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. Author manuscript; available in PMC: 2008 Jul 18.
Published in final edited form as: Science. 2008 May 22;320(5882):1496–1501. doi: 10.1126/science.1157535

Fig. 2.

Fig. 2

Effects of overexpressed RagBGTP-containing heterodimers on the mTORC1 pathway and its response to leucine, amino acids, or insulin. Effects of expressing the indicated proteins on the phosphorylation state of co-expressed S6K1 in response to deprivation and stimulation with (A) leucine, (B) total amino acids, or (C) insulin. Cell lysates were prepared from HEK-293T cells deprived for 50 minutes of serum, and (A) leucine or (B) amino acids, and then, where indicated, stimulated with leucine or amino acids for 10 minutes. HEK-293E cells (C) were deprived of serum for 50 minutes and where indicated stimulated with 150 nM insulin for 10 minutes. Lysates and FLAG-immunoprecipitates were analyzed for the levels of the specified proteins and the phosphorylation state of S6K1. (D) Effects of amino acid deprivation on insulin-mediated activation of mTORC1. HEK-293E cells were starved for serum and amino acids or just serum for 50 minutes, and where specified, stimulated with 10 or 150 nM insulin. Cell lysates were analyzed for the level and phosphorylation state of S6K1.