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International audience; Senescence determines plant organ lifespan depending on aging and environmental cues. During the endosymbiotic interaction with rhizobia, legume plants develop a specific organ, the root nodule, which houses nitrogen (N)-fixing bacteria. Unlike earlier processes of the legume-rhizobium interaction (nodule formation, N fixation), mechanisms controlling nodule senescence remain poorly understood. To identify nodule senescence-associated genes, we performed a dual plant-bacteria RNA sequencing approach on Medicago truncatula-Sinorhizobium meliloti nodules having initiated senescence either naturally (aging) or following an environmental trigger (nitrate treatment or salt stress). The resulting data allowed the identification of hundreds of plant and bacterial genes differentially regulated during nodule senescence, thus providing an unprecedented comprehensive resource of new candidate genes associated with this process. Remarkably, several plant and bacterial genes related to the cell cycle and stress responses were regulated in senescent nodules, including the rhizobial RpoE2-dependent general stress response. Analysis of selected core nodule senescence plant genes allowed showing that MtNAC969 and MtS40, both homologous to leaf senescence-associated genes, negatively regulate the transition between N fixation and senescence. In contrast, overexpression of a gene involved in the biosynthesis of cytokinins, well-known negative regulators of leaf senescence, may promote the transition from N fixation to senescence in nodules.

Reactive oxygen species such as hydrogen peroxide (H

Legumes can meet their nitrogen requirements through root nodule symbiosis, which could also trigger plant systemic resistance against pests. The pea aphid Acyrthosiphon pisum , a legume pest, can harbour different facultative symbionts (FS) influencing various traits of their hosts. It is therefore worth determining if and how the symbionts of the plant and the aphid modulate their interaction. We used different pea aphid lines without FS or with a single one ( Hamiltonella defensa , Regiella insecticola, Serratia symbiotica ) to infest Medicago truncatula plants inoculated with Sinorhizobium meliloti (symbiotic nitrogen fixation, SNF) or supplemented with nitrate (non-inoculated, NI). The growth of SNF and NI plants was reduced by aphid infestation, while aphid weight (but not survival) was lowered on SNF compared to NI plants. Aphids strongly affected the plant nitrogen fixation depending on their symbiotic status, suggesting indirect relationships between aphid- and plant-associated microbes. Finally, all aphid lines triggered expression of Pathogenesis-Related Protein 1 ( PR1 ) and Proteinase Inhibitor (PI) , respective markers for salicylic and jasmonic pathways, in SNF plants, compared to only PR1 in NI plants. We demonstrate that the plant symbiotic status influences plant–aphid interactions while that of the aphid can modulate the amplitude of the plant's defence response.

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In the legume-rhizobia symbiosis, atmospheric dinitrogen (N2) fixation by rhizobia takes place in a specific root organ, called the nodule. During biological N2 fixation, the plant provides photosynthetic carbohydrates to bacteria and, in exchange, bacteria feed the plant with reduced nitrogen (N) under ammonia (NH3) form. Like most other organs, the root nodule has a limited lifespan, and eventually enters a senescence process characterized by a decline of N2 fixation and the coordinated death of both bacteria and plant cells. This senescence process characterized by a metabolic switch from a carbon sink to a nutrient source either results from nodule aging (developmental nodule senescence) or can be triggered prematurely by adverse environmental conditions. Accumulating evidence suggests that redox and hormone signaling contribute to the triggering of senescence. Indeed, nodule senescence is a developmentally programmed process in which antioxidants, reactive oxygen and N species (ROS/RNS), hormones and proteinases play key roles. This chapter reviews recent knowledge on nodule senescence and proposes models by which ROS, RNS, and likely hormones intervene in the orchestration of this process.

The interaction between legumes and bacteria of rhizobia type results in a beneficial symbiotic relationship characterized by the formation of new root organs, called nodules. Within these nodules the bacteria, released in plant cells, differentiate into bacteroids and fix atmospheric nitrogen through the nitrogenase activity. This mutualistic interaction has evolved sophisticated signaling networks to allow rhizobia entry, colonization, bacteroid differentiation and persistence in nodules. Nodule cysteine rich (NCR) peptides, reactive oxygen species (ROS), reactive nitrogen species (RNS), and toxin-antitoxin (TA) modules produced by the host plants or bacterial microsymbionts have a major role in the control of the symbiotic interaction. These molecules described as weapons in pathogenic interactions have evolved to participate to the intracellular bacteroid accommodation by escaping control of plant innate immunity and adapt the functioning of the nitrogen-fixation to environmental signalling cues.

