Tomato SlGSTU38 interacts with the PepMV coat protein and promotes viral infection.

Pepino mosaic virus (PepMV) is pandemic in tomato crops, causing important economic losses world-wide. No PepMV-resistant varieties have been developed yet. Identification of host factors interacting with PepMV proteins is a promising source of genetic targets to develop PepMV-resistant varieties. The interaction between the PepMV coat protein (CP) and the tomato glutathione S-transferase (GST) SlGSTU38 was identified in a yeast two-hybrid (Y2H) screening and validated by directed Y2H and co-immunoprecipitation assays. SlGSTU38-knocked-out Micro-Tom plants (gstu38) generated by the CRISPR/Cas9 technology together with live-cell imaging were used to understand the role of SlGSTU38 during infection. The transcriptomes of healthy and PepMV-infected wild-type (WT) and gstu38 plants were profiled by RNA-seq analysis. SlGSTU38 functions as a PepMV-specific susceptibility factor in a cell-autonomous manner and relocalizes to the virus replication complexes during infection. Besides, knocking out SlGSTU38 triggers reactive oxygen species accumulation in leaves and the deregulation of stress-responsive genes. SlGSTU38 may play a dual role: On the one hand, SlGSTU38 may exert a proviral function depending on its specific interaction with the PepMV CP; and on the other hand, SlGSTU38 may delay PepMV-infection sensing by participating in the redox intracellular homeostasis in a nonspecific manner.

Comparative metabolomic analysis between tomato and halophyte plants under intercropping conditions.

Halophyte-based intercropping appears nowadays as a valuable approach in soil remediation and agriculture. In this work, intercropping between the halophyte Arthrocaulon macrostachyum and tomato (Solanum lycopersicum var. Sargento) was studied in both plant species using comparative mass spectrometry-based metabolomics coupled to metabolic pathway predictions. A significant number of changes in metabolites was observed in the halophyte. In terms of alteration of specific metabolic pathways, intercropping conditions stimulated sugar and starch metabolisms in tomato, whereas in the halophyte, intercropping mainly altered amino acid-related pathways. In addition, arginine and proline metabolism were commonly affected in both tomato and halophyte plants. Moreover, metabolomic changes were associated with physiological alterations in tomato. In this sense, mild oxidative stress was induced in intercropped tomato plants, which, in turn, could trigger signaling events leading to plant adjustment to intercropping conditions. This study represents the first approach toward understanding intercropping interactions at the metabolome level and its effect on plant physiology, opening up prospects for further characterization of this crop cultivation strategy.

The apoplastic antioxidant system and altered cell wall dynamics influence mesophyll conductance and the rate of photosynthesis.

Mesophyll conductance (gm ), the diffusion of CO2 from substomatal cavities to the carboxylation sites in the chloroplasts, is a highly complex trait driving photosynthesis (net CO2 assimilation, AN ). However, little is known concerning the mechanisms by which it is dynamically regulated. The apoplast is considered as a 'key information bridge' between the environment and cells. Interestingly, most of the environmental constraints affecting gm also cause apoplastic responses, cell wall (CW) alterations and metabolic rearrangements. Since CW thickness is a key determinant of gm , we hypothesize that other changes in this cellular compartiment should also influence gm . We study the relationship between the antioxidant apoplastic system and CW metabolism and the gm responses in tobacco plants (Nicotiana sylvestris L.) under two abiotic stresses (drought and salinity), combining in vivo gas-exchange measurements with analyses of antioxidant activities, CW composition and primary metabolism. Stress treatments imposed substantial reductions in AN (58-54%) and gm (59%), accompanied by a strong antioxidant enzymatic response at the apoplastic and symplastic levels. Interestingly, apoplastic but not symplastic peroxidases were positively related to gm . Leaf anatomy remained mostly stable; however, the stress treatments significantly affected the CW composition, specifically pectins, which showed significant relationships with AN and gm . The treatments additionally promoted a differential primary metabolic response, and specific CW-related metabolites including galactose, glucosamine and hydroxycinnamate showed exclusive relationships with gm independent of the stress. These results suggest that gm responses can be attributed to specific changes in the apoplastic antioxidant system and CW metabolism, opening up more possibilities for improving photosynthesis using breeding/biotechnological strategies.

