Figure 6.

Inhibition of PAWR sustains autophagic phenotypes in CaP cells. (A) Representative fluorescence microscopy images show PC-3 cells were initially transiently transfected with control siRNA, PAWR siRNA, GFP and GFP-PAWR for 48 h and then treated with 0.75 and 1.0 μM of 3-AWA for 12 h. After 12 h, cells were stained with MDC to detect autophagosomes accumulated in cells. Original magnification 40×, scale bar = 100 μm. (B) Quantification of MDC positive cells by fluorescence microscopy was determined in 3 independent experiments (for details see text). Three random fields representing 100 cells were counted. (C) After transfection with the indicated constructs, cells were treated with the indicated concentrations of 3-AWA for 12 h and then subjected to apoptosis measurement by ANXA5 and propidium iodide staining. Representative dot plots display the number in each top and bottom right quadrant and represent the percentage of apoptotic cells positive for ANXA5. The data shown are representative of 3 individual experiments. The data represent the mean value ± SE of 3 independent experiments, **P < 0.01.