vFLIP K13 blocks growth factor withdrawal–induced apoptosis by blocking loss of mitochondrial membrane potential and caspase-activation.

(A) vFLIP K13–expressing cells maintain their mitochondrial potential in the absence of GM-CSF. TF-1 vector– and vFLIP-K13–expressing cells were grown in the absence or presence of GM-CSF for 24 hours. Cells were stained with 100 nM TMRE for 30 minutes at 37°C and fluorescence determined using a flow cytometer. (B) Inhibition of caspase-9, -3, and -8 cleavage during GM-CSF withdrawal–induced apoptosis in TF-1 vFLIP K13 cells. TF-1 vector and vFLIP K13 cells were grown in the presence and absence of GM-CSF for the indicated time periods. Cell lysates were analyzed for cleavage of caspase-9, -3, and -8 by Western blot analysis using antibodies that can recognize their cleaved forms. The blot was reprobed with a polyclonal antibody against actin to show equal loading of all lanes. (C) Inhibition of caspase-6 cleavage during GM-CSF withdrawal–induced apoptosis in TF-1 vFLIP K13 cells. The experiment was performed essentially as described for panel B, except expression of caspase-6 was analyzed using an antibody that recognizes its full-length form.