Article: Soil Biology and Biochemistry

Tree roots select specific bacterial communities in the subsurface critical zone O Nicolitch, Y Colin, MP Turpault, L Fauchery, S Uroz. Soil Biology and Biochemistry 115, 109-123

Abstract

In soils characterized by nutrient-poor conditions trees have developed strategies to maximize the exploration of the environment through their root system. Notably, in shallow soils, trees adopt a deep-rooting strategy to access appropriate levels of water and nutrients from the bedrock. Despite the critical importance of microorganisms in nutrient access in topsoil, understanding their involvement in subsoil was rarely addressed. Our study provides the first comprehensive picture of the bacterial communities colonizing deep roots at the bedrock interface. Particularly, we aimed at deciphering if the subsoil edaphic conditions allowed the enrichment of specific bacterial communities in the rhizosphere. To answer such questioning, we focused on a shallow soil dominated by deep-rooting beech trees (Fagus sylvatica). The taxonomic and functional structures of bacterial communities were investigated through 16S rRNA-pyrosequencing analyses and in vitro bioassays on culturable representatives isolated from the saprolite, the limestone rocks and the roots penetrating those two compartments at the bedrock interface. Our taxonomic analyses revealed a rhizosphere effect, with no difference between the limestone- and saprolite-rhizosphere bacterial communities. Notably, our functional assays highlighted a significant enrichment of bacteria effective at mineral weathering in the limestone-rhizosphere compared to the surrounding environment, whereas organic matter decomposing bacteria were exclusively enriched in the saprolite-rhizosphere. Altogether our results suggest that tree roots select specific bacterial communities in subsoil as potential allies to improve nutrient availability and tree nutrition.

Article: J Plant Biochem Physiol

Covalent and Non-Covalent Associations Mediate MED28 Homo J Shaikhali, N Rouhier, A Hecker, K Brännström, G Wingsle.  J Plant Biochem Physiol 5:189. doi:10.4172/2329-9029.1000189

Abstract

The Mediator is a multi-protein complex that plays a key role in modulating gene expression. Our previous studies suggested that the MED10a, MED28, MED32 complex subunits could be subject to redox regulation. In this study we tested the capacity of different thioredoxins (TRXs) from poplar (TRX-H3 and TRX-H5) and Arabidopsis thaliana (TPR repeat-containing thioredoxin, TDX) as well as glutaredoxins (GRXs) from poplar (GRX-C3 and GRX-C4) to reduce MED28 oligomers in vitro and found that these proteins were less efficient than the the previously tested poplar TRX-H1 and Arabidopsis GRX-C1. Concerning the susceptibility of MED28 to oxidation, both hydrogen peroxide (H2O2) and glutathione disulfide (GSSG) are efficiently mediating the formation of intermolecular disulfides. In fact, MED28 forms homo- oligomers in vivo as assessed by yeast two-hybrid experiments but also in vitro in solution as shown by size-exclusion chromatography, the latter also demonstrated the formation of non- covalent homo-oligomers. These findings suggest that both the redox-dependent and – independent MED28 oligomerization could regulate its biological activities, could it be linked or not to the Mediator. In particular, it would be important to assess MED28 oligomerization state during senescence considering the previously observed phenotype of med28 plants.

Article: Photorespiration

In Vitro Alkylation Methods for Assessing the Protein Redox State. F Zannini, J Couturier, O Keech, N Rouhier. Photorespiration, 51-64

Abstract

Cysteines are important residues for protein structure, function, and regulation. Owing to their modified reactivity, some cysteines can undergo very diverse redox posttranslational modifications, including the reversible formation of disulfide bonds, a widespread protein regulatory process as well exemplified in plant chloroplasts for Calvin-Benson cycle enzymes. Both core- and peripheral-photorespiratory enzymes possess conserved cysteines, some of which have been identified as being subject to oxidative modifications. This is not surprising considering their presence in subcellular compartments where the production of reactive species can be important. However, in most cases, the types of modifications and their biochemical effect on protein activity have not been validated, meaning that the possible impact of these modifications in a complex physiological context, such as photorespiration, remains obscure.

We here describe a detailed set of protocols for alkylation methods that have been used so far to (1) study the protein cysteine redox state either in vitro by submitting purified recombinant proteins to reducing/oxidation treatments or in vivo by western blots on protein extracts from plants subject to environmental constraints, and its dependency on the two major reducing systems in the cell, i.e., the thioredoxin and glutathione/glutaredoxin systems, and (2) determine two key redox parameters, i.e., the cysteine pKa and the redox midpoint potential.

