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SummaryIn forests ecosystems, fungi and bacteria are key actors in wood degradation. However, few studies have focused on the impact of fungal pre-degradationon bacterial and fungal communities. Coniferous forests are dominated by Brown rot fungi that are involved in earliest phase of lignocellulose breakdown, and therefore, influencing the second phase of microbial colonization. This study aimed to investigate the first microbial species colonizing after intermediate and advanced pre-degradation by the brown rot fungus Gloeophyllum trabeum. Using Illumina metabarcoding, bacterial and fungal communities were monitored after 70 days from Picea abies wood blocks placed between forest floor and topsoil. Chemical composition of the wood samples was determined for each of the pre-degradation stages. We observed significant changes in the bacterial and fungal communities associated with the pre-degradation of P. abies, and that wood substrate condition acted as a gatekeeper for both microbial communities. Our data also suggested that fungal and bacterial communities could interact and act in a synergistic way during the wood decomposition process.

Fomitiporia mediterranea M. Fisch. (Fmed) is a basidiomycete first described in 2002, and was considered up to then as part of Fomitiporia punctata (P. Karst) Murrill. This fungus can degrade lignocellulosic biomass, causing white rot and leaving bleached fibrous host residues. In Europe Fmed is considered the main grapevine wood rot (Esca) agent within the Esca disease complex, which includes some of the most economically important Grapevine Trunk Diseases (GTDs). This review summarises and evaluates published research on Fmed, on white rot elimination by curettage or management by treatments with specific products applied to diseased grapevines, and on the relationship between wood symptoms and Grapevine Leaf Stripe Disease (GLSD) in the Esca disease complex. Information is also reviewed on the fungus biology, mechanisms of pathogenicity, and their possible relationships with external foliar symptoms of the Esca disease complex. Information on Fmed control strategies is also reviewed.

Eutypa dieback and Esca are serious grapevine trunk diseases (GTDs) caused by fungal consortia causing large economic losses in vineyards. Depending on the disease the species involved include Eutypa lata, Phaeoacremonium minimum, and Phaeomoniella chlamydospora. There is a need to understand the complex pathogenesis mechanisms used by these causative fungi to develop treatments for the diseases they cause. Low molecular weight metabolites (LMW) are known to be involved in non-enzymatic oxygen radical generation in fungal degradation of wood by some Basidiomycota species, and as part of our work to explore the basis for fungal consortia pathogenesis, LMW metabolite involvement by the causal GTD fungi was explored. The GTD fungal pathogens examined, Eutypa lata, Phaeoacremonium minimum and Phaeomoniella chlamydospora, were found to produce low molecular weight iron binding metabolites that preferentially reduced iron or redox cycled to produce hydrogen peroxide. Uniquely, different LMW metabolites isolated from the GTD fungi promoted distinct chemistries that are important in a type of non-enzymatic catalysis known as chelator-mediated Fenton (CMF) reactions. CMF chemistry promoted by LMW metabolites from these fungi allowed for the generation of highly reactive hydroxyl radicals under conditions promoted by the fungi. We hypothesize that this new reported mechanism may help to explain the necrosis of woody grapevine tissue as a causal mechanism important in pathogenesis in these two grapevine trunk diseases. IMPORTANCEUnderstanding the pathogenesis of grape trunk diseases (GTDs) is the key to the development of disease control and treatment. While fungal extracellular enzyme systems are typically cited relative to their fungal mechanisms in pathogenesis, non-enzymatic mechanisms have been less studied in this regard and the role of low molecular weight (LMW) fungal metabolites in GTD development is quite limited. In this article, we demonstrate that GTD-causative fungal pathogens Eutypa lata, Phaeoacremonium minimum and Phaeomoniella chlamydospore produce LMW phenolic metabolites under iron-restricted conditions. These metabolites undergo a series of redox reactions, with different fungi producing metabolites that preferentially either reduce iron, or generate hydrogen peroxide, under conditions simulating grapevine woody tissue. These conditions have the potential to promote generation of highly damaging hydroxyl radicals through a mechanism that appears to be similar to non-enzymatic chelator-mediated Fenton (CMF) chemistry which is involved in fungal degradation of wood by non-related fungal orders. This is the first report of CMF chemistry promoted by GTD-causative fungi under laboratory conditions and the research suggests an alternate pathway that may contribute to pathogenesis in GTDs, and a potential target for vine protection.