Interactions between legumes and rhizobia lead to the establishment of a symbiotic relationship characterized by the formation of a new organ, the nodule, which facilitates the fixation of atmospheric nitrogen (N2) by nitrogenase through the creation of a hypoxic environment. Significant amounts of nitric oxide (NO) accumulate at different stages of nodule development, suggesting that NO performs specific signaling and/or metabolic functions during symbiosis. NO, which regulates nodule gene expression, accumulates to high levels in hypoxic nodules. NO accumulation is considered to assist energy metabolism within the hypoxic environment of the nodule via a phytoglobin–NO-mediated respiration process. NO is a potent inhibitor of the activity of nitrogenase and other plant and bacterial enzymes, acting as a developmental signal in the induction of nodule senescence. Hence, key questions concern the relative importance of the signaling and metabolic functions of NO versus its toxic action and how NO levels are regulated to be compatible with nitrogen fixation functions. This review analyses these paradoxical roles of NO at various stages of symbiosis, and highlights the role of plant phytoglobins and bacterial hemoproteins in the control of NO accumulation.

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.

International audience; Background: Nitrogen-fixing symbiosis between Rhizobium bacteria and legumes leads to the formation of a new organ, the root nodule. The development of the nodule requires the differentiation of plant root cells to welcome the endosymbiotic bacterial partner. This development includes the formation of an efficient vascular tissue which allows metabolic exchanges between the root and the nodule, the formation of a barrier to oxygen diffusion necessary for the bacterial nitrogenase activity and the enlargement of cells in the infection zone to support the large bacterial population. Inside the plant cell, the bacteria differentiate into bacteroids which are able to reduce atmospheric nitrogen to ammonia needed for plant growth in exchange for carbon sources. Nodule functioning requires a tight regulation of the development of plant cells and bacteria. Scope of the review: Nodule functioning requires a tight regulation of the development of plant cells and bacteria. The importance of redox control in nodule development and N-fixation is discussed in this review. The involvement of reactive oxygen and nitrogen species and the importance of the antioxidant defense are analyzed. Major conclusions: Plant differentiation and bacterial differentiation are controlled by reactive oxygen and nitrogen species, enzymes involved in the antioxidant defense and antioxidant compounds. General significance: The establishment and functioning of nitrogen-fixing symbiosis involve a redox control important for both the plant-bacteria crosstalk and the consideration of environmental parameters. This article is part of a Special Issue entitled Redox regulation of differentiation and de-differentiation.

Glutathione (GSH) is a major antioxidant molecule in plants. It is involved in regulating plant development and responses to abiotic and biotic environment changes. In leguminous plants, a GSH homolog, homoglutathione is also found. Most legumes can develop a symbiotic interaction with soil bacteria of the rhizobium family under nitrogen deficiency. This symbiosis allows the reduction of atmospheric nitrogen by the bacteria in plant organs called root nodules. In this chapter, we summarize studies that describe the synthesis and the roles of GSH and hGSH in the nitrogen-fixing symbiosis.

The symbiotic interaction between legumes and bacteria of Rhizobium type leads to the formation of new organs, called nodules, which provides a niche for bacterial nitrogen (N2) fixation. In the nodules, bacteria differentiate into bacteroids able to fix atmospheric N2 through nitrogenase activity. As nitrogenase is strongly inhibited by oxygen, N2-fixation is made possible thanks to microaerophilic conditions prevailing in the nodules, characterized by a balance between low oxygen contents and adjusted nitric oxide (NO) contents. NO was shown to be produced by both the plant and bacterial partners during symbiosis, from early interaction steps between the plant and the bacteria to N2-fixing and senescence steps in mature nodules. NO is required for an optimal establishment of the symbiotic interaction. Transcriptomic analysis at early stage of the symbiosis showed that NO regulates about 2030 genes including genes involved in the induction of cell dedifferentiation and organogenesis, and in the repression of plant defense reactions, favouring the establishment of the plant–microbe interaction. In mature nodules, NO was found to act as both an inhibitor of N2-fixation, a regulator of nitrogen metabolism, a beneficial metabolic intermediate for the maintenance of the energy metabolism under hypoxic conditions and a signal triggering nodule senescence. The present review provides an overview of NO sources and degradation pathways and of its multifaceted functions throughout the whole symbiotic process. It additionally analyses NO crosstalks with reactive oxygen species and hormones and presents particularly promising issues to decipher the roles of NO in N2-fixing symbioses.

