4E-BP1 phosphorylation is mediated by the FRAP-p70(s6k) pathway and is independent of mitogen-activated protein kinase

It has previously been argued that the repressor of protein synthesis initiation factor 4E, 4E-BP1, is a direct in vivo target of p42(mapk). However, the immunosuppressant rapamycin blocks serum-induced 4E-BP1 phosphorylation and, in parallel, p70(s6k) activation, with no apparent effect on p42(mapk) activation. Consistent with this finding, the kinetics of serum-induced 4E-BP1 phosphorylation closely follow those of p70(s6k) activation rather than those of p42(mapk). More striking, insulin, which does not induce p42(mapk) activation in human 293 cells or Swiss mouse 3T3 cells, induces 4E-BP1 phosphorylation and p70(s6k) activation in both cell types. Anisomycin, which, like insulin, does not activate p42(mapk) promotes a small parallel increase in 4E-BP1 phosphorylation and p70(s6k) activation. The insulin effect on 4E-BP1 phosphorylation and p70(s6k) activation in both cell types is blocked by SQ20006, wortmannin, and rapamycin. These three inhibitors have no effect on p42(mapk) activation induced by phorbol 12-tetradecanoate 13-acetate, though wortmannin partially suppresses both the p70(s6k) response and the 4E-BP1 response. Finally, in porcine softie endothelial cells stably transfected with either the wild-type platelet-derived growth factor receptor or a mutant receptor bearing the double point mutation 740F/751F, p42(mapk) activation in response to platelet-derived growth factor is unimpaired, but increased 4E-BP1 phosphorylation is ablated, as previously reported for p70(s6k). The data presented here demonstrate that 4E-BP1 phosphorylation is mediated by the FRAP-p70(s6k) pathway and is independent of mitogen-activated protein kinase.