Caractérisation et rôle des NADP-thiorédoxines réductases et des thiorédoxines au cours de la germination des graines de Medicago truncatula

Afin de cerner leur rôle dans la germination des légumineuses, les NADPH-thioredoxine réductases (NTR) et thiorédoxines (Trx) ont été caractérisées chez Medicago truncatula Tandis que MtNTRA a les mêmes propriétés que ses orthologues, MtNTRC a des propriétés uniques. Elle réduit efficacement BAS1 en présence de NADH ou NADPH. Comme elle est présente avec BAS1 dans les tissus verts et les graines, le couple pourrait éliminer les peroxydes produits dans ces organes. Les sept Trx h caractérisées ont des propriétés similaires aux Trx h déjà décrites. Seules Mth1, h2 et h6 auraient un rôle dans la germination. Un nouveau type de Trx associé à la symbiose a également été découvert. Enfin, 111 cibles potentielles des Trx ont été identifiées, la moitié étant nouvelle. Ainsi, les Trx participent à la mobilisation des protéines de réserves et sont impliquées dans la tolérance aux stress. Une régulation par les Trx de la synthèse des phénylpropanoïdes et de la gestion des stocks de certains métabolites a aussi été découverte.To determine the role of NADPH-thioredoxin reductases (NTR) and thioredoxins (Trx) in legume germination, they were characterised and their expression analysed in seed of Medicago truncatula. While MtNTRA has the same properties as its orthologs, MtNTRC has unique properties. Indeed, it reduces efficiently BAS1 using either NADH or NADPH. As BAS1, it is present in green tissues and seeds, suggesting that NTRC/BAS1 may be involved in the detoxication of peroxides produced in these organs. Seven out of 11 Trx h isoforms were characterised. Only Mth1, Mth2 and Mth6 may have a role during germination. Interestingly, a new type of Trx associated with symbiosis was also discovered in M. truncatula. Finally, 111 potential targets were identified in seeds, half being new. As in cereals, Trx are involved in storage protein mobilisation and stress tolerance. However, a new role of Trx in the regulation of the synthesis of DNA and phenylpropanoids as well as in the storage of certain metabolites was found in seed of M. TruncatulaANGERS-BU Lettres et Sciences (490072106) / SudocSudocFranceF

Identification and differential expression of two thioredoxin h isoforms in germinating seeds from pea

http://dx.doi.org/10.1104/pp.102.019562International audienc

Involvement of Glutaredoxin and Thioredoxin Systems in the Nitrogen-Fixing Symbiosis between Legumes and Rhizobia

Leguminous plants can form a symbiotic relationship with Rhizobium bacteria, during which plants provide bacteria with carbohydrates and an environment appropriate to their metabolism, in return for fixed atmospheric nitrogen. The symbiotic interaction leads to the formation of a new organ, the root nodule, where a coordinated differentiation of plant cells and bacteria occurs. The establishment and functioning of nitrogen-fixing symbiosis involves a redox control important for both the plant-bacteria crosstalk and the regulation of nodule metabolism. In this review, we discuss the involvement of thioredoxin and glutaredoxin systems in the two symbiotic partners during symbiosis. The crucial role of glutathione in redox balance and S-metabolism is presented. We also highlight the specific role of some thioredoxin and glutaredoxin systems in bacterial differentiation. Transcriptomics data concerning genes encoding components and targets of thioredoxin and glutaredoxin systems in connection with the developmental step of the nodule are also considered in the model system Medicago truncatula⁻Sinorhizobium meliloti

Identification and Differential Expression of Two Thioredoxin h Isoforms in Germinating Seeds from Pea

The NADPH/NADP-thioredoxin (Trx) reductase (NTR)/Trx system (NTS) is a redox system that plays a posttranslational regulatory role by reducing protein targets involved in crucial cellular processes in microorganisms and animals. In plants, the system includes several h type Trx isoforms and has been shown to intervene in reserve mobilization during early seedling growth of cereals. To determine whether NTS was operational during germination of legume seeds and which Trx h isoforms could be implicated, Trx h isoforms expression was monitored in germinating pea (Pisum sativum cv Baccara) seeds, together with the amount of NTR and NADPH. Two new isoforms were identified: Trx h3, similar to the two isoforms already described in pea but not expressed in seeds; and the more divergent isoform, Trx h4. Active recombinant proteins were produced in Escherichia coli and used to raise specific antibodies. The expression of new isoforms was analyzed at both mRNA and protein levels. The lack of correlation between mRNA and protein abundances suggests the occurrence of posttranscriptional regulation. Trx h3 protein amount remained constant in both axes and cotyledons of dry and imbibed seeds but then decreased 2 d after radicle protrusion. In contrast, Trx h4 was only expressed in axes of dry and imbibed seeds but not in germinated seeds or in seedlings, therefore appearing as closely linked to germination. The presence of NTR and NADPH in seeds suggests that NTS could be functional during germination. The possible role of Trx h3 and h4 in this context is discussed

