Fig. 6

ECM stiffness regulates YAP activity through the Rap1a/ARHGAP35/RhoA/F-actin pathway. a Immunofluorescence staining showing the expression of RhoA in ESCs. b Pull-down assays of total RhoA and activated RhoA (RHOA-GTP) in ESCs plated on 1 kPa and 40 kPa hydrogels for 24 h. Scale bars, 20 μm. c, d Confocal immunofluorescence images (c) and quantification of the nuclear and cytoplasmic subcellular localization of YAP (d) in ESCs. Cells were plated on 40 kPa fibronectin-coated hydrogels and treated with C3, Y27632, or Lat. A and Cyto. B for 24 h. Scale bars, 5 μm. e Lysates of ESCs treated with C3, Y27632, or Lat. A and Cyto. B for 24 h were analyzed for the presence of the indicated proteins. Data are means ± S.E.M. of at least three independent experiments. ***P < 0.001