Legume-Rhizobium nitrogen fixation is dramatically affected under drought and other environmental constraints. However, it has yet to be established as to whether such regulation of nitrogen fixation is only exerted at the whole-plant level (e.g. by a systemic nitrogen feedback mechanism) or can also occur at a local nodule level. To address this question, nodulated pea (Pisum sativum) plants were grown in a split-root system, which allowed for half of the root system to be irrigated at field capacity, while the other half was water deprived, thus provoking changes in the nodule water potential. Nitrogen fixation only declined in the water-deprived, half-root system and this result was correlated with modifications in the activities of key nodule's enzymes such as sucrose synthase and isocitrate dehydrogenase and in nodular malate content. Furthermore, the decline in nodule water potential resulted in a cell redox imbalance. The results also indicate that systemic nitrogen feedback signaling was not operating in these water-stressed plants, since nitrogen fixation activity was maintained at control values in the watered half of the split-root plants. Thus, the use of a partially draughted split-root system provides evidence that nitrogen fixation activity under drought stress is mainly controlled at the local level rather than by a systemic nitrogen signal. © 2007 American Society of Plant Biologists.

Three genes encoding heme hydroperoxidases (katA, katB, and katC) have been identified in the soil bacterium Sinorhizobium meliloti. The recombinant proteins were overexpressed in Escherichia coli and purified in order toachieve a spectral and kinetic characterization. The three proteins contain heme b with high-spin Fe(III). KatB is an acidic bifunctional homodimeric catalase-peroxidase exhibiting both catalase (k c a t = 2400 s - 1 ) and peroxidase activity and having a high affinity for hydrogen peroxide (apparent K M = 1.6 mM). KatA and KatC are acidic monofunctional homotetrameric catalases. Although different in size (KatA is a small subunit catalase while KatC is a large subunit catalase) both enzymes exhibit the same heme type and a similar affinity for H 2 O 2 (apparent K M values of 160 and 150 mM). However, the turnover rate of KatA (k c a t = 279000 s - 1 ) exceeds that of KatC (k c a t = 3100 s - 1 ) significantly. The kinetic parameters are in good agreement with the physiological role of these heme proteins. KatB is the housekeeping hydroperoxidase exhibiting the highest affinity for hydrogen peroxide, while KatA has the lowest H 2 O 2 affinity but the highest k c a t /K M value (1.75 x 10 6 M - 1 s - 1 ), in agreement with the hydrogen peroxide inducibility of the encoding gene. Moreover, the lower catalytic efficiency of KatC (2.1 × 10 4 M - 1 s - 1 ) appears to be enough for growing in the stationary phase and/or under heat or salt stress (conditions that are known to favor katC expression).

Haem oxygenases (HO) are ubiquitous enzymes catalysing the oxidative degradation of haem into biliverdin, iron and carbon monoxide. Whereas animal HOs participate in multiple cellular functions including haemoglobin catabolism, antioxidant defence and iron homeostasis, to date, plant HOs have so far only been involved in phytochrome metabolism. The expression of the HO1 gene was studied in Medicago sativa, especially during the interaction with its symbiotic partner, Sinorhizobium meliloti. Transcript accumulation was higher in mature root nodules than in roots and leaves and was correlated to HO1 protein immunodetection. The analysis of HO1 expression following alfalfa root inoculation with S. meliloti indicates that transcripts do not accumulate during the early steps of symbiosis, but rather in the mature nodules. These results correlate with the expression of the leghaemoglobin gene, which encodes the major haem-containing protein present in the nodule. Contrary to its animal counterpart, alfalfa HO1 was not induced by pro- oxidant compounds including H(2)O(2), paraquat and sodium nitroprusside, suggesting that it is not involved in the antioxidant defence. The results suggest that HO1 could play a role in the alfalfa mature nodule and its involvement in leghaemoglobin metabolism is hypothesized.

