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Figure 4
Rapamycin activates Ras-Raf1-MEK-ERK in vitro. (A) Effect of rapamycin treatment (20 nM, 24 h) on ERK, AKT, and RpS6 (S6) phosphorylation in MEFs with different genetic modifications (n = 3). p-S6, phosphorylated RpS6. (B) ERK, AKT, and RpS6 phosphorylation status in SV40-immortalized MEFs upon acute mTOR genetic deletion. (C) Raf1, ERK, and RpS6 phosphorylation status in MCF7 cells upon rapamycin treatment (20 nM, 24 h; n = 3). (D) Effect of rapamycin (20 nM, 24 h) and/or MEK inhibitor UO126 (10 μM, 24 h) on ERK and RpS6 phosphorylation in MCF7 cells (n = 3). (E) ERK and RpS6 phosphorylation in MCF7 transfected for 24 hours with an empty vector (mock) or a dominant-negative form of Ras (Ras^N17) and treated with rapamycin (20 nM, 24 h; n = 3). Numbers indicate the ratio of the phosphorylated protein related to total protein levels.