Nitrate reductase is required for the transcriptional modulation and bactericidal activity of nitric oxide during the defense response of Arabidopsis thaliana against Pseudomonas syringae.

Nitrate reductase (NR) has emerged as a potential NO source in plants. Indeed, the Arabidopsis thaliana NR double-deficient mutant (nia1 nia2) produces low NO and develops abnormal susceptibility to bacterial infection. We have employed quantitative real-time polymerase chain reactions to analyze the effects of NO gas on the expression of defense-related genes in wild-type and nia1 nia2 A. thaliana plants that were inoculated with an avirulent strain of Pseudomonas syringae pv. tomato. The pathogenesis-related gene 1 (PR1) was up-regulated by bacterial infection, and its expression was higher in the wild type than in nia1 nia2. Fumigation with NO attenuated the expression of PR1 and other salicylic acid-related genes in plants that had been inoculated with P. syringae. Nevertheless, NO inhibited the most intense bacterial growth and disease symptoms in nia1 nia2 leaves. The NO fumigation also directly modulated lignin biosynthesis-related gene expression (CAD1) and parts of the auxin (TIR1, ILL1, GH3) and ethylene (ACCS7) pathways, among other defense-related genes, and their modulation was more intense in the NR-deficient mutant. Pathogen inoculation induced delayed but intense H2O2 production in mutant leaves in comparison with the wild type. Hydrogen peroxide potentiated the microbicidal effects of NO against bacterial cultures. These results suggest that NO has a direct microbicidal effect in combination with H2O2 to allow for the attenuation of the SA-mediated defense response, thereby reducing the energy expenditure associated with defense-related gene transcription. Overall, these results highlight the importance of NR-dependent NO production in the establishment of disease resistance.