Reagents
Cell culture and SILAC labeling
Cell viability assay
Immunoprecipitation
RNAi and transfection
Caspase-3/7 activity measurements
Sample preparation for MS-based interactome analysis
Sample preparation for MS-based phosphotyrosine interactome analysis
Sample preparation for MS-based phosphoproteome analysis
TMT labeling and phosphopeptide enrichment
Sample preparation for MS-based proteome analysis
Liquid chromatography–tandem mass spectrometry
MS data analysis
Bioinformatic analysis
Statistical analysis
NB1, SH-SY5Y, CLBGA, NBL-S, and H3122 cells were cultured in complete medium and treated for the indicated time points with inhibitor (TAE684, LDK378, crizotinib, lorlatinib, LY294002, or U0126) as indicated, with their respective IC50 values reported. Control cells were treated with similar amount of DMSO as the treated cultures (0.1% DMSO). IGF-1 and insulin stimulations were performed using 100 ng/ml for 10 min after ALK inhibitor pretreatment. Cell lysates, protein concentration determination, and Western blotting were carried out as previously described (29).