The major Grapevine Trunk Diseases (GTD) gather diseases associated with either one particular fungal species, e.g., eutypiose, or with fungal species complexes, e.g., Botryosphaeriae dieback. GTD-associated fungi are part of different fungal classes, depending on their substrate degradation mechanisms or on their lifestyles. They are identified in grapevine wood and they lead to wood typical symptoms. They are sometimes associated to foliar symptoms too, despite not found in aerial parts, probably acting through fungal toxins or through physiological disorders, such as non-functional wood vessels. Grapevine is a liana, and as such shows structural and chemical differences compared to forest tree wood. However, knowledge on wood-degrading fungi growing on tree wood can contribute to a better understanding of grapevine wood adaptation mechanisms to fungal attack. For example, wood density, xylem vessels size and number, or lignin content and composition can play a role in wood sensitivity to fungi. Also, some fungal detoxification mechanisms against wood extractibles are host-specific. In a first part of this chapter, structure and chemical composition of grapevine wood are described, pointing out properties that might play a role in fungal growth. In a second part, GTD main associated fungi and their known mechanisms for wood degradation or wood chemical detoxification are discussed. Extracellular and intracellular enzymatic and non-enzymatic mechanisms are considered. These mechanisms, well described for their role in fungal adaptation to tree wood, have been much less studied for GTD associated fungi. Further studies about those mechanisms would contribute to understand fungal adaptation to grapevine.

International audience; Secreted proteins are key players in fungal physiology and cell protection against external stressing agents and antifungals. Oak stress-induced protein 1 (OSIP1) is a fungal-specific protein with unknown function. By using Podospora anserina and Phanerochaete chrysosporium as models, we combined both in vivo functional approaches and biophysical characterization of OSIP1 recombinant protein. The P. anserina OSIP1(Delta) mutant showed an increased sensitivity to the antifungal caspofungin compared to the wild type. This correlated with the production of a weakened extracellular exopolysaccharide/protein matrix (ECM). Since the recombinant OSIP1 from P. chrysosporium self-assembled as fibers and was capable of gelation, it is likely that OSIP1 is linked to ECM formation that acts as a physical barrier preventing drug toxicity. Moreover, compared to the wild type, the OSIP1(Delta) mutant was more sensitive to oak extractives including chaotropic phenols and benzenes. It exhibited a strongly modified secretome pattern and an increased production of proteins associated to the cell-wall integrity signalling pathway, when grown on oak sawdust. This demonstrates that OSIP1 has also an important role in fungal resistance to extractive-induced stress.

The bacteria of the genus Streptomyces and Basidiomycete fungi harbor many biosynthetic gene clusters (BGCs) that are at the origin of many bioactive molecules with medical or industrial interests. Nevertheless, most BGCs do not express in standard lab growth conditions, preventing the full metabolic potential of these organisms from being exploited. Because it generates biotic cues encountered during natural growth conditions, co-culture is a means to elicit such cryptic compounds. In this study, we explored 72 different Streptomyces-fungus interaction zones (SFIZs) generated during the co-culture of eight Streptomyces and nine fungi. Two SFIZs were selected because they showed an elicitation of anti-bacterial activity compared to mono-cultures. The study of these SFIZs showed that co-culture had a strong impact on the metabolic expression of each partner and enabled the expression of specific compounds. These results show that mimicking the biotic interactions present in this ecological niche is a promising avenue of research to explore the metabolic capacities of Streptomyces and fungi.

The understanding of soil organic matter (SOM) dynamics has considerably advanced in recent years. It was previously assumed that most SOM consisted of recalcitrant compounds, whereas the emerging view considers SOM as a range of polymers continuously processed into smaller molecules by decomposer enzymes. Mainstreaming these new insights in current models is challenging because of their ill-adapted framework. We propose the C-STABILITY model to resolve this issue. Its innovative framework combines compartmental and continuous modelling approaches to accurately reproduce SOM cycling processes. Model theoretical simulations highlight the role of SOM accessibility on its turnover. They reveal how enzyme depolymerization, decomposer community succession and carbon use efficiency are key drivers of SOM composition and quantity during decomposition and at steady-state. The mathematical structure of C-STABILITY may be tailored to different scales and is open to alternative formulations, thus offering a promising foundation for enhancing carbon cycling predictions.

