Inhibition of mammalian target of rapamycin (mTOR) induces phosphatidylinositol 3 kinase-dependent and Mnk-mediated eIF4E phosphorylation Free

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The initiation factor eukaryotic translation initiation factor 4E (eIF4E) plays a key regulatory role in initiating translation of mRNAs, including those encoding oncogenic proteins. Thus, eIF4E is considered a survival protein involved in cell cycle progression, cell transformation and apoptotic resistance. Phosphorylation of eIF4E (usually at Ser209) increases its affinity for the cap of mRNA, and may also favor its entry into initiation complexes. The best candidate for eIF4E phosphorylation is the MAPK-activated protein kinase Mnk1, which physically associates with eIF4F and directly phosphorylates eIF4E at Ser209. Mammalian target of rapamycin (mTOR) inhibitors suppress cap-dependent translation through inhibition of phosphorylation of eIF4E-binding protein 1 (4E-BP1). Paradoxically, we have shown that inhibition of mTOR signaling by mTOR inhibitors increases eIF4E phosphorylation in cancer cells (Sun et al. Cancer Research, 2005). In this study, we focused on revealing the mechanism by which mTOR inhibition increases eIF4E phosphorylation. Silencing of either mTOR or raptor could mimic mTOR inhibitors’ effects in increasing eIF4E phosphorylation. Moreover, knockdown of mTOR but not p70S6K abrogated rapamycin’s ability to increase eIF4E phosphorylation. Thus, these results indicate that mTOR inhibitor-induced eIF4E phosphorylation is secondary to mTOR/raptor inhibition and independent of p70S6K. mTOR inhibitors failed to increase eIF4E phosphorylation in phosphatidylinositol 3 kinase (PI3K) deficient cells, furthering the notion that mTOR inhibitors induce a PI3K-dependent eIF4E phosphorylation. We noted that mTOR inhibitors lost their activity in increasing eIF4E phosphorylation only in cells where both Mnk1 and Mnk2 are knocked out, indicating that mTOR inhibitors increase eIF4E phosphorylation through a Mnk-dependent mechanism. Since PI3K deficiency also inhibited rapamycin-induced Mnk phosphorylation, we conclude that mTOR inhibition increases eIF4E phosphorylation through a PI3K-dependent and Mnk-mediated mechanism. (Supported by GCC Distinguished Cancer Scholar award, NIH grant RO1 CA118450-01 and DOD grant W81XWH-05-0027-IMPACT).