Evidence for participation of the methionine sulfoxide reductase repair system in plant seed longevity

Seeds are in a natural oxidative context leading to protein oxidation. Although inevitable for proper progression from maturation to germination; protein oxidation at high levels is detrimental and associated with seed aging. Oxidation of methionine to methionine sulfoxide is a common form of damage observed during aging in all organisms. This damage is reversible through the action of methionine sulfoxide reductases (MSRs), which play key roles in lifespan control in yeast and animal cells. To investigate the relationship between MSR capacity and longevity in plant seeds, we first used two Medicago truncatula genotypes with contrasting seed quality. After characterizing the MSR family in this species, we analyzed gene expression and enzymatic activity in immature and mature seeds exhibiting distinct quality levels. We found a very strong correlation between the initial MSR capacities in different lots of mature seeds of the two genotypes and the time to a drop in viability to 50% after controlled deterioration. We then analyzed seed longevity in Arabidopsis thaliana lines, in which MSR gene expression has been genetically altered, and observed a positive correlation between MSR capacity and longevity in these seeds as well. Based on our data, we propose that the MSR repair system plays a decisive role in the establishment and preservation of longevity in plant seeds

Post-translational modifications of Medicago truncatula glutathione peroxidase 1 induced by nitric oxide

Plant glutathione peroxidases (Gpx) catalyse the reduction of various peroxides, such as hydrogen peroxide (H2O2), phospholipid hydroperoxides and peroxynitrite, but at the expense of thioredoxins rather than glutathione. A main function of plant Gpxs is the protection of biological membranes by scavenging phospholipid hydroperoxides, but some Gpxs have also been associated with H2O2 sensing and redox signal transduction. Nitric oxide (NO) is not only known to induce the expression of Gpx family members, but also to inhibit Gpx activity, presumably through the S-nitrosylation of conserved cysteine residues. In the present study, the effects of NO-donors on both the activity and S-nitrosylation state of purified Medicago truncatula Gpxl1 were analyzed using biochemical assay measurements and a biotin-switch/mass spectrometry approach. MtGpx1 activity was only moderately inhibited by the NO donors diethylamine-NONOate and S-nitrosoglutathione, and the inhibition may be reversed by DTT. The three conserved Cys of MtGpx1 were found to be modified through S-nitrosylation and S-glutathionylation, although to different extents, by diethylamine-NONOate and S-nitrosoglutathione, or by a combination of diethylamine-NONOate and reduced glutathione. The regulation of MtGpx1 and its possible involvement in the signaling process is discussed in the light of these results

A New Role for Plastid Thioredoxins in Seed Physiology in Relation to Hormone Regulation

International audienceIn Arabidopsis seeds, ROS have been shown to be enabling actors of cellular signaling pathways promoting germination, but their accumulation under stress conditions or during aging leads to a decrease in the ability to germinate. Previous biochemical work revealed that a specific class of plastid thioredoxins (Trxs), the y-type Trxs, can fulfill antioxidant functions. Among the ten plastidial Trx isoforms identified in Arabidopsis, Trx y1 mRNA is the most abundant in dry seeds. We hypothesized that Trx y1 and Trx y2 would play an important role in seed physiology as antioxidants. Using reverse genetics, we found important changes in the corresponding Arabidopsis mutant seeds. They display remarkable traits such as increased longevity and higher and faster germination in conditions of reduced water availability or oxidative stress. These phenotypes suggest that Trxs y do not play an antioxidant role in seeds, as further evidenced by no changes in global ROS contents and protein redox status found in the corresponding mutant seeds. Instead, we provide evidence that marker genes of ABA and GAs pathways are perturbed in mutant seeds, together with their sensitivity to specific hormone inhibitors. Altogether, our results suggest that Trxs y function in Arabidopsis seeds is not linked to their previously identified antioxidant roles and reveal a new role for plastid Trxs linked to hormone regulation

Functional characterization of the Medicago truncatula dual affinity nitrate transporter MtNRT1.3 and regulation of gene expression

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