The eukaryotic translation elongation factor 1Bγ (eEF1Bγ) is an atypical member of the glutathione transferase (GST) superfamily. Contrary to more classical GSTs having a role in toxic compound detoxification, eEF1Bγ is suggested to act as a scaffold protein, anchoring the elongation factor complex EF1B to the endoplasmic reticulum. In this study, we show that eEF1Bγ from the basidiomycete Phanerochaete chrysosporium is fully active as a glutathione transferase in vitro and undergoes conformational changes upon binding of oxidized glutathione. Using real-time analyses of biomolecular interactions, we show that GSSG allows eEF1Bγ to physically interact with other GSTs from the Ure2p class, opening new perspectives for a better understanding of the role of eEF1Bγ in cellular oxidative stress response.

Extensive evidence showed that the efficiency of fungal wood degradation is closely dependent on their ability to cope with the myriad of putative toxic compounds called extractives released during this process. By analysing global gene expression of Phanerochaete chrysosporium after short oak extractive treatment (1, 3 and 6 h), we show that the early molecular response of the fungus concerns first mitochondrial stress rescue followed by the oxidation and finally conjugation of the compounds. During these early responses, the lignolytic degradative system is not induced, rather some small secreted proteins could play an important role in cell protection or signaling. By focusing on the functional characterization of an hitherto uncharacterized glutathione transferase, we show that this enzyme interacts with wood molecules suggesting that it could be involved in the detoxification of some of them, or act as a scavenger to prevent their cytosolic toxicity and favour their transport.

International audience; The natural durability of wood is linked to its chemical composition and in particular the presence of metabolites called extractives that often possess chemical reactivity. For dealing with these compounds, wood degraders have developed detoxification systems usually involving enzyme families. Among these enzymes, glutathione transferases (GSTs) are involved in the decrease of the reactivity of toxic compounds. In this study, the hypothesis that the detoxification systems of wood decaying fungi could be indicators of the chemical reactivity of wood extracts has been tested. This approach has been evaluated using 32 wood extracts coming from French Guiana species, testing their antimicrobial ability, antioxidative properties, and reactivity against six GSTs from the white rot Trametes versicolor. From the obtained data, a significant correlation between the antimicrobial and antioxidative properties of the tested wood extracts and GST interactions was established. In addition, the chemical analysis performed on one of the most reactive extracts (an acetonic extract of Bagassa guianensis) has demonstrated oxyresveratrol as a major constituent. We were able to cocrystallize one GST with this commercially interesting compound. Taken together, the presented data support the hypothesis that detoxifying enzymes could be used to identify the presence of molecules of industrial interest in wood extracts.

Extractives are non-structural wood molecules that represent a minor fraction in wood.However, they are source of diverse molecules putatively bioactive. Inhibition of fungal growth is one of the most interesting properties of wood extractives in a context of wood preservation, crop protection or medical treatments.The antifungal effect of molecules isolated from wood extractives has been mainly attributed to various mechanisms such as metal and free radical scavenging activity, direct interaction with enzymes, disruption of membrane integrity and perturbation of ionic homeostasis. Lignolytic fungi, which are microorganisms adapted to wood substrates, have developed various strategies to protect themselves against this toxicity.A better knowledge of these strategies could help both developing new systems for extractive removal in biomass valorization processes and using these molecules as antifungal agents. (C) 2017 British Mycological Society.

The Target Of Rapamycin (TOR) signaling pathway is known to regulate growth in response to nutrient availability and stress in eukaryotic cells. In the present study, we have investigated the TOR pathway in the white-rot fungus Phanerochaete chrysosporium. Inhibition of TOR activity by rapamycin affects conidia germination and hyphal growth highlighting the conserved mechanism of susceptibility to rapamycin. Interestingly, the secreted protein content is also affected by the rapamycin treatment. Finally, homologs of the components of TOR pathway can be identified in P. chrysosporium. Altogether, those results indicate that the TOR pathway of P. chrysosporium plays a central role in this fungus.

