Decreased intracellular superoxide levels activate Sindbis virus- induced apoptosis

Infection of many cultured cell types with Sindbis virus (SV), an alphavirus, triggers apoptosis through a commonly utilized caspase activation pathway. However, the upstream signals by which SV activates downstream apoptotic effectors, including caspases, remain unclear. Here we report that in AT-3 prostate carcinoma cells, SV infection decreases superoxide (O₂/·^- ) levels within minutes of infection as monitored by an aconitase activity assay. This SV-induced decrease in O₂/·^- levels appears to activate or modulate cell death, as a recombinant SV expressing the O₂/·^- scavenging enzyme, copper/zinc superoxide dismutase (SOD), potentiates SV-induced apoptosis. A recombinant SV expressing a mutant form of SOD, which has reduced SOD activity, has no effect. The potentiation of SV-induced apoptosis by wild type SOD is because of its ability to scavenge intracellular O₂/·^- rather than its ability to promote the generation of hydrogen peroxide. Pyruvate, a peroxide scavenger, does not affect the ability of wild type SOD to potentiate cell death; and increasing the intracellular catalase activity via a recombinant SV vector has no effect on SV-induced apoptosis. Moreover, increasing intracellular O₂/·^- by treatment of 3T3 cells with paraquat protects them from SV-induced death. Altogether, our results suggest that SV may activate apoptosis by reducing intracellular superoxide levels and define a novel redox signaling pathway by which viruses can trigger cell death.