Figure 6: YAP regulates DNp63 expression through ZEB2. | Nature Communications

Figure 6: YAP regulates DNp63 expression through ZEB2.

From: YAP inhibits squamous transdifferentiation of Lkb1-deficient lung adenocarcinoma through ZEB2-dependent DNp63 repression

Figure 6

(a) Knockdown of TEADs reversed the decrease of DNp63 mRNA and protein levels caused by YAP 5SA ectopic expression in HTB-182 cells. (b) YAP S127A ectopic expression upregulated ZEB2 mRNA and protein levels in HTB-182 cells. (c) YAP RNA interference (RNAi) downregulated ZEB2 mRNA and protein levels in HTB-182 cells. (d) Ectopic expression of YAP 5SA but not YAP 5SA with C-terminal transactivation domain deletion significantly increased ZEB2 mRNA and protein levels in HTB-182 cells. (e) YAP S127A ectopic expression significantly increased the transcriptional activity of ZEB2 wild-type promoter but not mutant with deletion of TEADs-binding site in 293T cells. (f) ChIP assay confirmed YAP binding to ZEB2 promoter in A549 cells. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) served as negative control. (g) ZEB2 ectopic expression downregulated DNp63 mRNA and protein levels in HTB-182 cells. (h) ZEB2 RNAi upregulated DNp63 mRNA and protein levels in HTB-182 cells. (i) ZEB2 overexpression decreased the transcriptional activity of DNp63 wild-type promoter but not mutant with deletion of ZEB2-binding site in L78 cells. (j) ChIP assay confirmed ZEB2 binding to DNp63 promoter in FLAG-ZEB2-transfected HTB-182 and A549 cells. GAPDH served as negative control. (k,l) Knockdown of ZEB2 reversed the decrease of DNp63 mRNA (k) and protein (l) levels caused by YAP ectopic expression in HTB-182 cells. (m,n) Re-introduction of ZEB2 abolished the increase of DNp63 mRNA (m) and protein (n) levels resulted from YAP knockdown in HTB-182 cells. (o) Re-introduction of ZEB2 abolished the increase of DNp63 protein levels caused by YAP 5SA-ΔC ectopic expression in HTB-182 cells. The asterisk represents the YAP 5SA-ΔC mutant form with its C-terminal transactivation domain deletion. Each sample was assayed in triplicates by quantitative real-time PCR and reporter gene assay. Representative data of real-time PCR, western blotting, ChIP and reporter gene assay were shown from three independent experiments. Data are shown as mean±s.e.m. t-test, *P<0.05; **P<0.01; ***P<0.005.

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