Figure 2
Autophagy is required to maintain intracellular glutamine levels.
(A) PDAC cells were treated with CQ (10 μM) for 24 h. (B) Relative glutamine consumption rate of 8988 T cells expressing a control shRNA (shGFP) or a shRNA targeting ATG7. Western blot confirmed the knockdown of ATG7 expression. (C) WT MEFs and atg5−/− MEFs expressing KRas G12V were cultured in complete medium for 24 h. Western blot confirmed the knockdown of ATG5 expression. (A–C) Glutamine levels were measured in the medium using a metabolite analyzer (BioProfile Basic100 analyzer) to monitor glutamine consumption. The glutamine consumption rate was presented after normalization by the number of cells. (D) 8988 T cells were plated in complete medium, which was replaced by glutamine-free medium the following day and then incubated for another 24 h with or without CQ (10 μM). Glutamine levels were monitored by using LC-MS/MS. (E) 8988 T cells expressing a control (shGFP) or ATG7 shRNA were cultured in complete or glutamine-free medium for 24 h and glutamine levels were monitored by using LC-MS/MS. Error bars represent the s.d. of triplicate wells from a representative experiment. **p < 0.01.
