Tandem immunoprecipitation of phosphotyrosine-mass spectrometry (TIPY-MS) indicates C19ORF19 becomes tyrosine-phosphorylated and associated with activated epidermal growth factor receptor

To identify phosphotyrosine (pY) sites in the epidermal growth factor receptor (EGFR) network, a tandem immunoprecipitation-mass spectrometry method (TIPY-MS) was applied wherein protease-digested EGFR immune complexes were extracted with anti-pY after Rush et al. (Nat. Biotech. 2005, 23, 94) and analyzed by LC-MS/MS. New pY sites in the pathway were found, including SOS1 Y1065, SOS2 Y1275, CBL-B Y889, and in the EGFR regulatory protein Mig-6 Y458. The novel human C19orf19 gene product was found EGFR-associated and phosphorylated at 5 tyrosines in response to EGFR activation and, therefore, represents a new component of the EGFR signaling network.