Fig. 6: Partial activation of YAP by Merlin knockout can be reversed by increased contact inhibition.

a Percent p-Rb (all-cell gating) per local density bin for WT cells transfected with siRNA (48 h) against control (Ctrl), NF2/Merlin (NF2), and LATS2 (LATS2) in starvation (left) or serum (right) conditions. b Median YAP ratio (n/c) binned by local cell density for Merlin knockout (Merlin KO) cells in 0.1% (left) or 10% serum (right), compared with WT cells (same experiment as in Fig. 1e). Data are representative of 3 independent experiments. Shaded error are 95% confidence intervals. c TEAD-dependent hyperproliferation of Merlin KO cells. Percent p-Rb (G1-gated) per local density bin for Merlin knockout cells in 0.1% (left) or 10% serum (right) treated with DMSO or TEADi (0.5 μM GNE-7883), compared with DMSO-treated WT cells. Data are representative of 3 independent experiments. d Percent p-Rb (G1-gated) per local density bin for serum-starved Merlin KO cells treated with 0–20 μM gefitinib (EGFRi). Data are representative of 2 independent experiments. e Left: percent p-Rb (G1-gated) per local density bin for serum-starved Merlin KO cells with control (CTRL) or EGF (25 ng/mL) treatment for 14 h (data are representative of 2 independent experiments). Right: mean percent p-Rb (G1-gated) for starved Merlin KO cells treated with mock (CTRL), EGF, or serum (FBS) release for 12 h (n = 2 independent experiments). Student’s t test: p = 0.046 (EGF versus CTRL), 0.26 (FBS versus EGF). f Percent p-Rb (all-cell gating) per local density bin for Merlin KO (seeded at 11,000 cells/well) and conditioned in 0.1%, 1%, or 10% serum for 36 h. Data are representative of 3 independent experiments. g Normalized cyclin D1 (left) or p27 (right) protein levels for G1-gated Merlin KO cells in titrated serum. Protein levels are normalized to lowest density bin. Data are representative of 2 independent experiments. h Model for Hippo-pathway inhibition of YAP-TEAD activity, which regulates signaling through and downstream of receptor tyrosine kinases to control the cyclin D1/p27 ratio. i Model proposing transitions between low and high YAP-TEAD activity as contact inhibition is temporarily reduced then recovered after a delay during epithelial sheet regeneration. For all representative binned plots, data are pooled from at least 2 wells, n > 100 cells/bin, n > 10,000 cells/plot.