The effect of low ascorbic acid content on tomato fruit ripening.

MAIN CONCLUSION: The oxidant/antioxidant balance affects the ripening time of tomato fruit. Ripening of tomato fruit is associated with several modifications such as loss of cell wall firmness and transformation of chloroplasts to chromoplasts. Besides a peak in H2O2, reactive oxygen species (ROS) are observed at the transition stage. However, the role of different components of oxidative stress metabolism in fruit ripening has been scarcely addressed. Two GDP-L-galactose phosphorylase (GGP) Solanum lycopersicum L. cv Micro-Tom mutants which have fruit with low ascorbic acid content (30% of wild type) were used in this work to unravel the participation of ascorbic acid and H2O2 in fruit maturation. Both GGP mutants show delayed fruit maturation with no peak of H2O2; treatment with ascorbic acid increases its own concentration and accelerates ripening only in mutants to become like wild type plants. Unexpectedly, the treatment with ascorbic acid increases H2O2 synthesis in both mutants resembling what is observed in wild type fruit. Exogenous supplementation with H2O2 decreases its own synthesis delaying fruit maturation in plants with low ascorbic acid content. The site of ROS production is localized in the chloroplasts of fruit of all genotypes as determined by confocal microscopy analysis. The results presented here demonstrate that both ascorbic acid and H2O2 actively participate in tomato fruit ripening.

Plasticity of root growth and respiratory activity: Root responses to above-ground senescence, fruit removal or partial root pruning in soybean.

This work focuses on the alterations in soybean root growth and activity during whole plant senescence and the contribution of roots to source-sink relations during plant development. The experiments were designed to analyze the activity of roots in relation to: a) whole plant senescence, b) total pod removal and c) root pruning (15, 25 and 50% of DW) during seed growth stages. Roots can grow until an advanced R5 stage and their specific activity decreases along the reproductive development but whole root activity declines from R6. However root respiration is maintained at a basal level until R8. Depodded plants showed a large increase of root dry matter (about 470%) and a large increase of AOX protein. Root pruning treatments showed a proportional increase of specific root respiration in 25 and 50% treatments but no differences of whole root respiration and dry matter partitioning at R7. These results indicate that roots are under the control of the requirements of above ground organs until final stages of seed growth but, after this, roots may survive independently for some time. This suggests that roots do not suffer a senescence-like process as leaves do. Also, plants have a high capacity to buffer changes in root biomass production and specific root activity under pod removal or partial root pruning.

Qualitative and quantitative modifications of root mitochondria during senescence of above-ground parts of Arabidopis thaliana.

This work studied modifications experienced by root mitochondria during whole plant senescence or under light deprivation, using Arabidopsis thaliana plants with YFP tagged to mitochondria. During post-bolting development, root respiratory activity started to decline after aboveground organs (i.e., rosette leaves) had senesced. This suggests that carbohydrate starvation may induce root senescence. Similarly, darkening the whole plant induced a decrease in respiration of roots. This was partially due to a decrease in the number of total mitochondria (YFP-labelled mitochondria) and most probably to a decrease in the quantity of mitochondria with a developed inner membrane potential (ΔΨm, i.e., Mitotracker red- labelled mitochondria). Also, the lower amount of mitochondria with ΔΨm compared to YFP-labelled mitochondria at 10d of whole darkened plant, suggests the presence of mitochondria in a "standby state". The experiments also suggest that small mitochondria made the main contribution to the respiratory activity that was lost during root senescence. Sugar supplementation partially restored the respiration of mitochondria after 10d of whole plant dark treatment. These results suggest that root senescence is triggered by carbohydrate starvation, with loss of ΔΨm mitochondria and changes in mitochondrial size distribution.

Heat stress induces ferroptosis-like cell death in plants.

In plants, regulated cell death (RCD) plays critical roles during development and is essential for plant-specific responses to abiotic and biotic stresses. Ferroptosis is an iron-dependent, oxidative, nonapoptotic form of cell death recently described in animal cells. In animal cells, this process can be triggered by depletion of glutathione (GSH) and accumulation of lipid reactive oxygen species (ROS). We investigated whether a similar process could be relevant to cell death in plants. Remarkably, heat shock (HS)-induced RCD, but not reproductive or vascular development, was found to involve a ferroptosis-like cell death process. In root cells, HS triggered an iron-dependent cell death pathway that was characterized by depletion of GSH and ascorbic acid and accumulation of cytosolic and lipid ROS. These results suggest a physiological role for this lethal pathway in response to heat stress in Arabidopsis thaliana The similarity of ferroptosis in animal cells and ferroptosis-like death in plants suggests that oxidative, iron-dependent cell death programs may be evolutionarily ancient.

MiR393 regulation of auxin signaling and redox-related components during acclimation to salinity in Arabidopsis.

