Figure 1.

A schematic illustration of the approach to identify c-Src kinase substrates in PDGF receptor signaling using SILAC. Mouse embryonic fibroblasts (NIH 3T3) were grown in three different stable isotope-labeled media. One population of cells growing in ¹²C₆-arginine-containing medium was left unstimulated, another population of cells growing in ¹³C₆-arginine-containing medium was stimulated with 100 ng/ml PDGF-BB ligand and the third population of cells growing in ¹³C₆-¹⁵N₄-arginine-containing medium was pretreated with SU6656 for 1 hour and then stimulated with 100 ng/ml PDGF-BB ligand. The cells were lyzed and tyrosine phosphorylated proteins were enriched and analyzed as described in materials and methods.