NADPH-dependent thioredoxin reductases (NTRs) are key regulatory enzymes determining the redox state of the thioredoxin system. The Arabidopsis thaliana genome has two genes coding for NTRs (NTRA and NTRB), both of which encode mitochondrial and cytosolic isoforms. Surprisingly, plants of the ntra ntrb knockout mutant are viable and fertile, although with a wrinkled seed phenotype, slower plant growth, and pollen with reduced fitness. Thus, in contrast with mammals, our data demonstrate that neither cytosolic nor mitochondrial NTRs are essential in plants. Nevertheless, in the double mutant, the cytosolic thioredoxin h3 is only partially oxidized, suggesting an alternative mechanism for thioredoxin reduction. Plant growth in ntra ntrb plants is hypersensitive to buthionine sulfoximine (BSO), a specific inhibitor of glutathione biosynthesis, and thioredoxin h3 is totally oxidized under this treatment. Interestingly, this BSO-mediated growth arrest is fully reversible, suggesting that BSO induces a growth arrest signal but not a toxic accumulation of activated oxygen species. Moreover, crossing ntra ntrb with rootmeristemless1, a mutant blocked in root growth due to strongly reduced glutathione synthesis, led to complete inhibition of both shoot and root growth, indicating that either the NTR or the glutathione pathway is required for postembryonic activity in the apical meristem.

Thioredoxins (Trx) and glutaredoxins (Grx) constitute families of thiol oxidoreductases. Our knowledge of Trx and Grx in plants has dramatically increased during the last decade. The release of the Arabidopsis genome sequence revealed an unexpectedly high number of Trx and Grx genes. The availability of several genomes of vascular and nonvascular plants allowed the establishment of a clear classification of the genes and the chronology of their appearance during plant evolution. Proteomic approaches have been developed that identified the putative Trx and Grx target proteins which are implicated in all aspects of plant growth, including basal metabolism, iron/sulfur cluster formation, development, adaptation to the environment, and stress responses. Analyses of the biochemical characteristics of specific Trx and Grx point to a strong specificity toward some target enzymes, particularly within plastidial Trx and Grx. In apparent contradiction with this specificity, genetic approaches show an absence of phenotype for most available Trx and Grx mutants, suggesting that redundancies also exist between Trx and Grx members. Despite this, the isolation of mutants inactivated in multiple genes and several genetic screens allowed the demonstration of the involvement of Trx and Grx in pathogen response, phytohormone pathways, and at several control points of plant development. Cytosolic Trxs are reduced by NADPH-thioredoxin reductase (NTR), while the reduction of Grx depends on reduced glutathione (GSH). Interestingly, recent development integrating biochemical analysis, proteomic data, and genetics have revealed an extensive crosstalk between the cytosolic NTR/Trx and GSH/Grx systems. This crosstalk, which occurs at multiple levels, reveals the high plasticity of the redox systems in plants.

Plant mitochondria have a fully operational tricarboxylic acid (TCA) cycle that plays a central role in generating ATP and providing carbon skeletons for a range of biosynthetic processes in both heterotrophic and photosynthetic tissues. The cycle enzymeencoding genes have been well characterized in terms of transcriptional and effector-mediated regulation and have also been subjected to reverse genetic analysis. However, despite this wealth of attention, a central question remains unanswered: "What regulates flux through this pathway in vivo?" Previous proteomic experiments with Arabidopsis discussed below have revealed that a number of mitochondrial enzymes, including members of the TCA cycle and affiliated pathways, harbor thioredoxin (TRX)-binding sites and are potentially redox-regulated. We have followed up on this possibility and found TRX to be a redox-sensitive mediator of TCA cycle flux. In this investigation, we first characterized, at the enzyme and metabolite levels, mutants of the mitochondrial TRX pathway in Arabidopsis: the NADP-TRX reductase a and b double mutant (ntra ntrb) and the mitochondrially located thioredoxin o1 (trxo1) mutant. These studies were followed by a comparative evaluation of the redistribution of isotopes when 13 Cglucose, 13 C-malate, or 13 C-pyruvate was provided as a substrate to leaves of mutant or WT plants. In a complementary approach, we evaluated the in vitro activities of a range of TCA cycle and associated enzymes under varying redox states. The combined dataset suggests that TRX may deactivate both mitochondrial succinate dehydrogenase and fumarase and activate the cytosolic ATP-citrate lyase in vivo, acting as a direct regulator of carbon flow through the TCA cycle and providing a mechanism for the coordination of cellular function.Arabidopsis | redox regulation | thioredoxin TCA cycle regulation | citric acid cycle regulation | ATP-citrate lyase A s in animals and aerobic microorganisms (1, 2), the tricarboxylic acid (TCA) cycle of plant mitochondria is composed of a set of eight enzymes that oxidize pyruvate and malate formed in the cytosol to CO 2 and NADH (3). The CO 2 is released and the NADH is oxidized by the electron transport chain for the generation of ATP. Recent years have witnessed major advances in our understanding of the cycle in plants, including its different modes of operation and properties of its constituent enzymes (4-6). We also now understand a great deal about the physiological role, kinetic features, and transcriptional and posttranslational regulation of enzymes participating in the cycle.In addition to these studies, experiments have focused on functional interactions taking place between mitochondria and the other organelle that generates energy in plant cells, namely, the chloroplast (7, 8). The results suggest that the two compartments are tightly linked by regulatory mechanisms acting at the levels of the gene and interorganellar metabolite transport (9-13). Further, a long-standing body of evidence indicates that the cycle is regul...

