2,3,5-Tris(glutathion-S-yl)hydroquinone (TGHQ), a metabolite of benzene, catalyzes the generation of reactive oxygen species (ROS) and caspase-dependent apoptosis in human promyelocytic leukemia (HL-60) cells. We now report that TGHQ induces severe DNA damage, as evidenced by DNA ladder formation and H2AX phosphorylation. The subsequent activation of the DNA nick sensor enzyme, poly(ADP-ribose) polymerase-1 (PARP-1), leads to the rapid depletion of ATP and NAD and the concomitant formation of poly(ADP-ribosylated) proteins (PARs). PJ-34 (a PARP-1 inhibitor) completely prevented the formation of PARs, partially attenuated TGHQ-mediated ATP depletion, but had little effect on NAD depletion. Intriguingly, although z-vad-fmk (a pan-caspase inhibitor) attenuated TGHQ-induced apoptosis, cotreatment with PJ-34 led to a further decrease in apoptosis, suggesting that PARP-1 participates in caspase-dependent apoptosis. Indeed, PARP-1 inhibition reduced TGHQ-induced caspase-3, -7, and -9 activation, at least partially by attenuating cytochrome c translocation from mitochondria to the cytoplasm. In contrast, PJ-34 potentiated TGHQ-induced caspase-8 activation, suggesting that PARP-1 plays a dual role in regulating TGHQ-induced apoptosis via opposing effects on the intrinsic (mitochondrial) and extrinsic (death-receptor) pathways. PARP-1 knockdown in HL-60 cells confirmed that PARP-1 participates in effector caspase activation. Finally, PJ-34 also inhibited TGHQ-induced apoptosis-inducing factor (AIF) nuclear translocation, but neither c-jun NH(2)-terminal kinase nor p38 MAPK (p38 mitogen-activated protein kinase) activation was required for AIF translocation. In summary, TGHQ-induced apoptosis of HL-60 cells is accompanied by PARP-1, caspase activation, and AIF nuclear translocation. TGHQ-induced apoptosis appears to primarily occur via engagement of the mitochondrial-mediated pathway in a process amenable to PARP inhibition. Residual cell death in the presence of PJ-34 is likely mediated via the extrinsic apoptotic pathway.
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