Relationship between H2O2 and jasmonic acid in pea leaf wounding response

The relationship of H₂O₂ and jasmonic acid (JA) in wound-induced defense response was investigated in the leaves of pea (Pisum sativum L.) plants. The results showed that both wounding and JA treatment led to a significant increase in activities of plasma membrane NADPH oxidase and phenylalanine ammonialyase. However, such an increase was blocked by the pretreatment with plasma membrane NADPH oxidase inhibitors, O ₂ ^? scavengers, or H₂O₂ scavenger, implying that H₂O₂ functions downstream of JA. Furthermore, wounding treatment activated two key enzymes of JA biosynthesis, lipoxygenase and allene oxide synthase, while JA biosynthetic inhibitors impaired the wounding-induced H₂O₂ burst. Thus, it is suggested that H₂O₂ burst depends on JA production in plant wounding response.     

Phloem exudate metabolic content reflects the response to water-deficit stress in pea plants (Pisum sativum L.)

Drought stress impacts the quality and yield of Pisum sativum. Here, we show how short periods of limited water availability during the vegetative stage of pea alters phloem sap content and how these changes are connected to strategies used by plants to cope with water deficit. We have investigated the metabolic content of phloem sap exudates and explored how this reflects P. sativum physiological and developmental responses to drought. Our data show that drought is accompanied by phloem-mediated redirection of the components that are necessary for cellular respiration and the proper maintenance of carbon/nitrogen balance during stress. The metabolic content of phloem sap reveals a shift from anabolic to catabolic processes as well as the developmental plasticity of P. sativum plants subjected to drought. Our study underlines the importance of phloem-mediated transport for plant adaptation to unfavourable environmental conditions. We also show that phloem exudate analysis can be used as a useful proxy to study stress responses in plants. We propose that the decrease in oleic acid content within phloem sap could be considered as a potential marker of early signalling events mediating drought response.     

Specificity of strain and genotype in the susceptibility of pea to Agrobacterium tumefaciens

Summary To determine the best combination for potential use in transformation of Pisum sativum L., 13 genotypes were inoculated with wild-type Agrobacterium tumefaciens strains A281, C58 and Ach5. A281 appeared to be the most virulent strain, as determined by size and number of tumours, followed by C58 and Ach5. Genotypes differed considerably in their response to inoculation and genotype x strain interaction was evident. Genotypes also responded differently to in vivo or in vitro inoculation. Axenic calli from tumours could be grown on hormone-free medium and the presence of the specific opines for each strain in the callus indicated successful transfer and expression of T-DNA. Southern blot analysis of DNA from callus of A281-inoculated material showed that both T_R and T_L T-DNA had been incorporated into the pea genome.NRCC No. 30265     

Replacement of nitrate by ammonium as the nitrogen source increases the salt sensitivity of pea plants. II. Inter- and intracellular solute compartmentation in leaflets

Pea plants (Pisum sativum L. cv. ‘Kleine Rheinländerin’) grown on ammonium or nitrate as the sole nitrogen source were treated with 50 mol m⁻³ NaCl. Four days after salt addition, ammonium-grown plants developed the first visible damage symptoms (wilting of leaflets, starting from the margins). Salt-treated, nitrate-grown plants were not affected during the experimental period. In order to obtain a better understanding of this differential salt sensitivity, we investigated the inter- and intracellular ion compartmentation of leaflets under both nutritional conditions by analysing ion concentrations in the apoplastic space, in chloroplasts and in protoplasts. When the leaves of nitrate- and ammonium-grown plants had attained similar sodium and chloride contents (after different times of exposure to salinity), the latter had a considerably lower chloroplastic chloride (and also sulphate) concentration. The results suggest that the intracellular compartmentation capacity of ammonium-grown plants is considerably lower than that of nitrate-grown plants. Ion toxicity appeared to initiate breakdown of metabolism in parts of the mesophyll tissue of ammonium-grown plants, causing an abrupt release of solutes into the apoplast, which coincided with the appearance of visible damage. Although the ammonium concentrations in leaves increased dramatically in the later phases of damage development, they were too low to cause the collapse of electrochemical gradients at the time at which damage became visible. Thus, the reason for a lower compartmentation capacity under ammonium nutrition remains as yet unclear.     