Article: Plant, Cell & Environment

Involvement of Arabidopsis glutaredoxin S14 in the maintenance of chlorophyll content P Rey, N Becuwe, S Tourrette, N Rouhier Plant, Cell & Environment

Abstract

Plant class-II glutaredoxins (GRXs) are oxidoreductases carrying a CGFS active site signature and are able to bind iron-sulfur clusters in vitro. In order to explore the physiological functions of the two plastidial class-II isoforms, GRXS14 and GRXS16, we generated knockdown and overexpression Arabidopsis thaliana lines and characterized their phenotypes using physiological and biochemical approaches. Plants deficient in one GRX did not display any growth defect, whereas the growth of plants lacking both was slowed. Plants overexpressing GRXS14 exhibited reduced chlorophyll content in control, high light and high salt conditions. However, when exposed to prolonged darkness, plants lacking GRXS14 showed accelerated chlorophyll loss compared to WT and overexpression lines. We observed that the GRXS14 abundance and the proportion of reduced form were modified in WT upon darkness and high salt. The dark treatment also resulted in decreased abundance of proteins involved in the maturation of iron-sulfur proteins. We propose that the phenotype of GRXS14-modified lines results from its participation in the control of chlorophyll content in relation with light and osmotic conditions, possibly through a dual action e.g. regulating the redox status of biosynthetic enzymes and contributing to the biogenesis of iron-sulfur clusters, which are essential cofactors in chlorophyll metabolism.

Article: Industrial Crops and Products

Characterization of bark extractives of different industrial Indonesian wood species for potential valorization. NA Rosdiana, S Dumarçay, C Gérardin, H Chapuis, FJ Santiago-Medina, … Industrial Crops and Products 108, 121-127

Abstract

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), Paraserianthes 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.

Article: FEBS Journal

Structural plasticity among glutathione transferase Phi members: natural combination of catalytic residues confers dual biochemical activities. H Pégeot, S Mathiot, T Perrot, F Gense, A Hecker, C Didierjean, … The FEBS journal

Abstract

The glutathione transferase (GST) gene family is divided into fourteen classes in photosynthetic organisms. Among them, the Phi class (GSTF) is composed of a large number of genes that are often induced in response to environmental constraints due to their ability to detoxify xenobiotics, to their peroxidase activity and to their involvement in the biosynthesis and/or transport of secondary metabolites. However, the exact functions of GSTFs from many plants including Populus trichocarpa are unknown. Here, following GSTF1 characterization, we have performed a comparative analysis of the seven other GSTFs found in poplar by systematically evaluating the biochemical and enzymatic properties of the corresponding recombinant proteins and of variants mutated for active site residues and by determining the three-dimensional structures of several representatives. Owing to the presence of a cysteine with a pKa value around 5 in their active site, GSTF3, F7 and F8 displayed a thiol transferase activity in addition to the usual glutathione transferase and peroxidase activities. From structural analyses, it appeared that these dual biochemical properties originate from the existence of a certain variability in the β1-α1 loop. This allows positioning of several active site residues at proximity of the glutathione molecule, which itself remains unchanged in GSTF three-dimensional structures. These results highlight the promiscuity of some GSTFs and that changes of active site residues in some isoforms during evolution generated functional diversity by modifying their activity profile.

Article: Science

Ancestral alliances: Plant mutualistic symbioses with fungi and bacteria FM Martin, S Uroz, DG Barker Science 356 (6340), eaad4501

Abstract

Within the plant microbiota, mutualistic fungal and bacterial symbionts are striking examples of microorganisms playing crucial roles in nutrient acquisition. They have coevolved with their hosts since initial plant adaptation to land. Despite the evolutionary distances that separate mycorrhizal and nitrogen-fixing symbioses, these associations share a number of highly conserved features, including specific plant symbiotic signaling pathways, root colonization strategies that circumvent plant immune responses, functional host-microbe interface formation, and the central role of phytohormones in symbiosis-associated root developmental pathways. We highlight recent and emerging areas of investigation relating to these evolutionarily conserved mechanisms, with an emphasis on the more ancestral mycorrhizal associations, and consider to what extent this knowledge can contribute to an understanding of plant-microbiota associations as a whole.

