*4.5. Immunoblotting*

Cells were lysed in RIPA (radioimmunoprecipitation assay) buffer (Elpis Biotech, Daejeon, Korea) supplemented with a protease inhibitor cocktail (Calbiochem, La Jolla, CA, USA) and protein phosphatase inhibitors (Calbiochem). Protein concentrations were determined using a BCA assay kit (Pierce, Rockford, IL, USA). Proteins (10 μg/sample) were resolved in an SDS-PAGE gel and transferred to a nitrocellulose membrane (Millipore Corp., Billerica, MA, USA). Membranes were blocked with 5% skim milk prior to western blot analysis. Chemiluminescence was detected using an ECL kit (Advansta Corp., Menlo Park, CA, USA) and the Amersham Imager 600 (GE Healthcare Life Sciences, Little Chalfont, UK). The following primary antibodies were used on fresh individual membranes to minimize interference by stripping and reprobing: phospho-JNK (Thr183/Tyr185), JNK, phospho-p38 MAPK (Thr180/Tyr182), p38 MAPK, phospho-ERK1/2 (Thr202/Tyr204), ERK1/2, phospho-Src (Tyr416), Src, phospho-Lyn (Tyr507), Lyn, phospho-Akt (Ser473), Akt, phospho-PI3K p85 (Tyr458/Tyr199), PI3K p85, PI3K p110α, PI3K p110β, PI3K p110γ, PI3K p110δ, phospho-PTEN (Ser380/Thr382/383), PTEN, and Fyn (Cell Signaling Technology, Beverly, MA, USA); and β-actin and phospho-Fyn (Santa Cruz Biotechnology, Santa Cruz, CA, USA). All the raw data from immunoblotting experiments are presented in Figure S4.
