Glutathione: a key modulator of plant defence and metabolism through multiple mechanisms.

Redox reactions are fundamental to energy conversion in living cells, and also determine and tune responses to the environment. Within this context, the tripeptide glutathione plays numerous roles. As an important antioxidant, glutathione confers redox stability on the cell and also acts as an interface between signalling pathways and metabolic reactions that fuel growth and development.

Thioredoxins m regulate plastid glucose-6-phosphate dehydrogenase activity in Arabidopsis roots under salt stress.

Plants contain several NADPH-producing enzymes including glucose-6-phosphate dehydrogenases (G6PDH) with different sub-cellular localizations. The activity of plastidial G6PDHs is redox-regulated by thioredoxins (TRX). Although specific TRXs are known to regulate chloroplastic isoforms of G6PDH, little information is available for plastidic isoforms found in heterotrophic organs or tissues.

Quantitative Measurements of Biochemical and Molecular Markers of Oxidative Stress Signaling and Responses.

Increases in cellular oxidation are a part of most plant responses to challenging conditions and are commonly described as oxidative stress. While this phenomenon is closely related to the accumulation of reactive oxygen species, these latter compounds can be difficult to measure. Complementary measurements to assess cellular redox state are, therefore, very useful in studies of plant responses to stress.

S-Nitrosylation of the histone deacetylase HDA19 stimulates its activity to enhance plant stress tolerance in Arabidopsis.

Arabidopsis histone deacetylase HDA19 is required for gene expression programs of a large spectrum of plant developmental and stress-responsive pathways. How this enzyme senses cellular environment to control its activity remains unclear. In this work, we show that HDA19 is post-translationally modified by S-nitrosylation at 4 Cysteine (Cys) residues.

Metabolite modification in oxidative stress responses: A case study of two defense hormones.

Studies of the Arabidopsis cat2 mutant lacking the major leaf isoform of catalase have allowed the potential impact of intracellular HO on plant function to be studied. Here, we report a robust analysis of modified gene expression associated with key families involved in metabolite modification in cat2. Through a combined transcriptomic and metabolomic analysis focused on the salicylic acid (SA) and jasmonic acid (JA) pathways, we report key features of the metabolic signatures linked to oxidative stress-induced signaling via these defence hormones and discuss the enzymes that are likely to be involved in determining these features.

Measuring Stress-Induced Changes in Defense Phytohormones and Related Compounds.

Measuring quantitative changes in plant hormones and derivatives is crucial to understand how reactive oxygen species trigger signaling cascades to regulate stress responses. In this chapter, we describe the liquid chromatography-mass spectrometry procedure that we use to extract and quantify salicylic acid (SA), jasmonic acid (JA), and related compounds in common extracts of Arabidopsis tissue. The method can provide quantitative data on SA, SA glucosides, and JA, as well as information on oxidized and conjugated forms of these compounds and related derivatives of benzoic acid.

Measurement of NAD(P)H and NADPH-Generating Enzymes.

Pyridine nucleotides (NAD(H) and NADP(H)) are key redox carriers in cells and may also have other functions related to stress. These two molecules are crucial in linking metabolism to electron transport chains in photosynthesis and respiration, but they are also critical for ensuring redox signaling and homeostasis during episodes of stress. This is especially the case for NADPH, which must be generated from its oxidized form, NADP, by key dehydrogenases.

Quantitative Measurement of Ascorbate and Glutathione by Spectrophotometry.

Ascorbate and glutathione are key chemical antioxidants present at relatively high concentrations in plant cells. They are also reducing cofactors for enzymes that process hydrogen peroxide in the ascorbate-glutathione pathway. Due to these two related biochemical functions, the compounds form an interface between reactive oxygen species and sensitive cellular components.

Redox Homeostasis and Signaling in a Higher-CO World.

Rising CO concentrations and their effects on plant productivity present challenging issues. Effects on the photosynthesis/photorespiration balance and changes in primary metabolism are known, caused by the competitive interaction of CO and O at the active site of ribulose-1,5-bisphosphate carboxylase/oxygenase. However, impacts on stress resistance are less clear.

Glutathione-dependent denitrosation of GSNOR1 promotes oxidative signalling downstream of H O.

Photorespiratory hydrogen peroxide (H O ) plays key roles in pathogenesis responses by triggering the salicylic acid (SA) pathway in Arabidopsis. However, factors linking intracellular H O to activation of the SA pathway remain elusive. In this work, the catalase-deficient Arabidopsis mutant, cat2, was exploited to elucidate the impact of S-nitrosoglutathione reductase 1 (GSNOR1) on H O -dependent signalling pathways.

PROTEIN PHOSPHATASE 2A-B' Controls Resistance and Developmental Leaf Senescence.

Plants optimize their growth and survival through highly integrated regulatory networks that coordinate defensive measures and developmental transitions in response to environmental cues. Protein phosphatase 2A (PP2A) is a key signaling component that controls stress reactions and growth at different stages of plant development, and the PP2A regulatory subunit PP2A-B'γ is required for negative regulation of pathogenesis responses and for maintenance of cell homeostasis in short-day conditions. Here, we report molecular mechanisms by which PP2A-B'γ regulates resistance and leaf senescence in Arabidopsis ().

Cytosolic Isocitrate Dehydrogenase from Is Regulated by Glutathionylation.

