Title : Effects of low cAMP levels on plant immunity after infection with an avirulent strain of Pseudomonas syringae
Abstract:
Plants had to constantly deal with several environmental factors such as biotic and abiotic stresses, which can alter the oxidative metabolism and photosynthetic efficiency thus affecting plant growth and development. Despite remarkable phenotypic and metabolic differences of defence responses implemented by plants after stress exposure, only few chemical species are involved in the signalling pathways inducing them.
Plant defence responses seem to be mainly regulated by the amounts, timings and cellular compartmentalization of the produced stress-related metabolites. Among the signalling molecules inducing the activation of defence pathways, calcium ions (Ca2+), reactive oxygen species (ROS) and cyclic nucleotides (cNMP) play pivotal roles. Recent evidence suggests that specific calcium signatures, followed by Ca2+-mediated apoplastic oxidative bursts, are likely involved in both local and long-distance signalling. The amount, timing and type of ROS produced under stress conditions as well as their cellular localization, seem to be also critical for activating defence pathways in response to environmental stresses. In particular, hydrogen peroxide (H2O2) is one of the most interesting ROS for its action as signalling molecule due to its stability, higher than that of other ROS, and its capability to cross biological membranes. In higher plants, the presence and the physiological relevance of 3’-5’-cyclic adenosine monophosphate (cAMP) was becoming known only in the last decades, whereas its mechanism of action is still poorly characterized.
In this study, a more reliable approach has been developed, by generating Arabidopsis thaliana plants overexpressing the ‘cAMP-sponge’ (cAS plants), with the aim to genetically and specifically buffer cAMP levels.
We decided to use this genetic tool to investigate on cAMP role in plant immune response triggered by the avirulent pathogen Pseudomonas syringae pv. tomato DC3000 carrying the avirulence gene AvrB (PstAvrB). A higher bacterial growth and a reduced hypersensitive cell death were observed in cAS plants compared to wild-type plants. The genetically controlled lowering of intracellular cAMP levels results in the inability of cAS plants to manage the correct equilibrium between ROS production and scavenging thereby inducing an impairment of the redox homeostasis after pathogen infection. Consistently, the low cAMP availability induces a significant delay in the timing of cytosolic Ca2+ and H2O2 increase following PstAvrB exposure. Analyses at biochemical and molecular levels have been also carried out in both control and infected plants over treatment time. In particular, ascorbate (ASC) and glutathione (GSH) metabolisms, as well as expression and activity of redox enzymes involved in ROS scavenging were investigated in both genotypes 4 h and 24 h post-infection. Moreover, a comparative proteomic analysis performed on WT and cAS plants 24 h post-infection reveals the occurrence of a transcriptional reprogramming that involve a core set of proteins regulated in both genotypes and other proteins uniquely modulated in WT and cAS plants. Concluding, our data confirmed that low cAMP levels affect the timing and intracellular levels of key signalling molecules thus altering the induction of defence redox systems and consequently compromising the plant immune responses triggered by pathogen infection.
