Manipulation of salicylate content in <i>Arabidopsis thaliana</i> by the expression of an engineered bacterial salicylate synthase

Salicylic acid (SA) plays a central role as a signalling molecule involved in plant defense against microbial attack. Genetic manipulation of SA biosynthesis may therefore help to generate plants that are more disease-resistant. By fusing the two bacterial genes <i>pchA</i> and <i>pchB</i> from <i>Pseudomonas aeruginosa</i>, which encode isochorismate synthase and isochorismate pyruvate-lyase, respectively, we have engineered a novel hybrid enzyme with salicylate synthase (SAS) activity. The <i>pchB-A</i> fusion was expressed in <i>Arabidopsis thaliana</i> under the control of the constitutive cauliflower mosaic virus (CaMV) <i>35S</i> promoter, with targeting of the gene product either to the cytosol (c-SAS plants) or to the chloroplast (p-SAS plants). In p-SAS plants, the amount of free and conjugated SA was increased more than 20-fold above wild type (WT) level, indicating that SAS is functional in Arabidopsis. P-SAS plants showed a strongly dwarfed phenotype and produced very few seeds. Dwarfism could be caused by the high SA levels <i>per se</i> or, perhaps more likely, by a depletion of the chorismate or isochorismate pools of the chloroplast. Targeting of SAS to the cytosol caused a slight increase in free SA and a significant threefold increase in conjugated SA, probably reflecting limited chorismate availability in this compartment. Although this modest increase in total SA content did not strongly induce the resistance marker <i>PR-1</i>, it resulted nevertheless in enhanced disease resistance towards a virulent isolate of <i>Peronospora parasitica</i>. Increased resistance of c-SAS lines was paralleled with reduced seed production. Taken together, these results illustrate that SAS is a potent tool for the manipulation of SA levels in plants.