Indole-3-acetaldoxime delays root iron-deficiency responses and modify auxin homeostasis in Medicago truncatula.

Iron (Fe) is an essential plant micronutrient, being a major limiting growth factor in calcareous soils. To increase Fe uptake, plants induce lateral roots growth, the expression of a Fe(III)-chelate reductase (FCR), a Fe(II)-transporter and a H+-ATPase and the secretion of flavins. Furthermore, auxin hormone family is involved in the Fe-deficiency responses but the action mechanism remains elusive. In this work, we evaluated the effect of the auxin-precursor indole-3-acetaldoxime (IAOx) on hydroponically grown Medicago truncatula plants under different Fe conditions. Upon 4-days of Fe starvation, the pH of the nutrient solution decreased, while both the FCR activity and the presence of flavins increased. Exogenous IAOx increased lateral roots growth contributing to superroot phenotype, decreased chlorosis, and delayed up to 3-days the pH-decrease, the FCR-activity increase, and the presence of flavins, compared to Fe-deficient plants. Gene expression levels were in concordance with the physiological responses. RESULTS: showed that IAOx was immediately transformed to IAN in roots and shoots to maintain auxin homeostasis. IAOx plays an active role in iron homeostasis delaying symptoms and responses in Fe-deficient plants. We may speculate that IAOx or its derivatives remobilize Fe from root cells to alleviate Fe-deficiency. Overall, these results point out that the IAOx-derived phenotype may have advantages to overcome nutritional stresses.

The importance of the urea cycle and its relationships to polyamine metabolism during ammonium stress in Medicago truncatula.

The ornithine-urea cycle (urea cycle) makes a significant contribution to the metabolic responses of lower photosynthetic eukaryotes to episodes of high nitrogen availability. In this study, we compared the role of the plant urea cycle and its relationships to polyamine metabolism in ammonium-fed and nitrate-fed Medicago truncatula plants. High ammonium resulted in the accumulation of ammonium and pathway intermediates, particularly glutamine, arginine, ornithine, and putrescine. Arginine decarboxylase activity was decreased in roots, suggesting that the ornithine decarboxylase-dependent production of putrescine was important in situations of ammonium stress. The activity of copper amine oxidase, which releases ammonium from putrescine, was significantly decreased in both shoots and roots. In addition, physiological concentrations of ammonium inhibited copper amine oxidase activity in in vitro assays, supporting the conclusion that high ammonium accumulation favors putrescine synthesis. Moreover, early supplementation of plants with putrescine avoided ammonium toxicity. The levels of transcripts encoding urea-cycle-related proteins were increased and transcripts involved in polyamine catabolism were decreased under high ammonium concentrations. We conclude that the urea cycle and associated polyamine metabolism function as important protective mechanisms limiting ammonium toxicity in M. truncatula. These findings demonstrate the relevance of the urea cycle to polyamine metabolism in higher plants.

Volatile compounds other than CO2 emitted by different microorganisms promote distinct posttranscriptionally regulated responses in plants.

A "box-in-box" cocultivation system was used to investigate plant responses to microbial volatile compounds (VCs) and to evaluate the contributions of organic and inorganic VCs (VOCs and VICs, respectively) to these responses. Arabidopsis plants were exposed to VCs emitted by adjacent Alternaria alternata and Penicillium aurantiogriseum cultures, with and without charcoal filtration. No VOCs were detected in the headspace of growth chambers containing fungal cultures with charcoal filters. However, these growth chambers exhibited elevated CO2 and bioactive CO and NO headspace concentrations. Independently of charcoal filtration, VCs from both fungal phytopathogens promoted growth and distinct developmental changes. Plants cultured at CO2 levels observed in growth boxes containing fungal cultures were identical to those cultured at ambient CO2 . Plants exposed to charcoal-filtered fungal VCs, nonfiltered VCs, or superelevated CO2 levels exhibited transcriptional changes resembling those induced by increased irradiance. Thus, in the "box-in-box" system, (a) fungal VICs other than CO2 and/or VOCs not detected by our analytical systems strongly influence the plants' responses to fungal VCs, (b) different microorganisms release VCs with distinct action potentials, (c) transcriptional changes in VC-exposed plants are mainly due to enhanced photosynthesis signaling, and (d) regulation of some plant responses to fungal VCs is primarily posttranscriptional.

Review: Mechanisms of ammonium toxicity and the quest for tolerance.

