FIGURE 9:

Apoptotic cell death in COPI-depleted cells is attenuated by MitoQ and a JNK inhibitor and increased by treatment with external lipids. (A) Depletion of the COPI complex induced apoptosis. U2OS, PC3, and DU145 cells were transfected either with control or COPZ1 siRNAs in triplicates. The percentage of apoptotic cells was determined by TUNEL assay. Plot shows the percentage of TUNEL positive cells ±SD. (B) MitoQ effects on JNK phosphorylation and apoptosis. DU145, U2OS, or PC3 cells were transfected with siCONT, siCOZ1 or siCOPA. 48 h posttransfection cells were treated with vehicle control or MitoQ. At 72 h cells were lysed and PARP cleavage, JNK phosphorylation, total level of JNK, COPA, and COPZ1 expression were determined by WB with the corresponding antibodies. (C). Effects of JNK inhibitor on JNK phosphorylation and apoptosis. DU145, U2OS, or PC3 cells were transfected with siCONT, or siCOPZ1. 48 h posttransfection cells were treated with vehicle control or JNK inhibitor. At 72 h cells were lysed and PARP cleavage, JNK phosphorylation, and total level of JNK, COPA, and COPZ1 expression were determined by Western blotting with the corresponding antibodies. (D). Depletion of the COPI complex sensitizes cells to lipotoxicity. U20S cells were transfected with siCONT or siCOPA; 48 h posttransfection, cells were treated with different concentrations of oleic acids for 24 h. The percentage of dead cells was analyzed using live/dead viability/cytotoxicity assay. Plot represents dependence of percentage of alive ethidium homodimer-1 negative cells. Average of triplicates, ± SE.