Fig. 1
ET-1/ETAR axis induces the ZEB1/YAP interaction in HG-SOC cells. A, B Immunoblotting (IB) analysis of ZEB1, pYAP (S127), YAP, pTAZ (S89) and TAZ protein expression in the cytoplasmic and nuclear extracts of patient-derived HG-SOC PMOV10 (A) and OVCAR-3 (B) cells stimulated with ET-1 (100 nM) for the indicated times. Tubulin and PCNA were used as cytoplasmic and nuclear loading control, respectively. C Nuclear extracts of PMOV10 cells stimulated for 6 h with ET-1 and/or with macitentan (MAC, 1µM), a dual ET-1 receptor antagonist, were IB for ZEB1, YAP, and TAZ. PCNA was used loading control. D Nuclear extracts of PMOV10 cells stimulated for 6 h with ET-1 and/or with the selective ETAR antagonist BQ123 (1µM), the selective ETBR antagonist BQ788 (1µM), or macitentan (1µM), were IB for ZEB1, YAP and TAZ. PCNA was used loading control. E Nuclear extracts of PMOV10 cells stimulated as in C were immunoprecipitated (IP) for endogenous ZEB1 using anti-ZEB1 antibody (Ab) or anti-immunoglobulin G (IgG) Ab as control and IB using anti-ZEB1 and anti-YAP Abs. PCNA was used as loading control. F Representative images of proximity ligation assay (PLA) detection of direct protein–protein interaction between ZEB1 and YAP (red signals) in PMOV10 cells stimulated or not with ET-1 for 6 h. DAPI staining (blue) highlights the nucleus (Magnification: 63x; scale bar: 10 μm). Right graph represents the quantification of the ZEB1/YAP protein interaction. Bars are means ± SD (n = 3; **p < 0.01)