The Salt-Stress Response of the Transgenic Plum Line J8-1 and Its Interaction with the Salicylic Acid Biosynthetic Pathway from Mandelonitrile.

Salinity is considered as one of the most important abiotic challenges that affect crop productivity. Plant hormones, including salicylic acid (SA), are key factors in the defence signalling output triggered during plant responses against environmental stresses. We have previously reported in peach a new SA biosynthetic pathway from mandelonitrile (MD), the molecule at the hub of the cyanogenic glucoside turnover in Prunus sp. In this work, we have studied whether this new SA biosynthetic pathway is also present in plum and the possible role this pathway plays in plant plasticity under salinity, focusing on the transgenic plum line J8-1, which displays stress tolerance via an enhanced antioxidant capacity. The SA biosynthesis from MD in non-transgenic and J8-1 micropropagated plum shoots was studied by metabolomics. Then the response of J8-1 to salt stress in presence of MD or Phe (MD precursor) was assayed by measuring: chlorophyll content and fluorescence parameters, stress related hormones, levels of non-enzymatic antioxidants, the expression of two genes coding redox-related proteins, and the content of soluble nutrients. The results from in vitro assays suggest that the SA synthesis from the MD pathway demonstrated in peach is not clearly present in plum, at least under the tested conditions. Nevertheless, in J8-1 NaCl-stressed seedlings, an increase in SA was recorded as a result of the MD treatment, suggesting that MD could be involved in the SA biosynthesis under NaCl stress conditions in plum plants. We have also shown that the plum line J8-1 was tolerant to NaCl under greenhouse conditions, and this response was quite similar in MD-treated plants. Nevertheless, the MD treatment produced an increase in SA, jasmonic acid (JA) and reduced ascorbate (ASC) contents, as well as in the coefficient of non-photochemical quenching (qN) and the gene expression of Non-Expressor of Pathogenesis-Related 1 (NPR1) and thioredoxin H (TrxH) under salinity conditions. This response suggested a crosstalk between different signalling pathways (NPR1/Trx and SA/JA) leading to salinity tolerance in the transgenic plum line J8-1.

Metabolomics and Biochemical Approaches Link Salicylic Acid Biosynthesis to Cyanogenesis in Peach Plants.

Despite the long-established importance of salicylic acid (SA) in plant stress responses and other biological processes, its biosynthetic pathways have not been fully characterized. The proposed synthesis of SA originates from chorismate by two distinct pathways: the isochorismate and phenylalanine (Phe) ammonia-lyase (PAL) pathways. Cyanogenesis is the process related to the release of hydrogen cyanide from endogenous cyanogenic glycosides (CNglcs), and it has been linked to plant plasticity improvement. To date, however, no relationship has been suggested between the two pathways. In this work, by metabolomics and biochemical approaches (including the use of [13C]-labeled compounds), we provide strong evidences showing that CNglcs turnover is involved, at least in part, in SA biosynthesis in peach plants under control and stress conditions. The main CNglcs in peach are prunasin and amygdalin, with mandelonitrile (MD), synthesized from phenylalanine, controlling their turnover. In peach plants MD is the intermediary molecule of the suggested new SA biosynthetic pathway and CNglcs turnover, regulating the biosynthesis of both amygdalin and SA. MD-treated peach plants displayed increased SA levels via benzoic acid (one of the SA precursors within the PAL pathway). MD also provided partial protection against Plum pox virus infection in peach seedlings. Thus, we propose a third pathway, an alternative to the PAL pathway, for SA synthesis in peach plants.

Transformation of plum plants with a cytosolic ascorbate peroxidase transgene leads to enhanced water stress tolerance.

