FIGURE 2. Death receptor-induced Atg5^−/− cell death results from caspase-dependent apoptosis.

A, caspase 8 activity was assayed in wild-type (WT) and Atg5^−/− MEFs treated with Jo2 antibody (Fas) for 8 or 12 h. Results are from 4 experiments (*, p < 0.01 as compared with wild-type cells). Con, untreated. B and C, protein was isolated from untreated wild-type and Atg5^−/−cells and cells treated with Jo2 (Fas) (B) or TNF-α (C) for the indicated hours. Aliquots of protein were immunoblotted with antibodies for caspase 3 (casp 3), PARP, cIAP2, TRAF2, Atg5, Atg7, Beclin 1, phospho-mTOR (P-mTOR), and mTOR. For Atg5, the band corresponding to the Atg5/12 conjugate protein is shown. Stripped membranes were reprobed with β-actin as a measure of equivalent protein loading among samples. The caspase 3 and PARP cleavage products are indicated by arrows. Immunoblots are representative of 3 independent experiments. D, Atg5^−/− cells were pretreated with Me₂SO (DMSO) vehicle, 5 (QVD 5) or 10 (QVD 10) µm Q-VD-OPh, or 25 µm IDN-1529 (1529). Cells were then treated with Jo2 antibody (Fas) or TNF-α, and the percentage of cell death was determined by MTT assay at 24 h. Results are from 3 independent experiments (*, p < 0.00001 as compared with Me₂SO-reated cells).