Activation of the mammalian target of rapamycin complex 1 is both necessary and sufficient to stimulate eukaryotic initiation factor 2B epsilon mRNA translation and protein synthesis

In a previous study we demonstrated a requirement for activation of mTORC1 in the stimulation of eIF2B epsilon mRNA translation in skeletal muscle in response to resistance exercise. Although that study established the necessity of mTORC1 activation. the experimental model used did not lend itself readily to address the question of whether or not mTORC1 activation was sufficient to produce the response. Therefore, the present study was designed to address the sufficiency of mTORC1 activation, using cultures of Rat2 fibroblasts in which mTORC1 signaling was repressed by serum/leucine-depletion and stimulated by repletion of leucine and/or IGF-1. Repletion with leucine and IGF-1 caused a shift of eIF2B epsilon mRNA into actively translating polysomes and a stimulation of new eIF2B epsilon protein synthesis, but had no effect on mRNAs encoding the other four eIF2B subunits. Stimulation of eIF2B epsilon translation was reversed by pre-treatment with the mTORC1 inhibitor rapamycin. Exogenous overexpression of FLAG-Rheb, a proximal activator of mTORC1, also caused a re-distribution of eIF2B epsilon. mRNA into polysomes and a stimulation of eIF2B epsilon protein synthesis. The stimulation of eIF2B epsilon mRNA translation occurred in the absence of any effect on eIF2B epsilon mRNA abundance. RNAi-mediated knockdown of eIF2B epsilon resulted in reduced cellular proliferation, a result that phenocopied the known cytostatic effect of mTORC1 repression. Overall the results demonstrate that activation of mTORC1 is both necessary and sufficient to stimulate eIF2B epsilon mRNA translation and that this response may represent a novel mechanism through which mTORC1 can affect mRNA translation initiation, rates of protein synthesis, and cellular growth/proliferation. (C) 2008 Elsevier Ltd. All rights reserved.