siRNA-mediated inhibition of perifosine-induced upregulation of DR5 ( A ), but not DR4 ( B ), attenuates augmented induction of apoptosis by the combination of perifosine and TRAIL. M4e cells were transfected with control (Ctrl), DR5 (A) or DR4 (B) siRNA. After 48 h, the cells were treated without and with the combination of 5 μM perifosine and 12.5 ng/ml TRAIL (PRFS/TRAIL). After 24 h, the cells were harvested for preparation of whole-cell protein lysates and subsequent Western blot analysis to detect DR5, DR4 and caspase cleavage (A and C) and for detection of apoptosis with annexin V-flow cytometry (B and D). Columns in B and C are means ± SD of duplicated determinations.