Reactive oxygen species are generated in the first steps of the Rhizobium–legume symbiosis. Superoxide radicals and hydrogen peroxide have been detected in infection threads and there is also evidence of the presence of nitric oxide in young alfalfa nodules. Moreover, rhizobial mutants, with a reduced antioxidant defense, exhibit an impaired capacity to nodulate. The oxidative burst generated in response to symbiotic infection can be consistent with rhizobia being initially perceived as invaders by the plant; in this framework, it may be correlated with the existence of abortive infections. However, the burst appears to be also involved in the expression of early nodulins associated with successful infections. Thus, in parallel to its involvement in defense processes, a positive role for the oxidative burst (including nitric oxide) in the establishment of the symbiotic interaction can also be proposed. The burst could trigger the expression of plant and/or bacterial genes which are essential for the nodulation process. In this framework, glutathione and homoglutathione could be key intermediates for gene expression, via the modification of the redox balance. Thus, the oxidative burst may have a dual role in the establishment of the symbiosis. © 2002 Editions scientifiques et medicales Elsevier SAS. All rights reserved.

10 Págs., 2 Figs. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CCBY). The .pdf document is protected by copyright and was first published by Frontiers., In nitrogen poor soils legumes establish a symbiotic interaction with rhizobia that results in the formation of root nodules. These are unique plant organs where bacteria differentiate into bacteroids, which express the nitrogenase enzyme complex that reduces atmospheric N2 to ammonia. Nodule metabolism requires a tight control of the concentrations of reactive oxygen and nitrogen species (RONS) so that they can perform useful signaling roles while avoiding nitro-oxidative damage. In nodules a thiol-dependent regulatory network that senses, transmits and responds to redox changes is starting to be elucidated. A combination of enzymatic, immunological, pharmacological and molecular analyses has allowed us to conclude that glutathione and its legume-specific homolog, homoglutathione, are abundant in meristematic and infected cells, that their spatio-temporally distribution is correlated with the corresponding (homo)glutathione synthetase activities, and that they are crucial for nodule development and function. Glutathione is at high concentrations in the bacteroids and at moderate amounts in the mitochondria, cytosol and nuclei. Less information is available on other components of the network. The expression of multiple isoforms of glutathione peroxidases, peroxiredoxins, thioredoxins, glutaredoxins and NADPH-thioredoxin reductases has been detected in nodule cells using antibodies and proteomics. Peroxiredoxins and thioredoxins are essential to regulate and in some cases to detoxify RONS in nodules. Further research is necessary to clarify the regulation of the expression and activity of thiol redox-active proteins in response to abiotic, biotic and developmental cues, their interactions with downstream targets by disulfide-exchange reactions, and their participation in signaling cascades. The availability of mutants and transgenic lines will be crucial to facilitate systematic investigations into the function of the various proteins in the legume-rhizobial symbiosis., Research from our laboratories has been funded by grants from MAE and CNRS (Envimedprogram) to Pierre Frendo and the Spanish Ministry of Economy and Competitivity (AGL2011-24524, cofunded by FEDER) to Manuel Becana.

PRAF proteins are present in all plants, but their functions remain unclear. We investigated the role of one member of the PRAF family, MtZR1, on the development of roots and nitrogen-fixing nodules in Medicago truncatula. We found that MtZR1 was expressed in all M. truncatula organs. Spatiotemporal analysis showed that MtZR1 expression in M. truncatula roots was mostly limited to the root meristem and the vascular bundles of mature nodules. MtZR1 expression in root nodules was down-regulated in response to various abiotic stresses known to affect nitrogen fixation efficiency. The down-regulation of MtZR1 expression by RNA interference in transgenic roots decreased root growth and impaired nodule development and function. MtZR1 overexpression resulted in longer roots and significant changes to nodule development. Our data thus indicate that MtZR1 is involved in the development of roots and nodules. To our knowledge, this work provides the first in vivo experimental evidence of a biological role for a typical PRAF protein in plants.

