Fig. 5. IGF-I stimulates c-Src and Fyn kinase activities. A, proliferating 3T3-L1 cells were serum starved overnight, then stimulated with 10 nM IGF-I. Top panel, a representative autoradiogram of enolase phosphorylation by immunoprecipitated c-Src from cells stimulated with IGF-I for 0, 1, 2, 5, and 10 min. The graph is a compilation of densitometry results (mean + SD) of three time course experiments; results were corrected for a zero time of 1 densitometry unit. B, kinase assay autoradiograms with corresponding Western blots of c-Src or Fyn from cells stimulated with 10 nM IGF-I for 1 min. An aliquot of immunoprecipitated c-Src or Fyn was removed for Western blotting prior to measurement of kinase activity using enolase (arrow) as substrate. Immunoreactive c-Src and Fyn are seen just above immunoglobulin heavy chain at Mr = 60,000 and 58,000, respectively (arrows). Controls, which used IGF-I-stimulated cell lysates, included immunoprecipitation with nonimmune IgG (Control i.p.), No sample (omission of immunoprecipitate), and no enolase (No substrate).