Upregulated expression of both REDD1 and Sestrin2 contribute to resuppression of mTORC1 activity during prolonged Leu deprivation. REDD1+/+ (WT) and REDD1−/− MEFs stably expressing an shCont or shSN2 were generated as described in the Methods section. (A) Representative blots. (B–E) Quantitation of Western blots. p-p70S6K1, p70S6K1 phosphorylated on Thr389. Values are means ± SEM, n = 4. Labeled means without a common letter differ, P < 0.05. Leu, leucine; MEF, mouse embryo fibroblast; mTOR, mechanistic target of rapamycin; mTORC, mTOR complex; p70S6K1, 70 kDa ribosomal protein S6 kinase 1; REDD1, regulated in development and DNA damage responses 1; shCont, scrambled control shRNA; shRNA, short hairpin RNA; shSN2, shRNAs targeting Sestrin2; WT, wildtype.