DFO in lysosomes induces autophagy. (A) HeLa cells were incubated with or without FAC (10μM) for 18 hours followed by incubation with or without DFO (100μM) or dextran-DFO for 16 hours. Cells were harvested for measurement of ferritin by ELISA or fixed and processed for immunofluorescence for LC3B as described in the Figure 2 legend. The amount of ferritin in FAC-loaded cells was taken as 100%. Error bars represent SD from 3 different experiments. (B) HeLa cells were transfected with or without dynamin K44A and incubated in the presence or absence of FAC (10μM) for 18 hours. DFO and chloroquine were added to a final concentration of 100μM for 6 hours. Cells were fixed and processed for immunofluorescence as in panel A. Percentage of LC3B accumulation was determined by analyzing 100 cells. Error bars represent SD from 3 different experiments.