Fig. 4: AR regulates DRAM1 to promote both autophagy and BRAFi-resistance in melanoma.

DRAM1, LC3 and p62 protein expression were measured in A A375 and WM793 cells transfected with Flag-DRAM1, B A375R and WM793R cells transfected with si-DRAM1 or si-scramble C A375 and WM793 AR-OE cells transfected with si-DRAM1. D A375 cells expressing TF-LC3, AR-OE-TF-LC3 and AR-OE-TF-LC3 cells transfected with si-DRAM1 were imaged by fluorescence microscopy (Scale bars: 10 μm). White boxed regions in the panels are enlarged. Representative images of autolysosomes (red dots) and autophagosomes (yellow dots) are shown. Manually counted. Mean ± SEM, n = 30, two tailed Student’s t test, **p < 0.01, ***p < 0.001. E, F cell proliferation and colony formation assays were measured in cells from (D) treated with DAB as indicated. Mean ± SEM, n = 3, one-way ANOVA and Tukey’s post-hoc test. **p < 0.01. G, H WM793R cells transfected with/without si-DRAM1 were incubated with DAB (0.5 μM), and cell proliferation and colony formation assays were measured. Mean ± SEM, n = 3, two-tailed Student’s t test, **p < 0.01. E EV vs AR-OE **p = 0.0017, AR-OE + si-scramble vs AR-OE+si-DRAM1 **p = 0.0032, G DAB + si-scramble vs DAB + si-DRAM1 **p = 0.0047.