Benzothiadiazole and l-2-oxothiazolidine-4-carboxylic acid reduce the severity of Sharka symptoms in pea leaves: effect on antioxidative metabolism at the subcellular level

The effect of treatment with benzothiadiazole (BTH) or l -2-oxothiazolidine-4-carboxylic acid (OTC), and their interaction with Plum pox virus (PPV) infection, on antioxidative metabolism of pea plants was studied at the subcellular level. PPV infection produced a 20% reduction in plant growth. Pre-treatment of pea plants with OTC or BTH afforded partial protection against PPV infection, measured as the percentage of leaves showing symptoms, but neither BTH nor OTC significantly reduced the virus content. PPV infection caused oxidative stress, as monitored by increases in lipid peroxidation and protein oxidation in soluble and chloroplastic fractions. In leaves of non-infected plants, OTC increased the content of reduced glutathione (GSH) and total glutathione; accordingly, an increase in the redox state of glutathione was observed. An increase in oxidized glutathione (GSSG) was found in symptomatic leaves from infected plants. A similar increase in GSSG was also observed in asymptomatic leaves from infected, untreated plants. However, no changes in GSSG occurred in asymptomatic leaves from infected plants treated with BTH and OTC and, accordingly, a higher redox state of GSH was recorded in those leaves, which could have had a role in the reduction of symptoms, as observed in asymptomatic leaves from infected plants treated with BTH or OTC. Treatment with BTH or OTC had some effect on antioxidant enzymes in soluble and chloroplastic fractions from infected pea leaves. An increase in antioxidative mechanisms, such as GSH-related enzymes (DHAR, GR and G6PDH), as well as APX and POX, at the subcellular level was observed, which could play a role in reducing the severity of cellular damage induced by Sharka in pea leaves.     

Subcellular aspects of the protective effect of spermine against atrazine in pea plants

The action of the exogenously applied tetraamine spermine in reversing the effect of atrazine stress on intact pea plants (Pisum sativum L., cv. Koray) was investigated at the ultrastructural level. The results indicate that atrazine increases cell senescence by lipid peroxidation and loss of unsaturated fatty acids from thylakoid membranes of pea plant chloroplasts. Spermine acts as a polyfunctional effector. It stabilises the molecular composition of the membranes by preventing lipid peroxidation and as a consequence protects the conformation of the thylakoid system and structural integrity of the chloroplasts. Spermine treatment also contributes to the process of neutralisation of the free radicals by peroxisomes.     

Early stomatal closure in waterlogged pea plants is mediated by abscisic acid in the absence of foliar water deficits

Abstract Soil waterlogging decreased leaf conductance (interpreted as stomatal closure) of vegetative pea plants (Pisuin sativum L. cv. ‘Sprite’) approximately 24 h after the start of flooding, i.e. from the beginning of the second 16 h-long photo-period. Both adaxial and abaxial surfaces of leaves of various ages and the stipules were affected. Stomatal closure was sustained for at least 3 d with no decrease in foliar hydration measured as water content per unit area, leaf water potential or leaf water saturation deficit. Instead, leaves became increasingly hydrated in association with slower transpiration. These changes in the waterlogged plants over 3 d were accompanied by up to 10-fold increases in the concentration of endogenous abscisic acid (ABA). Waterlogging also increased foliar hydration and ABA concentrations in the dark. Leaves detached from non-waterlogged plants and maintained in vials of water for up to 3 d behaved in a similar way to leaves on flooded plants, i.e. stomata closed in the absence of a water deficit but in association with increased ABA content. Applying ABA through the transpiration stream to freshly detached leaflets partially closed stomata within 15 min. The extractable concentrations of ABA associated with this closure were similar to those found in flooded plants. When an ABA-deficient ‘wilty’ mutant of pea was waterlogged, the extent of stomatal closure was less pronounced than that in ordinary non-mutant plants, and the associated increase in foliar ABA was correspondingly smaller. Similarly, waterlogging closed stomata of tomato plants within 24 h, but no such closure was seen in ‘flacca’, a corresponding ABA-deficient mutant. The results provide an example of stomatal closure brought about by stress in the root environment in the absence of water deficiency. The correlative factor operating between the roots and shoots appeared to be an inhibition of ABA transport out of the shoots of flooded plants, causing the hormone to accumulate in the leaves.     

