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. 2021 Aug 19;297(4):101100. doi: 10.1016/j.jbc.2021.101100

Figure 1.

Figure 1

Amino acid stimulation at alkaline but not physiological pH increases mTORC2 and AMPK signaling.A, MEFs were cultured in complete media (DMEM/FBS) (left) or serum-starved overnight, ∼16 h (right). They were next amino acid–starved in D-PBS (50 min) with (left) or without (right) dFBS, pretreated with Torin 1 (T) and treated without (−) or with (+) a commercial amino acid solution (amino acid–sol) (10 min) to [1×] or [5×] final. Whole-cell lysates were immunoblotted as indicated. Note that [1×]f and [5×]f represents roughly the amino acids found in RPMI or DMEM, respectively. B, MEFs were cultured as in panel A but amino acid–starved in amino acid–free DMEM and refed with DMEM at pH 7.3 or 8.3 lacking (−) or containing (+) amino acids (10 min). C, MEFs were amino acid–starved in D-PBS/dFBS, pre-treated with Torin 1 (T), and stimulated with an amino acid solution at pH 10 or 7.4 (10 min). AMPK, AMP-activated protein kinase; D-PBS, Dulbecco's PBS; dFBS, dialyzed FBS; MEFs, mouse embryonic fibroblasts; mTOR, mechanistic target of rapamycin; mTORC2, mTOR complex 2.