Trametes versicolor glutathione transferase Omega 3S (TvGSTO3S) catalyzes the conjugation of isothiocyanates (ITC) with glutathione (GSH).Previously, this isoform was investigated in depth both biochemically and structurally. Structural analysis of complexes revealed the presence of a GSH binding site (G site) and a deep hydrophobic binding site (H site) able to bind plant polyphenols. In the present study, crystals of apo TvGSTO3S were soaked with glutathionyl-phenethylthiocarbamate, the product of the reaction between GSH and phenethyl isothiocyanate (PEITC).On the basis of this crystal structure, we show that the phenethyl moiety binds in a new site at loop beta(2)-alpha(2) while the glutathionyl part exhibits a particular conformation that occupies both the G site and the entrance to the H site. This binding mode is allowed by a conformational change of the loop beta(2)-alpha(2) at the enzyme active site. It forms a hydrophobic slit that stabilizes the phenethyl group at a distinct site from the previously described H site.Structural comparison of TvGSTO3S with drosophila DmGSTD2 suggests that this flexible loop could be the region that binds PEITC for both isoforms. These structural features are discussed in a catalytic context.

International audience; Key message Teak (Tectona grandis L.f.) is one of the most important tropical hardwood tree species, which is widely planted in Indonesia. Wood properties are strongly influenced by forest management conditioning further utilization of wood.Context In Indonesia, teak wood has been supplied from the state forests (Perhutani) for long rotation teak and from community teak plantations for short rotation teak. Short rotation teak has been harvested at 7-10 years and long rotation teak at 40-60 years.Aims This paper discusses the characterization of technical properties of short and long rotation teak wood based on the chemical, anatomical, physical, and mechanical properties.Methods The properties of short rotation and long rotation teak woods were characterized by measuring their density, extractive contents, chemical composition, swelling, wettability, water sorption isotherm, decay resistance, anatomical properties, bending strength (modulus of rupture (MOR), modulus of elasticity (MOE)), and hardness.Results The results indicate that short rotation teak was not particularly different in swelling, MOE and MOR, and Brinell hardness compared to long rotation teak, although it was less dense and less durable due to lower heartwood and extractive contents. Therefore, careful attention should be given to the use of short rotation teak in some wood-processing technologies.Conclusion Lower wood density and durability of the short rotation compared to the long rotation teak will restrict its utilization to some extent for both indoor and outdoor applications. Fast-growing teak from community cannot be used as usual heartwood teak from Perhutani because of the very low proportion of useful heartwood in the stem.

Trametes versicolor is a wood-inhabiting agaricomycete known for its ability to cause strong white-rot decay on hardwood and for its high tolerance of phenolic compounds. The goal of the present work was to gain insights into the molecular biology and biochemistry of the heme-including class II and dye-decolorizing peroxidases secreted by this fungus. Proteomic analysis of the secretome of T. versicolor BRFM 1218 grown on oak wood revealed a set of 200 secreted proteins, among which were the dye-decolorizing peroxidase Tv DyP1 and the versatile peroxidase Tv VP2. Both peroxidases were heterologously produced in Escherichia coli , biochemically characterized, and tested for the ability to oxidize complex substrates. Both peroxidases were found to be active against several substrates under acidic conditions, and Tv DyP1 was very stable over a relatively large pH range of 2.0 to 6.0, while Tv VP2 was more stable at pH 5.0 to 6.0 only. The thermostability of both enzymes was also tested, and Tv DyP1 was globally found to be more stable than Tv VP2. After 180 min of incubation at temperatures ranging from 30 to 50°C, the activity of Tv VP2 drastically decreased, with 10 to 30% of the initial activity retained. Under the same conditions, Tv DyP1 retained 20 to 80% of its enzyme activity. The two proteins were catalytically characterized, and Tv VP2 was shown to accept a wider range of reducing substrates than Tv DyP1. Furthermore, both enzymes were found to be active against two flavonoids, quercetin and catechin, found in oak wood, with Tv VP2 displaying more rapid oxidation of the two compounds. They were tested for the ability to decolorize five industrial dyes, and Tv VP2 presented a greater ability to oxidize and decolorize the dye substrates than Tv DyP1. IMPORTANCE Trametes versicolor is a wood-inhabiting agaricomycete known for its ability to cause strong white-rot decay on hardwood and for its high tolerance of phenolic compounds. Among white-rot fungi, the basidiomycete T. versicolor has been extensively studied for its ability to degrade wood, specifically lignin, thanks to an extracellular oxidative enzymatic system. The corresponding oxidative system was previously studied in several works for classical lignin and manganese peroxidases, and in this study, two new components of the oxidative system of T. versicolor , one dye-decolorizing peroxidase and one versatile peroxidase, were biochemically characterized in depth and compared to other fungal peroxidases.

