FIGURE 3.

Depletion of mTORC2 subunit Rictor attenuates phosphorylation of Akt at Ser-473 upon CXCR4 activation. A, time course of Akt activation promoted by stimulation of CXCR4. HeLa cells were treated with vehicle (PBS + 0.1% BSA) or CXCL12 for the indicated times, as described under “Experimental Procedures.” Samples were immunoblotted using anti-phosphospecific antibodies at Ser-473 (pAkt-S473) or Thr-308 (pAkt-T308) and total Akt. Immunoblots were analyzed by densitometry. Graph shows the average pAkt-S473 levels normalized to total Akt at each time point compared with control (5-min time point; set to 100%). Data are from three independent experiments, and error bars represent mean ± S.E. B, depletion of Rictor attenuates CXCR4-promoted phosphorylation of Akt at Ser-473. HeLa cells treated with siRNA directed against Rictor or luciferase (Ctrl) were serum-starved for 3 h followed by treatment with vehicle (minus symbol) or CXCL12 (plus symbol) for 5 min, as described under “Experimental Procedures.” Equal amounts of whole cell lysates were immunoblotted for the indicated proteins and their phosphorylation status. Representative blots from three independent experiments are shown. Arrow points to pAkt-T308 band. The asterisk represents nonspecific bands.