NADP-dependent (Nicotinamide Adénine Dinucléotide Phosphate-dependent) isocitrate dehydrogenases (NADP-ICDH) are metabolic enzymes involved in 2-oxoglutarate biosynthesis, but they also supply cells with NADPH. Different NADP-ICDH genes are found in among which a single gene encodes for a cytosolic ICDH (cICDH) isoform. Here, we show that cICDH is susceptible to oxidation and that several cysteine (Cys) residues are prone to S-nitrosylation upon nitrosoglutathione (GSNO) treatment.

Analysis of catalase mutants underscores the essential role of CATALASE2 for plant growth and day length-dependent oxidative signalling.

Three genes encode catalase in Arabidopsis. Although the role of CAT2 in photorespiration is well established, the importance of the different catalases in other processes is less clear. Analysis of cat1, cat2, cat3, cat1 cat2, and cat2 cat3 T-DNA mutants revealed that cat2 had the largest effect on activity in both roots and leaves.

Analyzing the Function of Catalase and the Ascorbate-Glutathione Pathway in HO Processing: Insights from an Experimentally Constrained Kinetic Model.

Significance: Plant stress involves redox signaling linked to reactive oxygen species such as hydrogen peroxide (HO), which can be generated at high rates in photosynthetic cells. The systems that process HO include catalase (CAT) and the ascorbate-glutathione pathway, but interactions between them remain unclear. Modeling can aid interpretation and pinpoint areas for investigation.

Measurement of Transcripts Associated with Photorespiration and Related Redox Signaling.

To study photorespiration and to characterize related components, gene expression analysis is a central approach. An overview of the experimental setup, protocols, and methods we use to investigate photorespiration-associated gene expression is presented. Within this chapter, we describe simple procedures to experimentally alter the photorespiratory flux and provide protocols for transcriptomic analysis with a focus on genes encoding photorespiratory proteins as well as those induced by photorespiratory hydrogen peroxide (HO).

Climate Change, CO, and Defense: The Metabolic, Redox, and Signaling Perspectives.

Ongoing human-induced changes in the composition of the atmosphere continue to stimulate interest in the effects of high CO on plants, but its potential impact on inducible plant defense pathways remains poorly defined. Recently, several studies have reported that growth at elevated CO is sufficient to induce defenses such as the salicylic acid pathway, thereby increasing plant resistance to pathogens. These reports contrast with evidence that defense pathways can be promoted by photorespiration, which is inhibited at high CO.

Glutathione oxidation in response to intracellular HO: Key but overlapping roles for dehydroascorbate reductases.

Glutathione is a pivotal molecule in oxidative stress, during which it is potentially oxidized by several pathways linked to HO detoxification. We have investigated the response and functional importance of 3 potential routes for glutathione oxidation pathways mediated by glutathione S-transferases (GST), glutaredoxin-dependent peroxiredoxins (PRXII), and dehydroascorbate reductases (DHAR) in Arabidopsis during oxidative stress. Loss-of-function gstU8, gstU24, gstF8, prxIIE and prxIIF mutants as well as double gstU8 gstU24, gstU8 gstF8, gstU24 gstF8, prxIIE prxIIF mutants were obtained.

ROS-related redox regulation and signaling in plants.

As sessile oxygenic organisms with a plastic developmental programme, plants are uniquely positioned to exploit reactive oxygen species (ROS) as powerful signals. Plants harbor numerous ROS-generating pathways, and these oxidants and related redox-active compounds have become tightly embedded into plant function and development during the course of evolution. One dominant view of ROS-removing systems sees them as beneficial antioxidants battling to keep damaging ROS below dangerous levels.

Functional analysis of the role of hydrogen sulfide in the regulation of dark-induced leaf senescence in Arabidopsis.

There is growing evidence that hydrogen sulfide (HS) is involved in many physiological processes in plants, but the role of HS in dark-induced leaf senescence remains unknown. In this work, we found that HS not only inhibited chlorophyll degradation but also caused the accumulation of photoreactive pheide a in detached leaves under extended darkness. Despite this, transcript levels of senescence-associated genes (SAGs) were less affected in HS-treated detached leaves compared with those in HS-untreated detached leaves.

Cytosolic and Chloroplastic DHARs Cooperate in Oxidative Stress-Driven Activation of the Salicylic Acid Pathway.

The complexity of plant antioxidative systems gives rise to many unresolved questions. One relates to the functional importance of dehydroascorbate reductases (DHARs) in interactions between ascorbate and glutathione. To investigate this issue, we produced a complete set of loss-of-function mutants for the three annotated Arabidopsis () DHARs.

Viewing oxidative stress through the lens of oxidative signalling rather than damage.

Concepts of the roles of reactive oxygen species (ROS) in plants and animals have shifted in recent years from focusing on oxidative damage effects to the current view of ROS as universal signalling metabolites. Rather than having two opposing activities, i.e.

High CO2 Primes Plant Biotic Stress Defences through Redox-Linked Pathways.

Industrial activities have caused tropospheric CO concentrations to increase over the last two centuries, a trend that is predicted to continue for at least the next several decades. Here, we report that growth of plants in a CO-enriched environment activates responses that are central to defense against pathogenic attack. Salicylic acid accumulation was triggered by high-growth CO in Arabidopsis (Arabidopsis thaliana) and other plants such as bean (Phaseolus vulgaris).