BACKGROUND AND AIMS: Water deficit is the most serious environmental factor limiting agricultural production. In this work, the tolerance to water stress (WS) of transgenic plum lines harbouring transgenes encoding cytosolic antioxidant enzymes was studied, with the aim of achieving the durable resistance of commercial plum trees. METHODS: The acclimatization process was successful for two transgenic lines: line C3-1, co-expressing superoxide dismutase (two copies) and ascorbate peroxidase (one copy) transgenes simultaneously; and line J8-1, harbouring four copies of the cytosolic ascorbate peroxidase gene (cytapx). Plant water relations, chlorophyll fluorescence and the levels of antioxidant enzymes were analysed in both lines submitted to moderate (7 d) and severe (15 d) WS conditions. Additionally, in line J8-1, showing the best response in terms of stress tolerance, a proteomic analysis and determination of the relative gene expression of two stress-responsive genes were carried out. KEY RESULTS: Line J8-1 exhibited an enhanced stress tolerance that correlated with better photosynthetic performance and a tighter control of water-use efficiency. Furthermore, this WS tolerance also correlated with a higher enzymatic antioxidant capacity than wild-type (WT) and line C3-1 plum plants. On the other hand, line C3-1 displayed an intermediate phenotype between WT plants and line J8-1 in terms of WS tolerance. Under severe WS, the tolerance displayed by J8-1 plants could be due to an enhanced capacity to cope with drought-induced oxidative stress. Moreover, proteomic analysis revealed differences between WT and J8-1 plants, mainly in terms of the abundance of proteins related to carbohydrate metabolism, photosynthesis, antioxidant defences and protein fate. CONCLUSIONS: The transformation of plum plants with cytapx has a profound effect at the physiological, biochemical, proteomic and genetic levels, enhancing WS tolerance. Although further experiments under field conditions will be required, it is proposed that J8-1 plants would be an interesting Prunus rootstock for coping with climate change.

Physiological and biochemical mechanisms of the ornamental Eugenia myrtifolia L. plants for coping with NaCl stress and recovery.

MAIN CONCLUSION: We studied the response of Eugenia myrtifolia L. plants, an ornamental shrub native to tropical and subtropical areas, to salt stress in order to facilitate the use of these plants in Mediterranean areas for landscaping. E. myrtifolia plants implement a series of adaptations to acclimate to salinity, including morphological, physiological and biochemical changes. Furthermore, the post-recovery period seems to be detected by Eugenia plants as a new stress situation. Different physiological and biochemical changes in Eugenia myrtifolia L. plants after being subjected to NaCl stress for up to 30 days (Phase I) and after recovery from salinity (Phase II) were studied. Eugenia plants proved to be tolerant to NaCl concentrations between 44 and 88 mM, displaying a series of adaptative mechanisms to cope with salt-stress, including the accumulation of toxic ions in roots. Plants increased their root/shoot ratio and decreased their leaf area, leaf water potential and stomatal conductance in order to limit water loss. In addition, they displayed different strategies to protect the photosynthetic machinery, including the limited accumulation of toxic ions in leaves, increase in chlorophyll content, changes in chlorophyll fluorescence parameters, leaf anatomy and antioxidant defence mechanisms. Anatomical modifications in leaves, including an increase in palisade parenchyma and intercellular spaces and decrease in spongy parenchyma, served to facilitate CO2 diffusion in a situation of reduced stomatal aperture. Salinity produced oxidative stress in Eugenia plants as evidenced by oxidative stress parameters values and a reduction in APX and ASC levels. Nevertheless, SOD and GSH contents increased. The post-recovery period is detected as a new stress situation, as observed through effects on plant growth and alterations in chlorophyll fluorescence and oxidative stress parameters.

Low glutathione regulates gene expression and the redox potentials of the nucleus and cytosol in Arabidopsis thaliana.

Reduced glutathione (GSH) is considered to exert a strong influence on cellular redox homeostasis and to regulate gene expression, but these processes remain poorly characterized. Severe GSH depletion specifically inhibited root meristem development, while low root GSH levels decreased lateral root densities. The redox potential of the nucleus and cytosol of Arabidopsis thaliana roots determined using roGFP probes was between -300 and -320 mV. Growth in the presence of the GSH-synthesis inhibitor buthionine sulfoximine (BSO) increased the nuclear and cytosolic redox potentials to approximately -260 mV. GSH-responsive genes including transcription factors (SPATULA, MYB15, MYB75), proteins involved in cell division, redox regulation (glutaredoxinS17, thioredoxins, ACHT5 and TH8) and auxin signalling (HECATE), were identified in the GSH-deficient root meristemless 1-1 (rml1-1) mutant, and in other GSH-synthesis mutants (rax1-1, cad2-1, pad2-1) as well as in the wild type following the addition of BSO. Inhibition of auxin transport had no effect on organ GSH levels, but exogenous auxin decreased the root GSH pool. We conclude that GSH depletion significantly increases the redox potentials of the nucleus and cytosol, and causes arrest of the cell cycle in roots but not shoots, with accompanying transcript changes linked to altered hormone responses, but not oxidative stress.

