Harpin Inactivates Mitochondria in Arabidopsis Suspension Cells

Abstract

Harpin is a well-known proteinaceous bacterial elicitor that can induce an oxidative burst and programmed cell death in various host plants. Given the demonstrated roles of mitochondria in animal apoptosis, we investigated the effect of harpin from Pseudomonas syringae on mitochondrial functions in Arabidopsis suspension cells in detail. Fluorescence microscopy in conjunction with double-staining for reactive oxygen species (ROS) and mitochondria suggested co-localization of mitochondria and ROS generation. Plant defense responses or cell death after pathogen attack have been suggested to be regulated by the concerted action of ROS and nitric oxide (NO). However, although Arabidopsis cells respond to harpin treatment with NO generation, time course analyses suggest that NO generation is not involved in initial responses but, rather, is a consequence of cellular decay. Among the fast responses we observed was a decrease of the mitochondrial membrane potential Δψm and, possibly as a direct consequence, of ATP production. Furthermore, treatment of Arabidopsis cells with harpin protein induced a rapid cytochrome C release from mitochondria into the cytosol, which is regarded as a hallmark of programmed cell death or apoptosis. Northern and DNA array analyses showed strong induction of protecting or scavenging systems such as alternative oxidase and small heat shock proteins, components that are known to be associated with cellular stress responses. In sum, the presented data suggest that harpin inactivates mitochondria in Arabidopsis cells.