Manipulation of salicylate content in Arabidopsis thaliana 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 pchA and pchB from Pseudomonas aeruginosa, which encode isochorismate synthase and isochorismate pyruvate-lyase, respectively, we have engineered a novel hybrid enzyme with salicylate synthase (SAS) activity. The pchB-A fusion was expressed in Arabidopsis thaliana under the control of the constitutive cauliflower mosaic virus (CaMV) 35S 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 per se 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 PR-1, it resulted nevertheless in enhanced disease resistance towards a virulent isolate of Peronospora parasitica. 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.
Mauch
F
F
Département de Biologie, Université de Fribourg, CH-1700 Fribourg, Switzerland.
Mauch-Mani
B
B
Gaille
C
C
Kull
B
B
Haas
D
D
Reimmann
C
C
D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000235:genetics / Q000502:physiology*; D001483:Base Sequence; D017796:Caulimovirus / Q000201:enzymology / Q000235:genetics; D003062:Codon; D019751:Intramolecular Transferases / Q000235:genetics / Q000378:metabolism; D008190:Lyases / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D010636:Phenols / Q000378:metabolism; D018515:Plant Leaves / Q000201:enzymology; D030821:Plants, Genetically Modified; D011550:Pseudomonas aeruginosa / Q000201:enzymology / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015183:Restriction Mapping; D012459:Salicylates / Q000378:metabolism*; D013844:Thiazoles
D003062:Codon; D010636:Phenols; D011993:Recombinant Fusion Proteins; D012459:Salicylates; D013844:Thiazoles; C025316:pyochelin; D008190:Lyases; C022620:salicylate synthetase; D019751:Intramolecular Transferases; C058486:isochorismate synthase