Sharka: how do plants respond to Plum pox virus infection?

Plum pox virus (PPV), the causal agent of sharka disease, is one of the most studied plant viruses, and major advances in detection techniques, genome characterization and organization, gene expression, transmission, and the description of candidate genes involved in PPV resistance have been described. However, information concerning the plant response to PPV infection is very scarce. In this review, we provide an updated summary of the research carried out to date in order to elucidate how plants cope with PPV infection and their response at different levels, including the physiological, biochemical, proteomic, and genetic levels. Knowledge about how plants respond to PPV infection can contribute to the development of new strategies to cope with this disease. Due to the fact that PPV induces an oxidative stress in plants, the bio-fortification of the antioxidative defences, by classical or biotechnological approaches, would be a useful tool to cope with PPV infection. Nevertheless, there are still some gaps in knowledge related to PPV-plant interaction that remain to be filled, such as the effect of PPV on the hormonal profile of the plant or on the plant metabolome.

Trichoderma harzianum T-78 supplementation of compost stimulates the antioxidant defence system in melon plants.

BACKGROUND: Compost is emerging as an alternative plant growing medium in efforts to achieve more sustainable agriculture. The addition of specific microorganisms such as Trichoderma harzianum to plant growth substrates increases yields and reduces plant diseases, but the mechanisms of such biostimulants and the biocontrol effects are not yet fully understood. In this work we investigated how the addition of citrus and vineyard composts, either alone or in combination with T. harzianum T-78, affects the antioxidant defence system in melon plants under nursery conditions. RESULTS: Compost application and/or Trichoderma inoculation modulated the antioxidant defence system in melon plants. The combination of citrus compost and Trichoderma showed a biostimulant effect that correlated with an increase in ascorbate recycling enzymes (monodehydroascorbate reductase, dehydroascorbate reductase) and peroxidase. Moreover, the inoculation of both composts with Trichoderma increased the activity of antioxidant enzymes, especially those involved in ascorbate recycling. CONCLUSION: Based on the long-established relationship between ascorbic acid and plant defence responses as well as plant growth and development, it can be suggested that ascorbate recycling activities play a major role in the protection provided by Trichoderma and its biostimulant effect and that these outcomes are linked to increases in antioxidant enzymes. We can conclude that the combination of citrus compost and T. harzianum T-78 constitutes a viable, environmentally friendly strategy for improving melon plant production.

Effect of halophyte-based management in physiological and biochemical responses of tomato plants under moderately saline greenhouse conditions.

Salinity, both in irrigation water and in soils, is one of the major abiotic constraints for agriculture activity worldwide. Phytodesalinization is a low-cost plant-based bioremediation strategy that can effectively amend salt-affected soils by cultivating salt tolerant plants. However, very few studies have evaluated the use of halophyte plants in crop management systems. In this work, we apply two different tomato crop management strategies involving the halophyte Arthrocaulon macrostachyum L. in a moderately saline soil: intercropping (mixed cultivation) and sequential cropping (cultivation of tomato where halophytes were previously grown). We investigated the effect of the different crop managements in some physiological and biochemical variables in tomato plants, including mineral nutrients content, photosynthesis, chlorophyll and flavonol contents, antioxidant metabolism and fruit production and quality. At soil level, both intercropping and sequential cropping decreased chloride content, sodium adsorption ratio and electrical conductivity, leading to reduced soil salinity. In tomato plants, halophyte-dependent management improved nutrient homeostasis and triggered a mild oxidative stress, whereas photosynthesis performance was enhanced by intercropping. In tomato fruits, the sequential cropping led to a 27% production increase and a slight decrease in the soluble sugar contents. We suggest the use of A. macrostachyum, and hence of halophyte plants, as an environmentally friendly phytoremediation strategy to improve plant performance while improving crop production, leading to a more sustainable agriculture and enhancing biodiversity.