One of the most striking aspects of plant plasticity is the modulation of development in response to environmental changes. Plant growth and development largely depend on the phytohormone auxin that exerts its function through a partially redundant family of F-box receptors, the TIR1-AFBs. We have previously reported that the Arabidopsis double mutant tir1 afb2 is more tolerant to salt stress than wild-type plants and we hypothesized that down-regulation of auxin signaling might be part of Arabidopsis acclimation to salinity. In this work, we show that NaCl-mediated salt stress induces miR393 expression by enhancing the transcription of AtMIR393A and leads to a concomitant reduction in the levels of the TIR1 and AFB2 receptors. Consequently, NaCl triggers stabilization of Aux/IAA repressors leading to down-regulation of auxin signaling. Further, we report that miR393 is likely involved in repression of lateral root (LR) initiation, emergence and elongation during salinity, since the mir393ab mutant shows reduced inhibition of emergent and mature LR number and length upon NaCl-treatment. Additionally, mir393ab mutant plants have increased levels of reactive oxygen species (ROS) in LRs, and reduced ascorbate peroxidase (APX) enzymatic activity compared with wild-type plants during salinity. Thus, miR393 regulation of the TIR1 and AFB2 receptors could be a critical checkpoint between auxin signaling and specfic redox-associated components in order to coordinate tissue and time-specific growth responses and tolerance during acclimation to salinity in Arabidopsis.

Impact of brassinosteroids and ethylene on ascorbic acid accumulation in tomato leaves.

Plant steroid hormones brassinosteroids (BRs) and the gaseous hormone ethylene (ET) alter the ascorbic acid-glutathione (AA-GSH) levels in tomato (Solanum lycopersicum L.) plants. The interaction of these hormones in regulating antioxidant metabolism is however unknown. The combined use of genetics (BR-mutants) and chemical application (BR/ET-related chemicals) shows that BRs and ET signalling pathways interact, to regulate leaf AA content and synthesis. BR-deficient (d(x)) leaves display low total AA but BR-accumulating (35S:D) leaves show normal total AA content. Leaves with either BR levels lower or higher than wild type plants showed a higher oxidised AA redox state. The activity of L-galactono-1,4-lactone dehydrogenase (L-GalLDH), the mitochondrial enzyme that catalyses the last step in AA synthesis is lower in d(x) and higher in 35S:D plants. BR-deficient mutants show higher ET production but it is restored to normal levels when BR content is increased in 35S:D plants. Suppression of ET signalling using 1-methylcyclopropene in d(x) and 35S:D plants restored leaf AA content and L-GalLDH activity, to the values observed in wild type. The suppression of ET action in d(x) and 35S:D leaves leads to the respective decreasing and increasing respiration, indicating an opposite response compared to AA synthesis. This inverse relationship is lacking in ET suppressed d(x) plants in response to external BRs. The modifications in the in vivo activity of L-GalLDH activity do not correlate with changes in the level of the enzyme. Taken together, these data suggest that ET suppresses and BRs promote AA synthesis and accumulation.

Glutathione and ascorbic acid protect Arabidopsis plants against detrimental effects of iron deficiency.

Iron is an essential micronutrient required for a wide variety of cellular functions in plant growth and development. Chlorosis is the first visible symptom in iron-deficient plants. Glutathione (GSH) and ascorbic acid (ASC) are multifunctional metabolites playing important roles in redox balancing. In this work, it was shown that GSH and ASC treatment prevented chlorosis and the accumulation of reactive oxygen species induced by iron deficiency in Arabidopsis leaves. In iron deficiency, GSH and ASC increased the activity of the heme protein ascorbate peroxidase at a similar level to that found in iron-sufficient seedlings. GSH was also able to preserve the levels of the iron-sulfur protein ferredoxin 2. GSH content decreased 25% in iron-deficient Arabidopsis seedlings, whereas the ASC levels were not affected. Taken together, these results showed that GSH and ASC supplementation protects Arabidopsis seedlings from iron deficiency, preserving cell redox homeostasis and improving internal iron availability.

Auxin signaling participates in the adaptative response against oxidative stress and salinity by interacting with redox metabolism in Arabidopsis.

Auxin regulates gene expression through direct physical interaction with TIR1/AFB receptor proteins during different processes of growth and development in plants. Here we report the contribution of auxin signaling pathway to the adaptative response against abiotic stress in Arabidopsis. Phenotypic characterization of tir1/afb auxin receptor mutants indicates a differential participation of each member under abiotic stress. In particular, tir1 afb2 and tir1 afb3 mutants resulted more tolerant to oxidative stress. In addition, tir1 afb2 showed increased tolerance against salinity measured as chlorophyll content, germination rate and root elongation compared with wild-type plants. Furthermore, tir1 afb2 displayed a reduced accumulation of hydrogen peroxide and superoxide anion, as well as enhanced antioxidant enzymes activities under stress. A higher level of ascorbic acid was detected in tir1 afb2 compared with wild-type plants. Thus, adaptation to salinity in Arabidopsis may be mediated in part by an auxin/redox interaction.

Mitochondria are the main target for oxidative damage in leaves of wheat (Triticum aestivum L.).

Photosynthesis, respiration, and other processes produce reactive oxygen species (ROS) that can cause oxidative modifications to proteins, lipids, and DNA. The production of ROS increases under stress conditions, causing oxidative damage and impairment of normal metabolism. In this work, oxidative damage to various subcellular compartments (i.e. chloroplasts, mitochondria, and peroxisomes) was studied in two cultivars of wheat differing in ascorbic acid content, and growing under good irrigation or drought. In well-watered plants, mitochondria contained 9-28-fold higher concentrations of oxidatively modified proteins than chloroplasts or peroxisomes. In general, oxidative damage to proteins was more intense in the cultivar with the lower content of ascorbic acid, particularly in the chloroplast stroma. Water stress caused a marked increase in oxidative damage to proteins, particularly in mitochondria and peroxisomes. These results indicate that mitochondria are the main target for oxidative damage to proteins under well-irrigated and drought conditions.