FIG. 1.

Identification of Rictor as a target of mTORC1 signaling. (A) Schematic representation of the agonists and pharmacological inhibitors used in this study. (B) HEK293 cells were transfected with empty vector or HA-tagged Rictor; serum starved overnight; and stimulated for 10 or 20 min with PMA (50 ng/ml), insulin (100 nM), EGF (50 ng/ml), or fetal bovine serum (10%). Immunoprecipitated (IP) Rictor was then assayed for phosphorylation with a phospho-motif antibody that recognizes the RXRXXpS/T consensus motif. (C and D) As for panel B, except cells were pretreated with U0126 (20 μM), rapamycin (100 nM), or wortmannin (100 nM) for 30 min prior to PMA or insulin stimulation. (E and F) Endogenous Rictor was immunoprecipitated from HEK293 (E) or HeLa (F) cells and assayed as for panel B.