Ectopic expression of cytosolic superoxide dismutase and ascorbate peroxidase leads to salt stress tolerance in transgenic plums.

To fortify the antioxidant capacity of plum plants, genes encoding cytosolic antioxidants ascorbate peroxidase (cytapx) and Cu/Zn-superoxide dismutase (cytsod) were genetically engineered in these plants. Transgenic plum plants expressing the cytsod and/or cytapx genes in cytosol have been generated under the control of the CaMV35S promoter. High levels of cytsod and cytapx gene transcripts suggested that the transgenes were constitutively and functionally expressed. We examined the potential functions of cytSOD and cytAPX in in vitro plum plants against salt stress (100 mm NaCl). Several transgenic plantlets expressing cytsod and/or cytapx showed an enhanced tolerance to salt stress, mainly lines C5-5 and J8-1 (expressing several copies of sod and apx, respectively). Transformation as well as NaCl treatments influenced the antioxidative metabolism of plum plantlets, including enzymatic and nonenzymatic antioxidants. Transgenic plantlets exhibited higher contents of nonenzymatic antioxidants glutathione and ascorbate than nontransformed control, which correlated with lower accumulation of hydrogen peroxide. Overall, our results suggest that transformation of plum plants with genes encoding antioxidant enzymes enhances the tolerance to salinity.

Chloroplast protection in plum pox virus-infected peach plants by L-2-oxo-4-thiazolidine-carboxylic acid treatments: effect in the proteome.

Sharka, a disease caused by plum pox virus (PPV), has a significant economic impact on fruit tree production. In this work, we analysed the effect of (2,1,3)-benzothiadiazole (BTH) and L-2-oxo-4-thiazolidine-carboxylic acid (OTC) on plant growth and virus content. OTC reduced sharka symptom, stimulated plant growth and alleviated PPV-induced oxidative stress, indicated by a lack of changes in some oxidative stress parameters. PPV infection reduced chloroplast electron transport efficiency. However, in the presence of BTH or OTC, no changes in the chlorophyll fluorescence parameters were observed. PPV produced an alteration in chloroplast ultrastructure, giving rise to a decrease in starch contents that was less dramatic in OTC-treated plants. Furthermore, PPV reduced the abundance of proteins associated with photosynthesis, carbohydrate and amino acid metabolism and photorespiration. These changes did not take place in OTC-treated plants, and increases in the expression of proteins related with the aforementioned processes, including ADP-glucose pyrophosphorylase, were produced, which correlated with the lower decrease in starch contents observed in PPV-infected plants treated with OTC. The results suggested that OTC treatment provides protection to the photosynthetic machinery and/or the chloroplast metabolism in PPV-infected peaches. Thus, OTC could have practical implications in agriculture in improving the vigour of different plant species as well as in immunizing plants against pathogens.

Antioxidant enzyme activities and hormonal status in response to Cd stress in the wetland halophyte Kosteletzkya virginica under saline conditions.

Salt marshes constitute major sinks for heavy metal accumulation but the precise impact of salinity on heavy metal toxicity for halophyte plant species remains largely unknown. Young seedlings of Kosteletzkya virginica were exposed during 3 weeks in nutrient solution to Cd 5 µM in the presence or absence of 50 mM NaCl. Cadmium (Cd) reduced growth and shoot water content and had major detrimental effect on maximum quantum efficiency (F(v) /F(m) ), effective quantum yield of photosystem II (Y(II)) and electron transport rates (ETRs). Cd induced an oxidative stress in relation to an increase in O(2) (•-) and H(2) O(2) concentration and lead to a decrease in endogenous glutathione (GSH) and α-tocopherol in the leaves. Cd not only increased leaf zeatin and zeatin riboside concentration but also increased the senescing compounds 1-aminocyclopropane-1-carboxylic acid (ACC) and abscisic acid (ABA). Salinity reduced Cd accumulation already after 1 week of stress but was unable to restore shoot growth and thus did not induce any dilution effect. Salinity delayed the Cd-induced leaf senescence: NaCl reduced the deleterious impact of Cd on photosynthesis apparatus through an improvement of F(v) /F(m) , Y(II) and ETR. Salt reduced oxidative stress in Cd-treated plants through an increase in GSH, α-tocopherol and ascorbic acid synthesis and an increase in glutathione reductase (EC 1.6.4.2) activity. Additional salt reduced ACC and ABA accumulation in Cd+NaCl-treated leaves comparing to Cd alone. It is concluded that salinity affords efficient protection against Cd to the halophyte species K. virginica, in relation to an improved management of oxidative stress and hormonal status.