Up-regulation of the leaf mitochondrial and peroxisomal antioxidative systems in response to salt-induced oxidative stress in the wild salt-tolerant tomato species Lycopersicon pennellii

The response of the antioxidative systems of leaf cell mitochondria and peroxisomes of the cultivated tomato Lycopersicon esculentum (Lem) and its wild salt-tolerant related species Lycopersicon pennellii (Lpa) to NaCl 100 mM stress was investigated. Salt-dependent oxidative stress was evident in Lem mitochondria as indicated by their raised levels of lipid peroxidation and H2O2 content whereas their reduced ascorbate and reduced glutathione contents decreased. Concomitantly, SOD activity decreased whereas APX and GPX activities remained at control level. In contrast, the mitochondria of salt-treated Lpa did not exhibit salt-induced oxidative stress. In their case salinity induced an increase in the activities of superoxide dismutase (SOD), ascorbate peroxidase (APX), monodehydroascorbate reductase (MDHAR), dehydroascorbate reductase (DHAR) and glutathione-dependent peroxidase (GPX). Lpa peroxisomes exhibited increased SOD, APX, MDHAR and catalase activity and their lipid peroxidation and H2O2 levels were not affected by the salt treatment. The activities of all these enzymes remained at control level in peroxisomes of salt-treated Lem plants. The salt-induced increase in the antioxidant enzyme activities in the Lpa plants conferred cross-tolerance towards enhanced mitochondrial and peroxisomal reactive oxygen species production imposed by salicylhydroxamic acid (SHAM) and 3-amino-1,2,4-triazole (3-AT), respectively.     

Salicylic acid increases tolerance to oxidative stress induced by hydrogen peroxide accumulation in leaves of cadmium-exposed flax (Linum usitatissimum L.)

The aim of this study is to investigate the impacts of exogenous salicylic acid (SA) pretreatments on hydrogen peroxide (H₂O₂) accumulation, protein oxidation, and H₂O₂-scavenging enzymes in leaves of Cd-treated flax seedlings. Cd-enhanced H₂O₂ levels were related to increased activities of guaiacol peroxidase (POX, EC 1.11.1.7) and ascorbate peroxidase (APX, EC 1.11.1.11), and were independent of changes in catalase (CAT, EC 1.11.1.6) and superoxide dismutase (SOD, EC 1.15.1.1) activities. In control flax seedlings, exogenous SA pretreatments inhibited the activity of CAT, resulted in an enhanced production of H₂O₂ suggesting that SA requires H₂O₂ to initiate an oxidative stress. However, although leaves of Cd-free flax seedlings pretreated with SA accumulated in vivo H₂O₂ by 1.2-fold compared with leaves of Cd-only exposed ones; the damage to growth and proteins after the exposure to Cd was significantly less, indicating that SA can regulate the Cd-induced oxidative stress. Moreover, the Cd-treated seedlings primed with SA exhibited a higher level of total antioxidant capacities and increased activities of H₂O₂-detoxifying enzymes.     

Oxidative stress tolerance in plants: Novel interplay between auxin and reactive oxygen species signaling

Biotic and abiotic stress conditions produce reactive oxygen species (ROS) in plants causing oxidative stress damage. At the same time, ROS have additional signaling roles in plant adaptation to the stress. It is not known how the two seemingly contrasting functional roles of ROS between oxidative damage to the cell and signaling for stress protection are balanced. Research suggests that the plant growth regulator auxin may be the connecting link regulating the level of ROS and directing its role in oxidative damage or signaling in plants under stress. The objective of this review is to highlight some of the recent research on how auxin’s role is intertwined to that of ROS, more specifically H₂O₂, in plant adaptation to oxidative stress conditions.     

Antioxidative responses and proline level in leaves and roots of pea plants subjected to nickel stress

The activities of superoxide dismutase (SOD), ascorbate peroxidase (APX), catalase (CAT), glutathione S-transferase (GST) as well as proline content were studied in leaves and roots of 14 day-old pea plants treated with NiSO₄ (10, 100, 200 μm) for 1, 3, 6 and 9 days. Exposure of pea plants to nickel (Ni) resulted in the decrease in CuZnSOD as well as total SOD activities in both leaves and roots. The activity of APX in leaves of plants treated with 100 and 200 μm Ni increased following the 3rd day after metal application, while in roots at the end of the experiment the activity of this enzyme was significantly reduced. In both organs CAT activity generally did not change in response to Ni treatment. The activity of GST in plants exposed to high concentrations of Ni increased, more markedly in roots. In both leaves and roots after Ni application accumulation of free proline was observed, but in the case of leaves concentration of this amino acid increased earlier and to a greater extent than in roots. The results indicate that stimulation of GST activity and accumulation of proline in the tissues rather than antioxidative enzymes are involved in response of pea plants to Ni stress.     

