cells with transfected of plasmids carrying a wild-type or mutant promoter of snail in response to overexpressing YAP or knockdown of YAP using the siRNA in A549 and H1299 cells. g, h A549 cells were transfected with YAP or siYAP, respectively. The mRNA and protein levels of YAP and Snail were analyzed by RT-PCR, western blot, qPCR (g) and immunofluorescent staining (h) assays. i Representative IHC staining images from human lung cancer tissues for YAP and Snail (n = 10). (j) Quantitative ChIP analysis demonstrating that knockdown of YAP using the siRNA decreases but overexpressing YAP increases YAP levels within the promoter region of snail in A549 and H1299 cells. k, l RT-PCR, qPCR and western blot assays indicated that YAP increased the expression of Snail in a dose- (k) and time-dependent (l) manner in A549 cells. Results were presented as mean ± SD, and the error bars represent the SD of three independent experiments. *P < 0.05 or **P < 0.01 indicates a significant difference between the indicated groups (two-tailed, unpaired Student’s t-test or one-way analysis of variance). ns, not significant