PML enhances Fas-mediated apoptosis and PML-Fas association is common in noncancerous tissues. (A) U937 cells were transduced with scrambled shRNA or PML-specific shRNAs as described in “Experiments using shRNA.” After selection with puromycin, cells were analyzed for protein expression with anti-PML and anti–α-tubulin antibodies to confirm down-regulation of PML. The cells were incubated with 0 or 50 ng/mL of FasL for 24 hours and then stained with propidium iodide and analyzed for apoptosis by flow cytometry. The data represent the means ± SD of 3 independent experiments. (B) Cell extracts of various mouse tissues were immunoprecipitated using antibody against Fas or IgG followed by Western blot analysis with antibodies against PML and Fas. The precipitates from HEK293 cell extracts by normal IgG antibody were used as negative control; the precipitates from HEK 293 cell extracts by anti-PML or anti-Fas antibody were used as positive control, respectively.