Glutathione transferases (GSTs) from the Xi and Omega classes have a catalytic cysteine residue, which gives them reductase activities. Until now, they have been assigned distinct substrates. While Xi GSTs specifically reduce glutathionyl-(hydro)quinones, Omega GSTs are specialized in the reduction of glutathionyl-acetophenones. Here, we present the biochemical and structural analysis of TvGSTX1 and TvGSTX3 isoforms from the wood-degrading fungus Trametes versicolor. TvGSTX1 reduces GS-menadione as expected, while TvGSTX3 reduces both Xi and Omega substrates. An in-depth structural analysis indicates a broader active site for TvGSTX3 due to specific differences in the nature of the residues situated in the C-terminal helix α9. This feature could explain the catalytic duality of TvGSTX3. Based on phylogenetic analysis, we propose that this duality might exist in saprophytic fungi and ascomycetes.

Trametes versicolor is a wood-inhabiting agaricomycete known for its ability to cause strong white-rot decay on hardwood and for its high tolerance of phenolic compounds. The goal of the present work was to gain insights into the molecular biology and biochemistry of the heme-including class II and dye-decolorizing peroxidases secreted by this fungus. Proteomic analysis of the secretome of T. versicolor BRFM 1218 grown on oak wood revealed a set of 200 secreted proteins, among which were the dye-decolorizing peroxidase TvDyP1 and the versatile peroxidase TvVP2. Both peroxidases were heterologously produced in Escherichia coli, biochemically characterized, and tested for the ability to oxidize complex substrates. Both peroxidases were found to be active against several substrates under acidic conditions, and TvDyP1 was very stable over a relatively large pH range of 2.0 to 6.0, while TvVP2 was more stable at pH 5.0 to 6.0 only. The thermostability of both enzymes was also tested, and TvDyP1 was globally found to be more stable than TvVP2. After 180 min of incubation at temperatures ranging from 30 to 50°C, the activity of TvVP2 drastically decreased, with 10 to 30% of the initial activity retained. Under the same conditions, TvDyP1 retained 20 to 80% of its enzyme activity. The two proteins were catalytically characterized, and TvVP2 was shown to accept a wider range of reducing substrates than TvDyP1. Furthermore, both enzymes were found to be active against two flavonoids, quercetin and catechin, found in oak wood, with TvVP2 displaying more rapid oxidation of the two compounds. They were tested for the ability to decolorize five industrial dyes, and TvVP2 presented a greater ability to oxidize and decolorize the dye substrates than TvDyP1.

International audience; Barks are available as waste material and by-product of wood industry. They have been reported to contain interesting molecules and show some bioactivity such as antioxidant and antifungal. This study aimed at evaluating the amounts of extractives in Acacia mangium (acacia), Poraserianthes falcataria (sengon) and Swietenia mahagoni (mahoni) barks, to evaluate their extractive contents and the presence of potential valuable molecules. The extraction method used soxhlet with four different solvents. Antioxidant activity assays were carried out using methyl linoleate and 2,2-diphenyl-1-picrilhidrazyl (DPPH) and the antifungal activity was determinate by fungal growth inhibitions assays. 5.3%-18.5% extraction yields were obtained. All acetone and toluene ethanol extracts show high antioxidant activity by DPPH. The highest antioxidant value obtained by DPPH was obtained for mahoni bark acetone extract with 3.9 mg/L of EC50, followed by mahoni bark toluene ethanol 6.8 mg/L, acacia bark acetone 7 mg/L, and acacia bark toluene ethanol extract 7.4 mg/L. Sengon bark extracts had the greatest antifungal activity inhibition. The greatest antioxidant and antifungal activity were obtained with phenolic compounds which were contained in the extracts.