Elucidating hormonal/ROS networks during seed germination: insights and perspectives.

While authors have traditionally emphasized the deleterious effects of reactive oxygen species (ROS) on seed biology, their role as signaling molecules during seed dormancy alleviation and germination is now the focus of many studies around the world. Over the last few years, studies using "-omics" technologies together with physiological and biochemical approaches have revealed that seed germination is a very complex process that depends on multiple biochemical and molecular variables. The pivotal role of phytohormones in promoting germination now appears to be interdependent with ROS metabolism, involving mitogen-activated protein kinase cascade activation, gene expression and post-translational protein modifications. This review is, thus, an attempt to summarize the new discoveries involving ROS and seed germination. The study of these interactions may supply markers of seed quality that might eventually be used in breeding programs to improve crop yields.

Understanding the role of H(2)O(2) during pea seed germination: a combined proteomic and hormone profiling approach.

In a previous publication, we showed that the treatment of pea seeds in the presence of hydrogen peroxide (H(2)O(2)) increased germination performance as well as seedling growth. To gain insight into the mechanisms responsible for this behaviour, we have analysed the effect of treating mature pea seeds in the presence of 20 mm H(2)O(2) on several oxidative features such as protein carbonylation, endogenous H(2)O(2) and lipid peroxidation levels. We report that H(2)O(2) treatment of the pea seeds increased their endogenous H(2)O(2) content and caused carbonylation of storage proteins and of several metabolic enzymes. Under the same conditions, we also monitored the expression of two MAPK genes known to be activated by H(2)O(2) in adult pea plants. The expression of one of them, PsMAPK2, largely increased upon pea seed imbibition in H(2)O(2) , whereas no change could be observed in expression of the other, PsMAPK3. The levels of several phytohormones such as 1-aminocyclopropane carboxylic acid, indole-3-acetic acid and zeatin appeared to correlate with the measured oxidative indicators and with the expression of PsMAPK2. Globally, our results suggest a key role of H(2)O(2) in the coordination of pea seed germination, acting as a priming factor that involves specific changes at the proteome, transcriptome and hormonal levels.

A nuclear glutathione cycle within the cell cycle.

The complex antioxidant network of plant and animal cells has the thiol tripeptide GSH at its centre to buffer ROS (reactive oxygen species) and facilitate cellular redox signalling which controls growth, development and defence. GSH is found in nearly every compartment of the cell, including the nucleus. Transport between the different intracellular compartments is pivotal to the regulation of cell proliferation. GSH co-localizes with nuclear DNA at the early stages of proliferation in plant and animal cells. Moreover, GSH recruitment and sequestration in the nucleus during the G1- and S-phases of the cell cycle has a profound impact on cellular redox homoeostasis and on gene expression. For example, the abundance of transcripts encoding stress and defence proteins is decreased when GSH is sequestered in the nucleus. The functions of GSHn (nuclear GSH) are considered in the present review in the context of whole-cell redox homoeostasis and signalling, as well as potential mechanisms for GSH transport into the nucleus. We also discuss the possible role of GSHn as a regulator of nuclear proteins such as histones and PARP [poly(ADP-ribose) polymerase] that control genetic and epigenetic events. In this way, a high level of GSH in the nucleus may not only have an immediate effect on gene expression patterns, but also contribute to how cells retain a memory of the cellular redox environment that is transferred through generations.

H2O2-Elicitation of Black Carrot Hairy Roots Induces a Controlled Oxidative Burst Leading to Increased Anthocyanin Production.

