Death effector domain–containing initiator caspases and FADD are not required and do not enhance paclitaxel-induced cell death. (A) Lysates of Jurkat cells (WT) and their caspase-8– or FADD-deficient derivatives were subjected to immunoblot analysis. Protein bands specific for caspase-8, caspase-10, caspase-3, and FADD are indicated by arrowheads. (B) Wild-type, caspase-8–deficient, and FADD-deficient Jurkat cells were treated with 1 μM paclitaxel for 24 hours (middle panels) or left untreated (left panels). Hypodiploid nuclei indicative of paclitaxel-induced apoptosis were detected irrespectively of FADD or caspase-8 expression. The position of nuclei in G₀/G₁-phase (◁) and G₂/M-phase (◀) is marked. Cell-cycle analysis by a linear depiction of PI staining (right panels) revealed no blockage of the paclitaxel-induced G₂/M arrest due to lack of FADD expression. Gray histograms show the untreated controls, and black histograms represent paclitaxel-treated samples. Cells in the G₂/M phase are given as percentage of all vital, nonapoptotic cells. (C) Treatment of the different Jurkat cell clones with increasing concentrations of paclitaxel for 72 hours (left panel) or with 0.5 μM paclitaxel for the indicated time points (right panel) revealed a dose- and time-dependent increase in hypodiploid cells, regardless of caspase-8 or FADD expression. The results show the mean values (± SD). (D) Paclitaxel-induced caspase activation is independent of FADD and caspase-8. Immunoblot analysis of lysates from wild-type and FADD- or caspase-8–deficient Jurkat cells that were treated with 1 μM paclitaxel for 48 hours or left untreated. The positions of procaspase-8, procaspase-3, and β-actin are indicated with closed arrowheads. The cleaved fragments of active caspase-3 are marked with open arrowheads.