Generation of hydrogen peroxide is not required for harpin-induced apoptotic cell death in tobacco BY-2 cell suspension culture

To characterize molecular and biochemical mechanisms of hypersensitive reaction (HR) in plants, a tobacco suspension culture of BY-2 was treated with the proteinaceous HR elicitor harpin from several pathovars of Pseudomonas syringae. Tobacco BY-2 cells are sensitive to harpins derived from non-pathogenic pathovars of P. syringae, such as pvs. pisi, tomato and glycinea. These three harpins induce apoptotic cell death accompanied by DNA fragmentation in BY-2. Because the cell death was also accompanied by rapid generation of hydrogen peroxide (H2O2), one of the active oxygen species, we investigated the role of H2O2 in harpin-induced cell death. Although treatment with diphenylene iodium chloride (DPI) reduced, and catalase completely abolished, the harpin-induced generation of H2O2, the frequency of cell death was not affected at all. Treatment with superoxide dismutase (SOD) enhanced the generation of H2O2, but the cell death was unaffected. These results indicate that harpin-induced apoptotic cell death does not require oxidative burst.