International audience; The intracellular systems of detoxification are crucial for the survival of wood degrading fungi. Within these systems, glutathione transferases could play a major role since this family of enzymes is specifically extended in lignolytic fungi. In particular the Ure2p class represents one third of the total GST number in Phanerochaete chrysosporium. These proteins have been phylogenetically split into two subclasses called Ure2pA and Ure2pB. Ure2pB can be classified as Nu GSTs because of shared structural and functional features with previously characterized bacterial isoforms. Ure2pA can rather be qualified as Nu-like GSTs since they exhibit a number of differences. Ure2pA possess a classical transferase activity, a more divergent catalytic site and a higher structural flexibility for some of them, compared to Nu GSTs. The characterization of four members of this Ure2pA subclass (PcUre2pA4, PcUre2pA5, PcUre2pA6 and PcUre2pA8) revealed specific functional and structural features, suggesting that these enzymes have rapidly evolved and differentiated, probably to adapt to the complex chemical environment associated with wood decomposition.

International audience; Glutathionyl-hydroquinone reductases (GHRs) catalyze the deglutathionylation of quinones via a catalytic cysteine. The two GHR genes in the Populus trichocarpa genome, Pt-GHR1 and Pt-GHR2, are primarily expressed in reproductive organs. Both proteins are localized in plastids. More specifically, Pt-GHR2 localizes in nucleoids. At the structural level, Pt-GHR1 adopts a typical GHR fold, with a dimerization interface comparable to that of the bacterial and fungal GHR counterparts. Pt-GHR1 catalyzes the deglutathionylation of both reduced and oxidized glutathionylated quinones, but the enzyme is more catalytically efficient with the reduced forms.

International audience; Glutathione transferases (GSTs) constitute a widespread super-family of enzymes with highly specific to overlapping functions. These enzymes have been extensively studied in mammals because of their roles in xenobiotic conjugation. Despite this, GST functions are largely unexplored, especially in microorganisms. Our work focused on fungal glutathionyl-hydroquinone reductases (GHRs). These singular GSTs catalyze the reduction of hydroquinones-SG (Xun et al., 2010), and are present in all kingdoms but animals. The first GHR structure to be described in the saprotrophic fungus Phanerochaete chrysosporium (PcGHR1) has unveiled original features. An original dimerization mode led us to establish a new structural class named Xi (GSTX) (Meux et al., 2011). Here we report the crystal structures of two GHRs, TvGHR1 from the basidiomycete Trametes versicolor and ScECM4 from the yeast Saccharomyces cerevisiae, at 2.28 and 1.45 Å resolution, respectively. TvGHR1 and ScECM4 share the classical GST fold which consists in a thioredoxin N-terminal domain and an all α C-terminal domain. Besides the features that have been uncovered in PcGHR1, additional secondary structures were found in the yeast ScECM4 structure. Like its characterized homologs, ScEcM4 is specific to the reduction of quinones conjugated with glutathione (Lallement et al., 2014). TvGHR1 is more polyvalent because it also reduces substrates rather accepted by Omega-class GSTs like phenylacetophenone-SG. Compared with known GHR structures, TvGHR1 exhibits a larger substrate binding site with an original motif that could explain the substrate versatility. Phylogenetic analyses revealed that fungal GHRs could be separated in two families : the ‘classic’ GHRs and the ‘atypical’ GHRs that exhibit motif variations as observed in TvGHR1. Furthermore, the conservation of GHRs in fungi suggests that they could fulfill a crucial role. Their diversification in wood-degrading basidiomycetes agrees with the hypothesis that GHRs would play a role in the metabolism of phenolic compounds (Belchik & Xun, 2011).