Ammonium sensitivity of plants is a worldwide problem, constraining crop production. Prolonged application of ammonium as the sole nitrogen source may result in physiological and morphological disorders that lead to decreased plant growth and toxicity. The main causes of ammonium toxicity/tolerance described until now include high ammonium assimilation by plants and/or low sensitivity to external pH acidification. The various ammonium transport-related components, especially the non-electrogenic influx of NH3 (related to the depletion of (15)N) and the electrogenic influx of NH4(+), may contribute to ammonium accumulation, and therefore to NH3 toxicity. However, this accumulation may be influenced by increasing K(+) concentration in the root medium. Recently, new insights have been provided by "omics" studies, leading to a suggested involvement of GDP mannose-pyrophosphorylase in the response pathways of NH4(+) stress. In this review, we highlight the cross-talk signaling between nitrate, auxins and NO, and the importance of the connection of the plants' urea cycle to metabolism of polyamines. Overall, the tolerance and amelioration of ammonium toxicity are outlined to improve the yield of ammonium-grown plants. This review identifies future directions of research, focusing on the putative importance of aquaporins in ammonium influx, and on genes involved in ammonium sensitivity and tolerance.

Two Fe-superoxide dismutase families respond differently to stress and senescence in legumes.

Three main families of SODs in plants may be distinguished according to the metal in the active center: CuZnSODs, MnSOD, and FeSOD. CuZnSODs have two sub-families localized either in plant cell cytosol or in plastids, the MnSOD family is essentially restricted to mitochondria, and the FeSOD enzyme family has been typically localized into the plastid. Here, we describe, based on a phylogenetic tree and experimental data, the existence of two FeSOD sub-families: a plastidial localized sub-family that is universal to plants, and a cytosolic localized FeSOD sub-family observed in determinate-forming nodule legumes. Anti-cytosolic FeSOD (cyt_FeSOD) antibodies were employed, together with a novel antibody raised against plastidial FeSOD (p_FeSOD). Stress conditions, such as nitrate excess or drought, markedly increased cyt_FeSOD contents in soybean tissues. Also, cyt_FeSOD content and activity increased with age in both soybean and cowpea plants, while the cyt_CuZnSOD isozyme was predominant during early stages. p_FeSOD in leaves decreased with most of the stresses applied, but this isozyme markedly increased with abscisic acid in roots. The great differences observed for p_FeSOD and cyt_FeSOD contents in response to stress and aging in plant tissues reveal distinct functionality and confirm the existence of two immunologically differentiated FeSOD sub-families. The in-gel FeSOD activity patterns showed a good correlation to cyt_FeSOD contents but not to those of p_FeSOD. This indicates that cyt_FeSOD is the main active FeSOD in soybean and cowpea tissues. The diversity of functions associated with the complexity of FeSOD isoenzymes depending of the location is discussed.

High irradiance increases NH(4)(+) tolerance in Pisum sativum: Higher carbon and energy availability improve ion balance but not N assimilation.

The widespread use of NO(3)(-) fertilization has had a major ecological impact. NH(4)(+) nutrition may help to reduce this impact, although high NH(4)(+) concentrations are toxic for most plants. The underlying tolerance mechanisms are not yet fully understood, although they are thought to include the limitation of C, the disruption of ion homeostasis, and a wasteful NH(4)(+) influx/efflux cycle that carries an extra energetic cost for root cells. In this study, high irradiance (HI) was found to induce a notable tolerance to NH(4)(+) in the range 2.5-10mM in pea plants by inducing higher C availability, as shown by carbohydrate content. This capacity was accompanied by a general lower relative N content, indicating that tolerance is not achieved through higher net N assimilation on C-skeletons, and it was also not attributable to increased GS content or activity in roots or leaves. Moreover, HI plants showed higher ATP content and respiration rates. This extra energy availability is related to the internal NH(4)(+) content regulation (probably NH(4)(+) influx/efflux) and to an improvement of the cell ionic balance. The limited C availability at lower irradiance (LI) and high NH(4)(+) resulted in a series of metabolic imbalances, as reflected in a much higher organic acid content, thereby suggesting that the origin of the toxicity in plants cultured at high NH(4)(+) and LI is related to their inability to avoid large-scale accumulation of the NH(4)(+) ion.

Short term physiological implications of NBPT application on the N metabolism of Pisum sativum and Spinacea oleracea.

