Bacterial and H2O2 stress-induced apoptosis-like events in Cryptococcus neoformans

Cell death induced by adherence of Staphylococcus aureus was observed in Cryptococcus neoformans. To investigate this death mechanism, we examined three factors: actin dynamics using rhodamine-phalloidin staining, accumulation of reactive oxygen species (ROS) via conversion of 2′,7′-dichlorofluorescein diacetate to 2′,7′-dichlorofluorescein, and DNA fragmentation via terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL). In co-culture, actin visibly clumped and the percentage of ROS- and TUNEL-positive cells increased. The same cellular events were observed in C. neoformans after addition of hydrogen peroxide (H2O2), creating a bacterial cell-free system. After H2O2 treatment and cell disruption, C. neoformans proteins in soluble fraction were separated by two-dimensional (2-D) electrophoresis. A voltage-dependent ion-selective channel was identified as one of the stress-enhanced proteins. Thus, the death of C. neoformans was accompanied by decreased actin turnover, enhanced ROS accumulation, and DNA fragmentation. Cell death appeared similar to apoptosis via mitochondrial pathways.