Small secreted proteins (SSP) have been defined as proteins containing a signal peptide and a sequence of less than 300 amino acids. In this analysis, we have compared the secretion pattern of SSPs among eight aspergilli species in the context of plant biomass degradation and have highlighted putative interesting candidates that could be involved in the degradative process or in the strategies developed by fungi to resist the associated stress that could be due to the toxicity of some aromatic compounds or reactive oxygen species released during degradation. Among these candidates, for example, some stress-related superoxide dismutases or some hydrophobic surface binding proteins (HsbA) are specifically secreted according to the species.[br/] Since these latter proteins are able to recruit lytic enzymes to the surface of hydrophobic solid materials and promote their degradation, a synergistic action of HsbA with the degradative system may be considered and need further investigations.[br/] These SSPs could have great applications in biotechnology by optimizing the efficiency of the enzymatic systems for biomass degradation.

Key message This paper describes how the combination of genomics, genetic engineering, and 3D structural characterization has helped clarify the redox regulatory networks in poplar with consequences not only in system biology in plants but also in bacteria and mammalian systems.[br/] Context Tree genomes are increasingly available with a large number of orphan genes coding for proteins, the function of which is still unknown.[br/] Aims and methods Modern techniques of genome analysis coupled with recombinant protein technology and massive 3D structural determination of tree proteins should help elucidate the function of many of the proteins encoded by orphan genes. X-ray crystallography and NMR will be the methods of choice for protein structure determination.[br/] Results In this review, we provide examples illustrating how the above-mentioned techniques improved our understanding of redox regulatory circuits in poplar, the first forest tree species sequenced. We showed that poplar peroxiredoxins use either thioredoxin or glutaredoxin as electron donors to reduce hydrogen peroxide. That glutaredoxin could be a reductant was unknown at the time of this discovery even in other biological organisms and was later confirmed notably by the observation that the two genes are fused in some bacteria and by the resolution of the structure of the bacterial hybrid protein. Similarly, genome analysis coupled to in vitro analysis of enzymatic properties led to the discovery that some plant methionine sulfoxide reductases can also use both thioredoxins and glutaredoxins as electron donors. Besides their disulfide reductase activity, it has been demonstrated that some poplar glutaredoxins are also involved in iron-sulfur center biogenesis and assembly. The original 3D structure determination has been made with poplar glutaredoxin C1 and then confirmed in a variety of other biological organisms including human. Our work also showed that in plants, so-called glutathione peroxidases use thioredoxins and not glutathione as electron donors. This is true for all non-selenocysteine-containing glutathione peroxidases. Finally, connections between the thioredoxin and glutaredoxin systems have been elucidated through the study of atypical poplar thioredoxins.[br/] Conclusion Altogether, these data illustrate how the combination of genetic engineering and structural biology improves our understanding of biological processes and helps fuel systems biology for trees and other biological species.

International audience; Abstract Key messageContent and type of extractives vary considerably among woody species in the Congo Basin tropical forest. They influence the natural durability of wood with respect to fungal and insect degradation. Species containing higher amounts of extractives and Klason lignin were generally less susceptible to fungal colonization. Phenolic extractives with large anti-fungal and high antioxidant activities were the main chemical family of compounds extracted from the tested species, which suggests a potential chemical valorization of these extractives.ContextSeveral woody species from the Congo Basin are known for their natural durability, but the causes of this natural durability are not always fully elucidated. This is particularly the case for the resistance to white rotting fungi decay.AimsThe chemical composition as well as the antioxidant activity of the heartwood extractives of several species were quantified and correlated to the ability of different fungi to colonize wood sawdusts in order to better understand their decay resistance.MethodsThe chemical screening of extracts was conducted using colorimetric methods. The quantity of lignin, extracts and total phenolic compounds, as well as antioxidant activities were determined. Extracted and unextracted heartwood sawdusts were exposed to two white rot fungi to assess the effect of extractives on their colonization.ResultsExtractives and lignin contents ranged from 2.7 to 16.0 % and from 26.5 to 35.9 %, respectively. Antioxidant activity was directly connected to total phenolic contents. The phytochemical screening revealed the presence of different chemical groups in all extracts. Resistance to fungi was correlated to the amount of extractives and in most cases to the lignin content.ConclusionPhenolic compounds associated with antioxidant activities influenced the natural resistance of heartwood.

