Roles of superoxide anion and hydrogen peroxide during replication of two unrelated plant RNA viruses in Nicotiana benthamiana

Reactive oxygen species (ROS), including superoxide anion (O₂ ⁻), hydrogen peroxide (H₂O₂), and hydroxyl radical, act as signaling molecules to transduce biotic and abiotic stimuli into stress adaptations in plants. A respiratory burst oxidase homolog B of Nicotiana benthamiana (NbRBOHB) is responsible for O₂ ⁻ production to inhibit pathogen infection during plant innate immunity. RBOH-derived O₂ ⁻ can be immediately converted into H₂O₂ by the action of superoxide dismutase. Interestingly, we recently showed that red clover necrotic mosaic virus (RCNMV), a plant positive-strand RNA [(+)RNA] virus, hijacks the host's ROS-generating machinery during infection. An RCNMV replication protein associates with NbRBOHB and triggers intracellular ROS bursts. These bursts are required for robust viral RNA replication. However, what types of ROS are required for viral replication is currently unknown. Here, we found that RCNMV replication was sensitive to an O₂ ⁻ scavenger but insensitive to an H₂O₂ scavenger. Interestingly, replication of another plant (+)RNA virus, brome mosaic virus, was sensitive to both types of scavengers. These results indicate a virus-specific pattern requirement of O₂ ⁻ and H₂O₂ for (+)RNA virus replication and suggest a conserved nature of the roles of ROS in (+)RNA virus replication.