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. 2015 Jan 20;290(11):6810–6824. doi: 10.1074/jbc.M114.606699

FIGURE 6.

FIGURE 6.

DEPTOR depletion enhances and DEPTOR overexpression attenuates CXCR4-promoted Akt activation and signaling. A and B, role of ESCRTs in regulating the association of DEPTOR with mTORC2 and mTORC1. HeLa cells were treated with siRNA directed against STAM1 or luciferase (A), followed by transfection with FLAG-tagged DEPTOR and with Myc-tagged Raptor or empty vector (pcDNA) (B). Endogenous Rictor (A) or myc-Raptor (B) were immunoprecipitated (IP) and analyzed for endogenous DEPTOR (A) or FLAG-DEPTOR (B), respectively, and the indicated proteins. Representative blots from three independent experiments are shown. The asterisks in A indicate nonspecific bands. C and D, role of DEPTOR in regulating Akt activation promoted by CXCR4. HeLa cells were transfected FLAG-DEPTOR or shRNA targeting DEPTOR, followed by serum starvation and stimulation with CXCL12. Cell lysates were analyzed for the indicated proteins and their phosphorylation status. Data are from three independent experiments. E, role of CXCR4 activation on DEPTOR association with mTORC2. HeLa cells were stimulated with CXCL12 for 5 or 30 min or vehicle, followed by immunoprecipitation of endogenous Rictor and immunoblotting for DEPTOR and mTOR. Representative blots from two independent experiments are shown. Ctrl, control.