Fig. 2.
GST-MDA-7/IL-24 induces profound mitochondrial injury and caspase activation in human myeloid leukemia cells. A, U937 cells were exposed to the designated concentration of GST-MDA-7/IL-24 for 48 h, after which protein lysates were prepared and subjected to Western blot analysis to monitor caspase-3, caspase-8 cleavage/activation, and PARP degradation. B, HL60, MV4-11, and MLL/ENL cells were exposed to GST-MDA-7/IL-24 (100 nM for HL60 and MV4-11, and 50 nM for MLL/ENL) for the designated intervals, after which Western blot assays were performed as in A. C, U937, HL60, and MV4-11 cells were treated with GST-MDA-7/IL-24 (200 nM for U937 cells and 100 nM for HL60 and MV4-11 cells) for the designated intervals, after which mitochondria-free cytosolic fractions were obtained as described under Materials and Methods and subjected to Western blot analysis to monitor the release of cytochrome c and AIF into the cytosol. For this and all subsequent Western blot assays, lanes were loaded with 20 μg of protein; blots were subsequently reprobed with anti-tubulin (Tub) antibodies to document equivalent loading and transfer. The blots shown are representative of three or more independent experiments.