Maintenance of oxidative phosphorylation protects cells from Actinobacillus actinomycetemcomitans leukotoxin‐induced apoptosis

Subnanomolar concentrations (3 × 10 −10  M) of Actinobacillus actinomycetemcomitans leukotoxin (Ltx) trigger apoptosis of JY cells, as shown by a decrease in mitochondrial transmembrane potential (ΔΨ m ), hyperproduction of reactive oxygen species (ROS) and release of cytochrome c from the intermembrane space. When compared with heat‐inactivated leukotoxin (ΔI Ltx) controls, ATP levels in Ltx‐treated JY cells continued to decrease during a 24 h experiment while cytoplasmic ADP concentrations were increasing. These results suggest that a blockage occurred in ATP/ADP exchange. To maintain ATP/ADP exchange, JY cells were transfected with bcl ‐2 and bcl ‐x L and incubated with Ltx. ATP levels of the transfected cells decreased to 67% (JY/ bcl ‐2) and 73% (JY/ bcl‐x L ) after the experiment. Furthermore, cytochrome c remained localized to the mitochondrial fraction of Ltx‐treated JY/ bcl ‐2 and JY/ bcl‐x L cells, whereas its presence in the cytoplasmic fraction of JY/ gen cells suggests an uncoupling of electron transport. Expression of bcl ‐2 and bcl‐x L in cells inhibited downstream apoptotic events such as cleavage of poly(ADP‐ribose) polymerase, DNA fragmentation and activation of a family of caspases. The results indicate that Ltx induces apoptosis through a mitochondrial pathway that involves decreased levels of the ADP in the mitochondrial matrix, a lack of substrate for ATP synthetase and arrest of oxidative phosphorylation.