Insulin-like growth factor-I (IGF-I)-dependent activation of pp42/44 mitogen-activated protein kinase occurs independently of IGF-I receptor kinase activation and IRS-1 tyrosine phosphorylation

The proliferation and metabolism of H4IIE hepatoma cells is apparently medi ated through the insulin receptor. These cells, however, also have high-aff inity binding sites for insulin-like growth factor-I (IGF-I). Addition of i nsulin to H4IIE cells increased RNA synthesis, DNA synthesis and cell numbe r. IGF-I, on the other hand, was ineffective at concentrations equivalent t o the lowest effective insulin dose, although stimulation was observed with concentrations 100-fold higher. Similar results were obtained when glucose uptake was measured. Western blot analysis demonstrated that tyrosine phos phorylation patterns produced by insulin and IGF-I differed. In particular, phosphorylation of insulin receptor substrate-1 (IRS-1) was evident after treatment with insulin, but not after treatment with IGF-I. Correspondingly , insulin, but not IGF-I, stimulated receptor tyrosine kinase activity, In contrast with these results, both insulin and IGF-I induced mitogen-activat ed protein (MAP) kinase phosphorylation and activity at a concentration of 10 nM. The correlation between insulin-dependent and IGF-I-dependent MAP ki nase activation was confirmed by Western blot analysis of phosphorylated MA P kinase kinase (MEK). These results suggest that phosphorylation of IRS-1 is essential for both cell proliferation and glucose metabolism, but is unc oupled from the MAP kinase cascade. Furthermore, stimulation of MEK and MAP kinase is independent of receptor tyrosine kinase activity.