The application of urease inhibitors in conjunction with urea fertilizers as a means of reducing N loss due to ammonia volatilization requires an in-depth study of the physiological effects of these inhibitors on plants. The aim of this study was to determine how the urease inhibitor N-(n-butyl) thiophosphoric triamide (NBPT) affects N metabolism in pea and spinach. Plants were cultivated in pure hydroponic culture with urea as the sole N source. After 2 weeks of growth for pea, and 3 weeks for spinach, half of the plants received NBPT in their nutrient solution. Urease activity, urea and ammonium content, free amino acid composition and soluble protein were determined in leaves and roots at days 0, 1, 2, 4, 7 and 9, and the NBPT content in these tissues was determined 48h after inhibitor application. The results suggest that the effects of NBPT on spinach and pea urease activity differ, with pea being most affected by this treatment, and that the NBPT absorbed by the plant caused a clear inhibition of the urease activity in pea leaf and roots. The high urea concentration observed in leaves was associated with the development of necrotic leaf margins, and was further evidence of NBPT inhibition in these plants. A decrease in the ammonium content in roots, where N assimilation mainly takes place, was also observed. Consequently, total amino acid contents were drastically reduced upon NBPT treatment, indicating a strong alteration of the N metabolism. Furthermore, the amino acid profile showed that amidic amino acids were major components of the reduced pool of amino acids. In contrast, NBPT was absorbed to a much lesser degree by spinach plants than pea plants (35% less) and did not produce a clear inhibition of urease activity in this species.

High irradiance induces photoprotective mechanisms and a positive effect on NH4+ stress in Pisum sativum L.

Photosynthesis provides plant metabolism with reduced carbon (C) but is also the main source of oxidative stress in plants. Likewise, high doses of NH(4)(+) as sole N source have been reported to be toxic for most plants, resulting in reduced plant growth and restricting C availability. The combination of high photosynthetic photon flux densities (PPFD) and NH(4)(+) nutrition may provide higher C availability but could also have a detrimental effect on the plants, therefore the objective of this study is to evaluate whether NH(4)(+) induces photo-oxidative stress that is exacerbated under high light conditions. Pea plants (Pisum sativum cv. sugar-snap) were grown hydroponically with NH(4)(+) (0.5, 2.5, 5 and 10 mM) under high (750 micromol photons m(-2)s(-1)) or low PPFD conditions (350 micromol photons m(-2)s(-1)). High PPFD contributes to a higher tolerance to ammonium by pea plants, as it originated higher biomass content due to higher photosynthetic rates. However, a deficit of N (0.5 and 2.5 mM NH(4)(+)) under high PPFD conditions caused an antioxidant response, as indicated by increased photoprotective pigment and chloroplastic superoxide dismutase contents. Plants grown with higher doses of N and high PPFD showed less need for photoprotection. An increase in the specific leaf weight (SLW) ratio was observed associated not only with high PPFDs but also with the highest NH(4)(+) dose. Overall, these results demonstrate that, despite the activation of some photoprotective responses at high PPFD, there were no photoinhibitory symptoms and a positive effect on NH(4)(+) toxicity, thus suggesting that the harmful effects of NH(4)(+) are not directly related to the generation of photo-oxidative stress.

Nitrogen nutrition and antioxidant metabolism in ammonium-tolerant and -sensitive plants.

Ammonium nutrition is of interest as an alternative to that of using nitrate. However, the former has been reported as stressful to many plant species especially to some important crops, as most abiotic stresses may trigger oxidative imbalances in plants. In this work, we investigate the response of oxidative metabolism of two plant species, spinach (Spinacia oleracea L. cv. Gigante de invierno) and pea (Pisum sativum L. cv. Rondo), which have distinct tolerance to ammonium. Plants were grown in the presence of 1.5 and 3.0 mM N as ammonium and compared with equivalent nitrate nutrition. The antioxidant enzymes and metabolites as well as oxidative damage to proteins were determined. Protein and amino acid contents in both types of plants were also analysed. Ammonium nutrition in sensitive spinach or in the tolerant pea plants does not alter the redox status of ascorbate and glutathione or the phenolic contents, while no clear effect is seen in the antioxidant enzymes. The results showed that the stress originated from applying ammonium as the only N source is not an oxidative stress, independent of the ammonium tolerance of the plant species studied. Moreover, ammonium stress diminishes oxidative damage to proteins in the spinach plants. The data of the protein oxidation together with those from N metabolism highlight the relation between the stress induced by ammonium and an increased protein turnover.

Crystallization and preliminary X-ray diffraction studies of the eukaryotic iron superoxide dismutase (FeSOD) from Vigna unguiculata.

Eukaryotic iron superoxide dismutases (FeSODs) are homodimeric proteins that constitute a fundamental protection against free radicals, which can damage essential cellular mechanisms. The protein was cloned and overexpressed in Escherichia coli with an N-terminal His tag. Crystallization experiments of the protein resulted, after several refined screenings, in crystals suitable for X-ray diffraction analysis. The crystals belong to the monoclinic space group C2, with unit-cell parameters a = 82.54, b = 48.41, c = 64.28 A, alpha = gamma = 90, beta = 119.66 degrees, and contain one molecule per asymmetric unit. At cryogenic temperatures, the crystals diffracted to a resolution limit of 1.80 A using synchrotron radiation at the European Synchrotron Radiation Facility (ESRF).