Activated eIF4E-binding protein slows G1 progression and blocks transformation by c-myc without inhibiting cell...

Translation initiation is poised between global regulation of cell growth and specific regulation of cell division. The mRNA cap-binding protein (eIF4E) is a critical integrator of cell growth and division because it is rate-limiting for translation initiation and is also rate-limiting for G(1) progression. Translation initiation factor eIF4E is also oncogenic and a candidate target of c-myc. Recently, an activated inhibitory 4E-binding protein (4EBP) that blocks eIF4E was used to study its regulation of Drosophila growth. We adopted this approach in mammalian cells after identifying an autosensing mechanism that protects against increased levels of 4EBP1. Increased 4EBP1 induced a quantitative increase in the inactivated phosphorylated form of 4EBP1 in vitro and in vivo. To overcome this protective mechanism, we introduced alanine substitutions at four phosphorylation/inactivation sites in 4EBP1 to constitutively activate a 4EBP mu to block eIF4E. Overexpression of activated 4EBP mu inhibited cell proliferation and completely blocked transformation by both eIF4E and c-myc, although it did not block all tested oncogenes. Surprisingly, expression of the activated 4EBP mu increased cell size and protein content. Activated 4EBP mu blocked both cell proliferation and c-myc transformation by inhibiting G(1) progression and increasing apoptosis, without decreasing protein synthesis. Our results identify mammalian eIF4E as rate-limiting for cell cycle progression before it regulates cell growth. It further identifies G(1) control by translation initiation factors as an essential genetic target of c-myc that is necessary for its ability to transform cells.