During the 70s and 80s two plant thioredoxin systems were identified. The chloroplastic system is composed of a ferredoxin-dependent thioredoxin, with two thioredoxin types (m and f) regulating the activity of enzymes implicated in photosynthetic carbon assimilation. In the cytosol of heterotrophic tissues, an NADP dependent thioredoxin reductase and a thioredoxin (h) were identified. The first plant glutaredoxin was only identified later, in 1994. Our view of plant thioredoxins and glutaredoxins was profoundly modified by the sequencing programs which revealed an unexpected number of genes encoding not only the previously identified disulfide reductases, but also numerous new types. At the same time it became clear that plant genomes encode chloroplastic, cytosolic and mitochondrial peroxiredoxins, suggesting a major role for redoxins in anti-oxidant defense. Efficient proteomics approaches were developed allowing the characterization of numerous thioredoxin target proteins. They are implicated in different aspects of plant life including development and adaptation to environmental changes and stresses. The most important challenge for the next years will probably be to identify in planta which redoxin reduces which target, a question which remains unsolved due to the low specificities of redoxins in vitro and the numerous redundancies which in most cases mask the phenotype of redoxin mutants.

Wild-type Escherichia coli K-12 ferments glucose to a mixture of ethanol and acetic, lactic, formic, and succinic acids. In anoxic chemostat culture at four dilution rates and two different oxidoreduction potentials (ORP), this strain generated a spectrum of products which depended on ORP. Whatever the dilution rate tested, in low reducing conditions (؊100 mV), the production of formate, acetate, ethanol, and lactate was in molar proportions of approximately 2.5:1:1:0.3, and in high reducing conditions (؊320 mV), the production was in molar proportions of 2:0.6:1:2. The modification of metabolic fluxes was due to an ORP effect on the synthesis or stability of some fermentation enzymes; thus, in high reducing conditions, lactate dehydrogenasespecific activity increased by a factor of 3 to 6. Those modifications were concomitant with a threefold decrease in acetyl-coenzyme A (CoA) needed for biomass synthesis and a 0.5-to 5-fold decrease in formate flux. Calculations of carbon and cofactor balances have shown that fermentation was balanced and that extracellular ORP did not modify the oxidoreduction state of cofactors. From this, it was concluded that extracellular ORP could regulate both some specific enzyme activities and the acetyl-CoA needed for biomass synthesis, which modifies metabolic fluxes and ATP yield, leading to variation in biomass synthesis.A wealth of information is available on the response of Escherichia coli cellular metabolism to pH, water activity, or temperature variations, but little is known about the action of extracellular oxidoreduction potentials (ORP) on metabolism, although numerous reactions and regulations are of the oxidoreduction type. Previous studies have shown that substrates with different oxidation states yield a specific product spectrum. Thus, with glucose (oxidation state ϭ 0), the spectrum of main end products (formate:acetate:ethanol:lactate) is equal to 2:1:1:2, with glucitol (oxidation state ϭ Ϫ1), it is equal to 2:1:6:0.5, and with glucuronate (oxidation state ϭ 2), it is equal to 1:5:1:1, with small amounts of succinate also being produced in each case (1). Those metabolic flux modifications are also observed when the nature of external electron acceptors varies (7). In the same way, the NADH/NAD ϩ ratio, which is responsible for regulation of enzymes (8) or genes (16), can be influenced by the oxidation level of the substrate (36) or by the availability and nature of electron acceptors (7). In addition, protein folding and disulfide bond formation are regulated and modified by different oxidoreductase enzymes and oxidoreduction couples (glutathione, thioredoxin) (26). Recently, Taylor and Zhulin in their review have shown that ORP influences or could influence numerous regulations of cell functions controlled via Per-Arnt-Sim (PAS)-containing receptors (signaling modules that monitor changes in light, ORP, oxygen, and the overall energy level of a cell), transducers, and regulators (34). Thus, E. coli senses the medium ORP and swims to a preferred ORP niche by redox...

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The major known function of glutaredoxins (Grxs) is to reduce disulphide bridges. Recently, some have also been shown to interact with iron-sulphur clusters. These can be classified in two subgroups: class II Grx are found in all living organisms and are implicated in assembly of ironsulphur clusters, while class I Grx are represented by only two members known to form a holodimer structure containing a cluster in vitro, but with an unknown function different from class II. Here, we report that in eukaryotic plants, GRXC1 (class I) orthologs are exclusively present in dicotyledonous plants, suggesting a specific function. Indeed, in Arabidopsis thaliana, reducing activity of recombinant GRXC1 is regulated by redox-dependent stability of the cluster. In planta, GRXC1 has been found predominantly in a holodimeric form, indicating the presence of the cluster in vivo. This suggests that GRXC1 acts as a redox sensor, reducing downstream pathways under oxidative conditions. GRXC2, the closest homolog of GRXC1, is unable to form a cluster in vitro. Knock-out mutants in grxc1 or grxc2 are aphenotypic, but the double mutant produces a lethal phenotype at an early stage after pollinization, suggesting that GRXC1 and GRXC2 share redundant and vital functions.

Thiol reduction proteins are key regulators of the redox state of the cell, managing development and stress response programs. In plants, thiol reduction proteins, namely thioredoxin (TRX), glutaredoxin (GRX), and their respective reducers glutathione reductase (GR) and thioredoxin reductase (TR), are organized in complex multigene families. In order to decipher the function of the different proteins, it is necessary to have a clear picture of their respective expression profiles. By collecting information from gene expression databases, we have performed a comprehensive in silico study of the expression of all members of different classes of thiol reduction genes (TRX, GRX) in Arabidopsis thaliana. Tissue expression profiles and response to many biotic and abiotic stress conditions have been studied systematically. Altogether, the significance of our data is discussed with respect to published biochemical and genetic studies.