Signalling molecules responsive to ozone-induced oxidative stress in Salvia officinalis

Tropospheric ozone (O₃) is the most important gaseous pollutant and induces a mass of negative impacts on vegetation at functional and genic levels. The aim of the present study was to investigate the role of reactive oxygen species and signalling molecules in sage plants exposed to O₃ (200 ppb, 5 h). Ozone exposure induced only a transient oxidative burst, as confirmed by the rapid peak of anion superoxide during the first hours of exposure (+16% compared to controls). The spontaneous reaction of O₃ with membrane fatty acids stimulates peroxidative processes, as demonstrated by the rise of thiobarbituric acid reactive substances concentration starting after 1 h of exposure (+25%). The formation of lipid-based signalling molecules (e.g. jasmonic acid) may be regarded as a sort of O₃-perception. The concomitant accumulation of salicylic acid suggests that sage responds early to O₃ by inducing cellular antioxidants mechanisms in order to minimize O₃-oxidative burst. The transient increase of abscisic acid (+25% at the end of the treatment) twinned with the maximal ethylene emission (about two-fold higher than controls) could be interpreted as a first attempt by plants to regulate the signalling responses induced by O₃. In order to investigate the involvement of transcription factors in managing oxidative protection, BLASTX analysis against the Salvia miltiorrhiza sequence genome was carried out using Arabidopsis thaliana WRKY sequences as queries. Six gene sequences were identified for sage WRKYs and their relative gene expression analyses were characterized. WRKY4, WRKY5, WRKY11 and WRKY46 were up-regulated by O₃ at 2 and 5 h of exposure and they showed similarity with AtWRKY48, AtWRKY22 and AtWRKY53 in A. thaliana. These results suggest that WRKYs could play a pivotal role in the signalling mechanisms during the responses of plants to O₃.