Src l4a1

Crenolanib (Selleck Chemicals) and Dasatinib (TSZ Chemicals) was added to cells at final concentrations from 0.1–10μM. The following primary antisera were used for immunoblot analysis: PDGF Receptor (28E1 Cell Signaling, Danvers, MA), actin (C-11 Santa Cruz, Dallas, TX), phospho YAP^Y357 (ab62751 abcam, Cambridge, MA), phospho YAP^S127 (4911 Cell Signaling), total YAP (63.7 Santa Cruz), lamin B1 (D8P3U Cell Signaling), GAPDH (MAB374 Millipore, Temecula, CA), phospho Src^Y416 (2101 Cell Signaling), Src (L4A1 Cell Signaling), and Mcl-1 (S-19 Santa Cruz). The following primary monoclonal antibodies were used for immunofluorescence: total YAP (63.7 Santa Cruz) and anti-FLAG M2 (F1804 Sigma Adrich). For ChIP immunoprecipitation assays we employed an anti-YAP monoclonal antibody (63.7 Santa Cruz). ProLong Antifade with 4′,6-diamidino-2-phenylindole (DAPI, Life Technologies) was used for nuclear staining. The following primers used are listed in Table 1.

Cell culture lysates were analyzed with BCA Assay (Pierce) and mixed with Laemmli sample buffer and briefly boiled. Equal amounts of protein from each sample were separated using Criterion 10% SDS-PAGE (Bio-Rad), and transferred to nitrocellulose membranes. Membranes were then blocked with either 5% BSA or non-fat milk and probed with antibodies against pY1068 EGFR (1:1000, 3777S), pY845 EGFR (1:1000, 2231S), EGFR (1:1000, 2646S), p-Src Tyr 416 (1:1000; 2101S), Src (L4A1; 1:1000; 2110S) (Cell Signaling Technologies) and β-Actin (1:5000, A5441, Sigma), in manufacturer recommended diluent and supplemented with 0.05% Sodium Azide overnight at 4°C. Membranes were then probed with horseradish peroxidase (HRP)- conjugated secondary antibodies and detected using Pico or Femto chemiluminescence substrates (Fisher and GeneTex, respectively) and an AI600 Chemiluminescent Imager (GE Lifesciences).