CP Science 2017_

Article: Journal of Proteome Research

iTRAQ and RNA-Seq analyses provide new insights into regulation mechanism of symbiotic germination of Dendrobium officinale seeds (Orchidaceae). J Chen, S Liu, A Kohler, B Yan, HM Luo, X Chen, SX Guo. Journal of Proteome Research

Abstract

Mycorrhizal fungi colonize orchid seeds and induce germination. This so-called symbiotic germination is a critical developmental process in the lifecycle of all orchid species. However, the molecular changes that occur during orchid seed symbiotic germination remain largely unknown. To better understand the molecular mechanism of orchid seed germination, we performed a comparative transcriptomic and proteomic analysis of the Chinese traditional medicinal orchid Dendrobium officinale to explore the change in protein expression at the different developmental stages during asymbiotic and symbiotic germination and identify the key proteins that regulate the symbiotic germination of orchid seeds. Among 2256 identified plant proteins, 308 were differentially expressed across three developmental stages during asymbiotic and symbiotic germination, and 229 were differentially expressed during symbiotic germination compared to asymbiotic development. Of these, 32 proteins were coup-regulated at both the proteomic and transcriptomic levels during symbiotic germination compared to asymbiotic germination. Our results suggest that symbiotic germination of D. officinale seeds shares a common signaling pathway with asymbiotic germination during the early germination stage. However, compared to asymbiotic germination, fungal colonization of orchid seeds appears to induce higher and earlier expression of some key proteins involved in lipid and carbohydrate metabolism and thus improves the efficiency of utilization of stored substances present in the embryo. This study provides new insight into the molecular basis of orchid seed germination.

Article: Scientific reports

HqiA, a novel quorum-quenching enzyme which expands the AHL lactonase family.M Torres, S Uroz, R Salto, L Fauchery, E Quesada, I Llamas Scientific reports 7 (1), 943

Abstract

The screening of a metagenomic library of 250,000 clones generated from a hypersaline soil (Spain) allowed us to identify a single positive clone which confers the ability to degrade N-acyl homoserine lactones (AHLs). The sequencing of the fosmid revealed a 42,318 bp environmental insert characterized by 46 ORFs. The subcloning of these ORFs demonstrated that a single gene (hqiA) allowed AHL degradation. Enzymatic analysis using puri ed HqiA and HPLC/MS revealed that this protein has lactonase activity on a broad range of AHLs. The introduction of hqiA in the plant pathogen Pectobacterium carotovorum e ciently interfered with both the synthesis of AHLs and quorum-sensing regulated functions, such as swarming motility and the production of maceration enzymes. Bioinformatic analyses highlighted that HqiA showed no sequence homology with the known prototypic AHL lactonases or acylases, thus expanding the AHL-degrading enzymes with a new family related to the cysteine hydrolase (CHase) group. The complete sequence analysis of the fosmid showed that 31 ORFs out of the 46 identi ed were related to Deltaproteobacteria, whilst many intercalated ORFs presented high homology with other taxa. In this sense, hqiA appeared to be assigned to the Hyphomonas genus (Alphaproteobacteria), suggesting that horizontal gene transfer had occurred.

Article: Molecular Ecology Ressources

A new promising phylogenetic marker to study the diversity of fungal communities: the GLYCOSIDE HYDROLASE 63 gene L Pérez‐Izquierdo, E Morin, JP Maurice, F Martin, A Rincón, M Buée. Molecular Ecology Resources

Abstract

In molecular ecology, the development of efficient molecular markers for fungi remains an important research domain. Nuclear ribosomal internal transcribed spacer (ITS) region was proposed as universal DNA barcode marker for Fungi, but this marker was criticized for idel-induced alignment problems and its potential lack of phylogenetic resolution. Our main aim was to develop a new phylogenetic gene and a putative functional marker, from single-copy gene, to describe fungal diversity. Thus, we developed a series of primers to amplify a polymorphic region of the Glycoside Hydrolase GH63 gene, encoding exo-acting α-glucosidases, in Basidiomycetes. These primers were validated on 125 different fungal genomic DNAs and GH63 amplification yield was compared with that of already published functional markers targeting genes coding for laccases, N-acetylhexosaminidases, cellobiohydrolases and class II Peroxidases. Specific amplicons were recovered for 95% of the fungal species tested and GH63 amplification success was strikingly higher than rates obtained with other functional genes. We downloaded the GH63 sequences from 483 fungal genomes publicly available at the JGI MycoCosm database. GH63 was present in 461 fungal genomes belonging to all phyla, except Microsporidia and Neocallimastigomycotadivisions. Moreover, the phylogenetic trees built with both GH63 and Rpb1 protein sequences revealed that GH63 is also a promising phylogenetic marker. Finally, a very high proportion of GH63 proteins was predicted to be secreted. This molecular tool could be a new phylogenetic marker of fungal species as well as potential indicator of functional diversity of Basidiomycotes fungal communities in term of secretory capacities.