Superoxide-responsive gene expression in Arabidopsis thaliana and Zea mays

Superoxide (O₂^-) and other reactive oxygen species (ROS) are generated in response to numerous biotic and abiotic stresses. Different ROS have been reported to elicit different transcriptional responses in plants, and so ROS-responsive marker genes and promoter::reporter gene fusions have been proposed as indirect means of detecting ROS and discriminating among different species. However, further information about the specificity of transcriptional responses to O₂^- is needed in order to assess potential markers for this critical stress-responsive signaling molecule. Using qRT-PCR, the expression of 12 genes previously reported to be upregulated by O₂^- was measured in Arabidopsis thaliana plants exposed to elicitors of common stress-responsive ROS: methyl viologen (an inducer of O₂^-), rose bengal (an inducer of singlet oxygen, ¹ΔO₂), and exogenous hydrogen peroxide (H₂O₂). Surprisingly, Zinc-Finger Protein 12 (AtZAT12), which had previously been used as a reporter for H₂O₂, responded more strongly to O₂^- than to H₂O₂; moreover, the expression of an AtZAT12 promoter-reporter fusion (AtZAT12::Luc) was enhanced by diethyldithiocarbamate, which inhibits dismutation of O₂^- to H₂O₂. These results suggest that AtZAT12 is transcriptionally upregulated in response to O₂^-, and that AtZAT12::Luc may be a useful biosensor for detecting O₂^- generation in vivo. In addition, transcripts encoding uncoupling proteins (AtUCPs) showed selectivity for O₂^- in Arabidopsis, and an AtUCP homolog upregulated by methyl viologen was also identified in maize (Zea mays L.), indicating that there are O₂^--responsive members of this family in monocots. These results expand our limited knowledge of ROS-responsive gene expression in monocots, as well as O₂^--selective responses in dicots.