The nitric oxide donor sodium nitroprusside regulates polyamine and proline metabolism in leaves of Medicago truncatula plants

Abstract

Nitric oxide (NO), polyamines, and proline have all been suggested to play key roles in a wide spectrum of physiological processes and abiotic stress responses. Although exogenous application of polyamines has been shown to induce NO production, the effect of NO on polyamine biosynthesis has not yet been elucidated. Several reports exist that demonstrate the protective action of sodium nitroprusside (SNP), a widely used NO donor, which acts as a signal molecule in plants responsible for the regulation of the expression of many defense-related enzymes. This study attempted to provide a novel insight into the effects of application of low (100μΜ) and high (2.5mM) concentrations of SNP on the biosynthesis of two major abiotic stress response-related metabolites, polyamines and proline, in mature (40 day) and senescing (65 day) Medicago truncatula plants. Physiological data showed that long-term (24h), higher SNP concentration resulted in decreased photosynthetic rate and stomatal conductance followed by intracellular putrescine and proline accumulation, as a result of an increase in biosynthetic arginine decarboxylase (ADC) and Δ11Abbreviations used: ROS, reactive oxygen species; RNS, reactive nitrogen species; SNP, sodium nitroprusside; PA, polyamine; ODC, ornithine decarboxylase; ADC, arginine decarboxylase; SPDS, spermidine synthase; SPMS, spermine synthase; SAMDC, S-adenosylmethionine decarboxylase; DAO, diamine oxidase; PAO, polyamine oxidase; Put, putrescine; Spd, spermidine; Spm, spermine; P5CS, Δ1-pyrroline-5-carboxylate synthetase; P5CR, pyrroline-5-carboxylate reductase. -pyrroline-5-carboxylate synthetase (P5CS) enzymatic activity, respectively. Further analysis of polyamine oxidase (PAO)/diamine oxidase (DAO) polyamine catabolic enzymes indicated that DAO enzymatic activity increased significantly in correlation with putrescine accumulation, whereas PAO activity, involved in spermidine/spermine degradation, increased slightly. Moreover, transcriptional analysis of polyamine and proline metabolism genes (P5CS, P5CR, ADC, SPMS, SPDS, SAMDC, PAO, DAO) further supported the obtained data and revealed a complex SNP concentration-, time-, and developmental stage-dependent mechanism controlling endogenous proline and polyamine metabolite production. This is the first report to provide a global analysis leading to a better understanding of the role of the widely used NO donor SNP in the regulation of key stress-related metabolic pathways.