Phospho yap1

Antibodies against PDEF (Santa Cruz Biotechnology, Inc. sc-166846, 1:1000 dilution), GAPDH (Sigma G8795, 1:3000 dilution), YAP1 (Santa Cruz Biotechnology, Inc. sc-101199, 1:1000 dilution for western blots, 1:50 dilution for Immunofluorescence), and phospho-YAP1 (Cell Signaling Technology (CST) cst-13008, 1:1000 dilution) were purchased from respective vendors.

Cells were harvested in RIPA buffer (150 mM sodium chloride, 1% Triton X-100, 1% sodium deoxycholate, 0.1% SDS, 50 mM Tris–HCl, pH7.5 and 2 mM EDTA) with 1% protease and phosphatase inhibitor cocktails (Sigma). Equal amount of proteins were separated by SDS/PAGE and analysed by immunoblotting. Western blotting was prepared by standard procedures using anti-YAP1 (Santa Cruz clone H-9, Cat. No.sc-271134), phospho-YAP1 (Ser127; Cell Signaling Technology, Cat. No. 4911), anti-TAZ (BD Pharmingen clone M2-616), anti-cofilin (Abcam, Cat. No. ab42824), anti-cofilin (phospho-S3; Abcam, Cat. No. ab12866), anti-gelsolin (Santa Cruz ABS017, Cat. No. sc-57509), anti-phospho-p44/42 MAPK (phospho-ERK1/2, Thr202/Tyr204; Cell Signaling Technology, Cat. No. 9101), anti-p44/42 MAPK (ERK1/2; Cell Signaling Technology, Cat. No. 9102), anti-FAK (Cell Signaling Technology, Cat. No. 3285), anti-phospho-FAK (Tyr397; Cell Signaling Technology, Cat. No. 3283) and β-actin (Santa Cruz clone C4, Cat. No.sc-271134) antibodies. The original scans of the blots are shown in Supplementary Fig. 11.

Cells were lysed in a standard protein lysis buffer. Protein concentrations were measured with Bradford reagent (Bio-Rad, Germany). Protein was loaded in equal amounts and separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis, which followed by transferring to polyvinylidene difluoride membranes (Merck Millipore, Germany). Membranes were incubated with primary antibodies against YAP-1 (Abcam, UK; ab56701), CTGF (Abcam, UK; ab6992), α-SMA (Abcam, UK; ab5694), platelet-derived growth factor receptor-β (PDGFR-β; Cell Signaling Technology, USA; #3170), S100A4 (Abcam, UK; ab197896), Phospho-YAP-1 (Cell Signaling Technology, USA; #13008), Integrin αVβ6 (Bioss, USA; bs-5791R), TGF-β1 (Abcam, UK; ab215715), GAPDH (Abcam, UK; ab181602), Lamin B1 (Cell Signaling Technology, USA; #13435), and β-actin (Sigma-Aldrich, Germany; A2228), independently, which followed by the incubation with anti-mouse IgG-HRP (GE Healthcare UK Limited, UK; RPN4301) or anti-rabbit IgG-HRP (GE Healthcare UK Limited, UK; NA934) secondary antibodies. The bands were visualized by SuperSignal West Pico Chemiluminescent Substrate (Thermo Fisher Scientific, UK) and photographed with an image acquisition system of ECL ChemoCam Imager (Intas GmbH, Germany).