Hairy roots (HRs) grown in vitro are a powerful platform for plant biotechnological advances and for the bio-based production of metabolites of interest. In this work, black carrot HRs able to accumulate anthocyanin as major secondary metabolite were used. Biomass and anthocyanin accumulation were improved by modulating growth medium composition-different Murashige & Skoog (MS)-based media-and H2O2-elicitation, and the level of the main antioxidant enzymes on elicited HRs was measured. Higher growth was obtained on liquid 1/2 MS medium supplemented with 60 g/L sucrose for HRs grown over 20 days. In this medium, 200 µM H2O2 applied on day 12 induced anthocyanin accumulation by 20%. The activity of superoxide dismutase (SOD)-which generates H2O2 from O2•--increased by over 50%, whereas the activity of H2O2-scavenging enzymes was not enhanced. Elicitation in the HRs can result in a controlled oxidative burst, in which SOD activity increased H2O2 levels, whereas anthocyanins, as effective reactive oxygen species scavengers, could be induced to modulate the oxidative burst generated. Moreover, given the proven stability of the HR lines used and their remarkable productivity, this system appears as suitable for elucidating the interplay between antioxidant and secondary metabolism.

Interplay among Antioxidant System, Hormone Profile and Carbohydrate Metabolism during Bud Dormancy Breaking in a High-Chill Peach Variety.

(1) Background: Prunus species have the ability to suspend (induce dormancy) and restart growth, in an intricate process in which environmental and physiological factors interact. (2) Methods: In this work, we studied the evolution of sugars, antioxidant metabolism, and abscisic acid (ABA) and gibberellins (GAs) levels during bud dormancy evolution in a high-chill peach variety, grown for two seasons in two different geographical areas with different annual media temperature, a cold (CA) and a temperate area (TA). (3) Results: In both areas, starch content reached a peak at ecodormancy, and then decreased at dormancy release (DR). Sorbitol and sucrose declined at DR, mainly in the CA. In contrast, glucose and fructose levels progressively rose until DR. A decline in ascorbate peroxidase, dehydroascorbate reductase, superoxide dismutase and catalase activities occurred in both seasons at DR. Moreover, the H2O2-sensitive SOD isoenzymes, Fe-SOD and Cu,Zn-SOD, and two novel peroxidase isoenzymes, were detected. Overall, these results suggest the occurrence of a controlled oxidative stress during DR. GA7 was the major bioactive GA in both areas, the evolution of its levels being different between seasons and areas. In contrast, ABA content decreased during the dormancy period in both areas, resulting in a reduction in the ABA/total GAs ratio, being more evident in the CA. (4) Conclusion: A possible interaction sugars-hormones-ROS could take place in high-chill peach buds, favoring the DR process, suggesting that, in addition to sugar metabolism, redox interactions can govern bud DR, regardless of chilling requirements.

Oxidative stress induced in tobacco leaves by chloroplast over-expression of maize plastidial transglutaminase.

As part of a project aiming to characterize the role of maize plastidial transglutaminase (chlTGZ) in the plant chloroplast, this paper presents results on stress induced by continuous chlTGZ over-expression in transplastomic tobacco leaves. Thylakoid remodelling induced by chlTGZ over-expression in young leaves of tobacco chloroplasts has already been reported (Ioannidis et al. in Biochem Biophys Acta 1787:1215-1222, 2009). In the present work, we determined the induced alterations in the photosynthetic apparatus, in the chloroplast ultrastructure, and, particularly, the activation of oxidative and antioxidative metabolism pathways, regarding ageing and functionality of the tobacco transformed plants. The results revealed that photochemistry impairment and oxidative stress increased with transplastomic leaf age. The decrease in pigment levels in the transformed leaves was accompanied by an increase in H(2)O(2) and lipid peroxidation. The rise in H(2)O(2) correlated with a decrease in catalase activity, whereas there was an increase in peroxidase activity. In addition, chlTGZ over-expression lead to a drop in reduced glutathione, while Fe-superoxide dismutase activity was higher in transformed than in wild-type leaves. Together with the induced oxidative stress, the over-expressed chlTGZ protein accumulated progressively in chloroplast inclusion bodies. These traits were accompanied by thylakoid scattering, membrane degradation and reduction of thylakoid interconnections. Consequently, the electron transport between photosystems decrease in the old leaves. In spite of these alterations, transplastomic plants can be maintained and reproduced in vitro. These results are discussed in line with chlTGZ involvement in chloroplast functionality.