Seed set,pollen morphology and pollen surface composition response to heat stress in field pea

Pea (Pisum sativum L.) is a major legume crop grown in a semi‐arid climate in Western Canada, where heat stress affects pollination, seed set and yield. Seed set and pod growth characteristics, along with in vitro percentage pollen germination, pollen tube growth and pollen surface composition, were measured in two pea cultivars (CDC Golden and CDC Sage) subjected to five maximum temperature regimes ranging from 24 to 36 °C. Heat stress reduced percentage pollen germination, pollen tube length, pod length, seed number per pod, and the seed–ovule ratio. Percentage pollen germination of CDC Sage was greater than CDC Golden at 36 °C. No visible morphological differences in pollen grains or the pollen surface were observed between the heat and control‐treated pea. However, pollen wall (intine) thickness increased due to heat stress. Mid‐infrared attenuated total reflectance (MIR‐ATR) spectra revealed that the chemical composition (lipid, proteins and carbohydrates) of each cultivar's pollen grains responded differently to heat stress. The lipid region of the pollen coat and exine of CDC Sage was more stable compared with CDC Golden at 36 °C. Secondary derivatives of ATR spectra indicated the presence of two lipid types, with different amounts present in pollen grains from each cultivar.     

Behaviour of antioxidant defense system in the adaptive response to salt stress in Helianthus annuus L. cells

A relationship between the antioxidant defense system and salt tolerance in two types of sunflower calli differing in salt sensitivity was studied. No reduction in growth occurred in the NaCl-salt-adapted cell line (T) when grown on 175 mM NaCl but growth of the salt-stressed cell line (S) was reduced by 83%. Lipid peroxidation and protein oxidation increased during acute stress of salt stressed cells at 14 and 28 d of the experiment, while salt-adapted calli (T) remained similar to non-shocked (C) values. The antioxidant defense system of callus adapted to growth under NaCl responded differently to 175 mM of salt compared with the corresponding controls under shock treatment. Salt-adapted and salt-stressed calli showed a similar pattern in GSH content at day 14 but at day 28 in S calli, GSH content was increased 100% over the non-shocked calli, while T calli returned to the initial values. In the salt-stressed calli, a general decrease in all the antioxidant enzymes studied (except for glutathione reductase and dehydroascorbate reductase activities) was observed at day 28. Except for catalase, the antioxidant enzymes were elevated constitutively in adapted calli as compared to stressed cells, when both were grown in the absence of NaCl (time 0), and remained unaltered until 28 d after the beginning of the experiment. These results suggest the involvement of an enzymatic antioxidant defense system in the adaptive response to salt stress in Helianthus annuus L. cells.     

Arbuscular mycorrhizal inoculation improves growth and antioxidative response of Jatropha curcas (L.) under Na2SO4 salt stress

This study investigated the effect of arbuscular mycorrhizal (AM) fungal consortia on growth, photosynthetic pigments, solutes concentration (e.g., sugars and proline), and antioxidant responses at different levels of Na₂SO₄ stress (0–0.5%, w:w) in potted culture of Jatropha. Results showed that increasing salt levels caused a significant reduction in survival (%), growth parameters, leaf relative water content (LRWC) (%), and chlorophyll content with an increase in electrolyte leakage (%) and lipid peroxidation of membranes of Jatropha. AM inoculation improved biomass yields as well as other physiological parameters (LRWC (%), chlorophyll, proline, and soluble sugar) of salt-stressed Jatropha over noninoculated plants. Tolerance index of Jatropha was higher with AM fungi than without at all salt levels; however, a decline in its value was recorded with increased salinity levels. AM inoculation also enhanced the activities of antioxidant enzymes (e.g., superoxide dismutase, peroxidase, ascorbate peroxidase, and glutathione reductase) and decreased oxidative damage to lipids. In conclusion, results indicate that AM inoculation was capable of alleviating the damage caused by salinity stress on Jatropha plants by reducing lipid peroxidation of membrane and membrane permeability and increasing the accumulation of solutes and antioxidant enzyme activity.