Functional Analysis of Arabidopsis Mutants Points to Novel Roles for Glutathione in Coupling H 2 O 2 to Activation of Salicylic Acid Accumulation and Signaling

Abstract

AIMS: Through its interaction with H(2)O(2), glutathione is a candidate for transmission of signals in plant responses to pathogens, but identification of signaling roles is complicated by its antioxidant function. Using a genetic approach based on a conditional catalase-deficient Arabidopsis mutant, cat2, this study aimed at establishing whether GSH plays an important functional role in the transmission of signals downstream of H(2)O(2). RESULTS: Introducing the cad2 or allelic mutations in the glutathione synthesis pathway into cat2 blocked H(2)O(2)-triggered GSH oxidation and accumulation. While no effects on NADP(H) or ascorbate were observed, and H(2)O(2)-induced decreases in growth were maintained, blocking GSH modulation antagonized salicylic acid (SA) accumulation and SA-dependent responses. Other novel double and triple mutants were produced and compared with cat2 cad2 at the levels of phenotype, expression of marker genes, nontargeted metabolite profiling, accumulation of SA, and bacterial resistance. Most of the effects of the cad2 mutation on H(2)O(2)-triggered responses were distinct from those produced by mutations for GLUTATHIONE REDUCTASE1 (GR1) or NONEXPRESSOR OF PATHOGENESIS-RELATED GENES 1 (NPR1), and were linked to compromised induction of ISOCHORISMATE SYNTHASE1 (ICS1) and ICS1-dependent SA accumulation. INNOVATION: A novel genetic approach was used in which GSH content or antioxidative capacity was independently modified in an H(2)O(2) signaling background. Analysis of new double and triple mutants allowed us to infer previously undescribed regulatory roles for GSH. CONCLUSION: In parallel to its antioxidant role, GSH acts independently of NPR1 to allow increased intracellular H(2)O(2) to activate SA signaling, a key defense response in plants.