International audience; Reactive oxygen species (ROS), particularly hydrogen peroxide (H(2)O(2)), play an important role in signalling in various cellular processes. The involvement of H(2)O(2) in the Medicago truncatula-Sinorhizobium meliloti symbiotic interaction raises questions about its effect on gene expression. A transcriptome analysis was performed on inoculated roots of M. truncatula in which ROS production was inhibited with diphenylene iodonium (DPI). In total, 301 genes potentially regulated by ROS content were identified 2 d after inoculation. These genes included MtSpk1, which encodes a putative protein kinase and is induced by exogenous H(2)O(2) treatment. MtSpk1 gene expression was also induced by nodulation factor treatment. MtSpk1 transcription was observed in infected root hair cells, nodule primordia and the infection zone of mature nodules. Analysis with a fluorescent protein probe specific for H(2)O(2) showed that MtSpk1 expression and H(2)O(2) were similarly distributed in the nodule infection zone. Finally, the establishment of symbiosis was impaired by MtSpk1 downregulation with an artificial micro-RNA. Several genes regulated by H(2)O(2) during the establishment of rhizobial symbiosis were identified. The involvement of MtSpk1 in the establishment of the symbiosis is proposed.

Under nitrogen-limiting conditions, legumes interact with symbiotic rhizobia to produce nitrogen-fixing root nodules. We have previously shown that glutathione and homoglutathione [(h)GSH] deficiencies impaired Medicago truncatula symbiosis efficiency, showing the importance of the low M r thiols during the nodulation process in the model legume M. truncatula. In this study, the plant transcriptomic response to Sinorhizobium meliloti infection under (h)GSH depletion was investigated using cDNA-amplified fragment length polymorphism analysis. Among 6,149 expression tags monitored, 181 genes displayed significant differential expression between inoculated control and inoculated (h)GSH depleted roots. Quantitative reverse transcription polymerase chain reaction analysis confirmed the changes in mRNA levels. This transcriptomic analysis shows a down-regulation of genes involved in meristem formation and a modulation of the expression of stress-related genes in (h)GSH-depleted plants. Promoter-β-glucuronidase histochemical analysis showed that the putative MtPIP2 aquaporin might be up-regulated during nodule meristem formation and that this up-regulation is inhibited under (h)GSH depletion. (h)GSH depletion enhances the expression of salicylic acid (SA)-regulated genes after S. meliloti infection and the expression of SA-regulated genes after exogenous SA treatment. Modification of water transport and SA signaling pathway observed under (h)GSH deficiency contribute to explain how (h)GSH depletion alters the proper development of the symbiotic interaction.

Reactive Oxygen Species (ROS) are continuously produced as a result of aerobic metabolism or in response to biotic and abiotic stresses. ROS are not only toxic by-products of aerobic metabolism, but are also signaling molecules involved in plant growth and environmental adaptation. Antioxidants can protect the cell from oxidative damage by scavenging the ROS. Thus, they play an important role in optimizing cell function by regulating cellular redox state and modifying gene expression. This article aims to review recent studies highlighting the role of redox signals in establishing and maintaining symbiosis between rhizobia and legumes.

The symbiosis between legumes and Rhizobia leads to the formation of a new organ, the nodule, where nitrogen fixation takes place. The plant provides bacteria with energy and a micro-aerobic environment compatible with nitrogenase activity. In exchange, bacteria provide a nitrogen supply to the plant. Therefore, nodules represent a unique model for the study of developmental processes, plant–microbe, and carbon/nitrogen/oxygen metabolism interactions. Reactive oxygen species (ROS) have been described to be produced at every step of the symbiotic association: during the symbiosis establishment, during the nitrogen fixation and during the senescence of the nodule. ROS can be harmful provoking oxidative damage or can have a positive role in signalling processes. To deal with ROS production, nodules are fitted with a large panel of enzymatic and non-enzymatic antioxidant mechanisms. This review summarizes the present knowledge about ROS production and scavenging systems in legume nodules, and about their functional roles during the different steps of the symbiotic interaction.

Plants appear to generate reactive oxygen species (ROS) as signaling molecules to control various fundamental processes. With this background, this review aims to highlight the involvement of ROS, and their possible interactions with nitric oxide (NO) and glutathione (GSH) in the symbiosis between rhizobia and leguminous plants. This compatible interaction, which is very important for sustainable agriculture, leads to the formation of a novel organ capable of fixing atmospheric nitrogen. ROS are involved in the early steps of the symbiotic interaction: their presence is essential for the development of optimal symbiosis and points to a signaling role for ROS during the symbiotic process. ROS may also regulate nodule function by interacting with NO.

