Proteomics of NGF-TrkA signaling dynamics in neuroblastoma cells

We used quantitative mass spectrometry-based proteomics to unravel global nerve growth factor (NGF)-induced TrkA signaling dynamics at the interactome, phosphoproteome and proteome level. A tetracycline-inducible system for TrkA expression was generated in the human neuroblastoma cell line, SH-SY5Y. TrkA-induced cells were stimulated with NGF for different time points to follow phosphoproteome, interactome and proteome changes on a temporal scale. In a triple SILAC setup (Light: Lys0,Arg0; Medium: Lys4,Arg6; and Heavy: Lys8,Arg10), the samples were stimulated with NGF as indicated. Phosphoproteome: 0, 10, 45 min and 0, 120, 120 min+cycloheximide. Interactome: 0, 5, 10 min. Proteome: 0, 24, 48 h. All experiments were performed as biological replicates.