Background Glutathione transferases (GSTs) represent an extended family of multifunctional proteins involved in detoxification processes and tolerance to oxidative stress. We thus anticipated that some GSTs could play an essential role in the protection of fungal necrotrophs against plant-derived toxic metabolites and reactive oxygen species that accumulate at the host-pathogen interface during infection. Results Mining the genome of the necrotrophic Brassica pathogen Alternaria brassicicola for glutathione transferase revealed 23 sequences, 17 of which could be clustered into the main classes previously defined for fungal GSTs and six were ‘orphans’. Five isothiocyanate-inducible GSTs from five different classes were more thoroughly investigated. Analysis of their catalytic properties revealed that two GSTs, belonging to the GSTFuA and GTT1 classes, exhibited GSH transferase activity with isothiocyanates (ITC) and peroxidase activity with cumene hydroperoxide, respectively. Mutant deficient for these two GSTs were however neither more susceptible to ITC nor less aggressive than the wild-type parental strain. By contrast mutants deficient for two other GSTs, belonging to the Ure2pB and GSTO classes, were distinguished by their hyper-susceptibility to ITC and low aggressiveness against Brassica oleracea. In particular AbGSTO1 could participate in cell tolerance to ITC due to its glutathione-dependent thioltransferase activity. The fifth ITC-inducible GST belonged to the MAPEG class and although it was not possible to produce the soluble active form of this protein in a bacterial expression system, the corresponding deficient mutant failed to develop normal symptoms on host plant tissues. Conclusions Among the five ITC-inducible GSTs analyzed in this study, three were found essential for full aggressiveness of A. brassicicola on host plant. This, to our knowledge is the first evidence that GSTs might be essential virulence factors for fungal necrotrophs. Electronic supplementary material The online version of this article (doi:10.1186/s12866-015-0462-0) contains supplementary material, which is available to authorized users.

Deroy, Aurelie Saiag, Fanny Kebbi-Benkeder, Zineb Touahri, Nassim Hecker, Arnaud Morel-Rouhier, Melanie Colin, Francis Dumarcay, Stephane Gerardin, Philippe Gelhaye, Eric; White-rot fungi possess the unique ability to degrade and mineralize all the different components of wood. In other respects, wood durability, among other factors, is due to the presence of extractives that are potential antimicrobial molecules. To cope with these molecules, wood decay fungi have developed a complex detoxification network including glutathione transferases (GST). The interactions between GSTs from two white-rot fungi, Trametes versicolor and Phanerochaete chrysosporium, and an environmental library of wood extracts have been studied. The results demonstrate that the specificity of these interactions is closely related to the chemical composition of the extracts in accordance with the tree species and their localization inside the wood (sapwood vs heartwood vs knotwood). These data suggest that the fungal GSTome could reflect the chemical environment encountered by these fungi during wood degradation and could be a way to study their adaptation to their way of life.

Q beta phages infect Escherichia coli in the human gut by recognizing F-pili as receptors. Infection therefore occurs under reducing conditions induced by physiological agents (e.g. glutathione) or the intestinal bacterial flora. After excretion in the environment, phage particles are exposed to oxidizing conditions and sometimes disinfection. If inactivation does not occur, the phage may infect new hosts in the human gut through the oral route. During such a life cycle, we demonstrated that, outside the human gut, cysteines of the major protein capsid of Q beta phage form disulfide bonds. Disinfection with NaClO does not allow overoxidation to occur. Such oxidation induces inactivation rather by irreversible damage to the minor proteins. In the presence of glutathione, most disulfide bonds are reduced, which slightly increases the capacity of the phage to infect E. coli in vitro. Such reduction is reversible and barely alters infectivity of the phage. Reduction of all disulfide bonds by dithiothreitol leads to complete capsid destabilization. These data provide new insights into how the phages are impacted by oxidizing-reducing conditions outside their host cell and raises the possibility of the intervention of the redox during life cycle of the phage.Physiological redox conditions modify disulfide bonds of the capsid of Q beta phage without decreasing infectivity, whereas total reduction disrupts the capsid and chlorine oxidation disrupts especially minor proteins.Physiological redox conditions modify disulfide bonds of the capsid of Q beta phage without decreasing infectivity, whereas total reduction disrupts the capsid and chlorine oxidation disrupts especially minor proteins.