Several reactive oxygen and nitrogen species (ROS/RNS) are continuously produced in plants as by-products of aerobic metabolism or in response to stresses. Depending on the nature of the ROS and RNS, some of them are highly toxic and rapidly detoxified by various cellular enzymatic and non-enzymatic mechanisms. Whereas plants have many mechanisms with which to combat increased ROS/RNS levels produced during stress conditions, under other circumstances plants appear to generate ROS/RNS as signalling molecules to control various processes encompassing the whole lifespan of the plant such as normal growth and development stages. This review aims to summarize recent studies highlighting the involvement of ROS/RNS, as well as the low molecular weight thiols, glutathione and homoglutathione, during the symbiosis between rhizobia and leguminous plants. This compatible interaction initiated by a molecular dialogue between the plant and bacterial partners, leads to the formation of a novel root organ capable of fixing atmospheric nitrogen under nitrogen-limiting conditions. On the one hand, ROS/RNS detection during the symbiotic process highlights the similarity of the early response to infection by pathogenic and symbiotic bacteria, addressing the question as to which mechanism rhizobia use to counteract the plant defence response. Moreover, there is increasing evidence that ROS are needed to establish the symbiosis fully. On the other hand, GSH synthesis appears to be essential for proper development of the root nodules during the symbiotic interaction. Elucidating the mechanisms that control ROS/RNS signalling during symbiosis could therefore contribute in defining a powerful strategy to enhance the efficiency of the symbiotic interaction.

Legumes form a symbiotic interaction with bacteria of the Rhizobiaceae family toproduce nitrogen-fixing root nodules under nitrogen-limiting conditions. This process involves the recognition of the bacterial Nod factors by the plant which mediates the entry of the bacteria into the root and nodule organogenesis. We have examined the importance of the low molecular weight thiols, glutathione (GSH) and homoglutathione (hGSH), during the nodulation process in the model legume Medicago truncatula. Using both buthionine sulfoximine, a specific inhibitor of GSH and hGSH synthesis, and transgenic roots expressing GSH synthetase and hGSH synthetase in an antisense orientation, we showed that deficiency in GSH and hGSH synthesis inhibited the formation of the root nodules. This inhibition was not correlated to a modification in the number of infection events or to a change in the expression of the Rhizobium sp.-induced peroxidase rip1, indicating that the low level of GSH or hGSH did not alter the first steps of the infection process. In contrast, a strong diminution in the number of nascent nodules and in the expression of the early nodulin genes, Mtenod12 and Mtenod40, were observed in GSHand hGSH-depleted plants. In conclusion, GSH and hGSH appear to be essential for proper development of the root nodules during the symbiotic interaction.

Rhizobia form a symbiotic relationship with plants of the legume family to produce nitrogen-fixing root nodules under nitrogen-limiting conditions. We have examined the importance of glutathione (GSH) during free-living growth and symbiosis of Sinorhizobium meliloti. An S. meliloti mutant strain (SmgshA) which is unable to synthesize GSH due to a gene disruption in gshA, encoding the enzyme for the first step in the biosynthesis of GSH, was unable to grow under nonstress conditions, precluding any nodulation. In contrast, an S. meliloti strain (SmgshB) with gshB, encoding the enzyme involved in the second step in GSH synthesis, deleted was able to grow, indicating that γ- glutamylcysteine, the dipeptide intermediate, can partially substitute for GSH. However, the SmgshB strain showed a delayed-nodulation phenotype coupled to a 75% reduction in the nitrogen fixation capacity. This phenotype was linked to abnormal nodule development. Both the SmgshA and SmgshB mutant strains exhibited higher catalase activity than the wild-type S. meliloti strain, suggesting that both mutant strains are under oxidative stress. Taken together, these results show that GSH plays a critical role in the growth of S. meliloti and during its interaction with the plant partner., Facultad de Ciencias Exactas, Instituto de Biotecnologia y Biologia Molecular

Two catalases, KatA and KatB, have been detected in Sinorhizobium meliloti growing on rich medium. Here we characterize a new catalase gene encoding a third catalase (KatC). KatC activity was detectable only at the end of the stationary phase in S. meliloti growing in minimum medium, whereas KatA activity was found during the exponential phase. Analysis with a katC-lacZ fusion demonstrated that katC expression is mainly regulated at the transcription level. An increase of catalase activity correlating with KatA induction was detected in bacteroids. A dramatic decrease of nitrogen fixation capacity in a katA katC double mutant was observed, suggesting that these catalases are very important for the protection of the nitrogen fixation process.