A redox meeting will be held the 29-31 March in Amphitheater 8, Faculté des Sciences, Boulevard des Aiguillettes, Vandoeuvre.
Program redox meeting Nancy March 2017 FINAL
A paper entitled “Redox Regulation : Historical Background and Future Perspectives”
http://theconversation.com/biologie-retour-sur-lhistoire-de-la-regulation-redox-74656
Congratulations Leticia for your brilliant PhD defense
A New Method for Qualitative Multi-scale Analysis of Bacterial Biofilms on Filamentous Fungal Colonies Using Confocal and Electron Microscopy. CM Guennoc, C Rose, F Guinnet, I Miquel, J Labbé, A Deveau. JoVE (Journal of Visualized Experiments), e54771-e54771
Summary
Bacterial biofilms frequently form on fungal surfaces and can be involved in numerous bacterial-fungal interaction processes, such as metabolic cooperation, competition, or predation. The study of biofilms is important in many biological fields, including environmental science, food production, and medicine. However, few studies have focused on such bacterial biofilms, partially due to the difficulty of investigating them. Most of the methods for qualitative and quantitative biofilm analyses described in the literature are only suitable for biofilms forming on abiotic surfaces or on homogeneous and thin biotic surfaces, such as a monolayer of epithelial cells.
While laser scanning confocal microscopy (LSCM) is often used to analyze in situ and in vivo biofilms, this technology becomes very challenging when applied to bacterial biofilms on fungal hyphae, due to the thickness and the three dimensions of the hyphal networks. To overcome this shortcoming, we developed a protocol combining microscopy with a method to limit the accumulation of hyphal layers in fungal colonies. Using this method, we were able to investigate the development of bacterial biofilms on fungal hyphae at multiple scales using both LSCM and scanning electron microscopy (SEM). This report describes the protocol, including microorganism cultures, bacterial biofilm formation conditions, biofilm staining, and LSCM and SEM visualizations.
Desirée Gütle défendra publiquement sa thèse mercredi 29 mars à 9h (amphi 7 FST).
“ Characterization of the ferredoxin/thioredoxin system and its targets in Physcomitrella patens ”
Résumé
La régulation redox est un mécanisme ancien présent chez les organismes biologiques et impliquée dans diverses voies métaboliques. En particulier chez les organismes photosynthétiques elle est responsable des mécanismes d’adaptation rapide dans un environnement constamment modifié. Dans les chloroplastes le système ferrédoxine/thiorédoxine est la cascade redox principale qui relie l’activité de plusieurs enzymes plastidiales à la source lumineuse. Le rôle central dans ce système est joué par la ferrédoxine-thiorédoxine réductase (FTR), une protéine hétérodimérique qui récupère des électrons à partir de la ferrédoxine photoréduite et les transfère pour réduire des thiorédoxines plastidiales. Ces protéines peuvent alors réduire des enzymes cibles, requérant l’accessibilité de paires de cystéines dans un disulfure dont la réduction résulte en une activation/ inactivation de la cible. Jusqu’à présent des plantes viables n’ont pu être obtenues en l’absence de ce système de régulation. Dans cette thèse des secteurs du système redox ont été explorés chez la plante modèle Physcomitrella patens (une mousse). Par manipulation de gènes l’influence de l’enzyme FTR sur la croissance et le développement de la plante a été analysée suivant différents paramètres. De manière à impacter la fonction de la réductase des changements nucléotidiques simples ont été introduits au niveau des codons programmant les cystéines catalytiques et dans un deuxième temps le gène complet a été supprimé. De façon inattendue nous n’avons observé aucun effet significatif sur la viabilité et le développement des plantes mutantes. De plus, nous avons détecté dans P. patens des thiorédoxines additionnelles absentes chez les plantes à graine qui sont fonctionnelles vis à vis des enzymes cibles mais non-réduites par la FTR. Ceci rend possible un scénario de compensation chez les mutants via un système de réduction FTR-indépendant qui reste à caractériser. Deux des cibles photorégulées, la fructose-1,6-bisphosphatase (FBPase) et la sédoheptulose-1,7-bisphosphatase (SBPase), fonctionnent dans la phase de régénération du cycle de Calvin-Benson cycle et elles possèdent plusieurs caractéristiques de catalyse et de régulation similaires. En combinant des approches biochimiques et structurales, une comparaison fonctionnelle et structurale des deux phosphatases de P. patens a été conduite. De plus l’analyse phylogénétique a révélé une origine procaryotique indépendante des deux séquences en dépit de leurs similitudes structurales et catalytiques.
De plus trois articles de revue résument la plasticité et la représentativité du modèle P. patens pour la recherche forestière, les principes généraux de la régulation redox relativement aux aspects évolutifs et fonctionnels chez les plantes ainsi que l’ état de l’art de lé régulation redox chez les espèces ligneuses en utilisant principalement le peuplier comme modèle.
Mot clès: Régulation redox, FTR système, FBPase, SBPase, Physcomitrella patens
Abstract
Redox regulation is an ancient mechanism present in biological organisms and is involved in diverse cellular pathways. In particular in photosynthetic organisms it is responsible for fast adaption mechanisms to a constantly changing environment. In chloroplasts the ferredoxin/thioredoxin system represents the main redox regulatory cascade which links the activity of several plastid enzymes to the energy source, light. A central role in this system is played by the heterodimeric ferredoxin-thioredoxin reductase (FTR), which gains electrons from the photo-reduced ferredoxin and transfers those further on via reduction to plastidal thioredoxins. Those proteins in turn reduce their target enzymes and require therefore the availability of redox sensitive cysteine pairs whose reduction results in an inactivation/activation switch of the targets.
So far no viable plants could be obtained in complete absence of this redox regulation system. In this thesis single sections of the system were explored in the model plant Physcomitrella patens. Through gene manipulation the influence of the FTR enzyme on plant growth and development was analysed. In order to impact on the function of the reductase, firstly single nucleotide exchange of the catalytic cysteines was performed and later on the gene was completely deleted. Surprisingly, no significant effect could be observed on the viability and development of mutant lines compared to WT plants. Furthermore we found that P. patens possesses in contrast to seed plants additional thioredoxins which are functional for reduction of FTR target enzymes but are most likely not supplied with electrons by this reductase. Thus a possible rescue scenario independent of FTR could be assumed for P. patens and also by other redox regulation systems present in chloroplasts.
Two of the FTR target enzymes, fructose-1,6-bisphosphatase and sedoheptulose-1,7-bisphosphatase, are functional in the regeneration phase of the Calvin-Benson cycle and share similar characteristics in regulation and catalysis. By combining biochemical and structural approaches a functional comparison of both phosphatases was conducted using cDNAs from P. patens. A more strict TRX-dependent regulation and catalytic cleavage ability for both substrates, FBP and SBP, could be observed for SBPase, whereas FBPase is only capable of cleaving FBP. By obtaining the oxidized x-ray structure of both enzymes these observations can be associated with the distinct positions of regulatory sites and the various sizes of the substrate binding pocket. In addition, the phylogenetic analysis revealed an independent prokaryotic origin for both phosphatases.
Furthermore we summarized in three review articles the amenability of P. patens as model plant for forest research, the general principles of redox regulation in respect of evolution and functional mechanisms in plants, and the current state of the art in forest redox regulation using poplar as exemplary model.
Research engineer in bioinformatics and data analysis from microbial metagenomics research using microbial genomic resources
(INRA permanent position open in 2017 – application before 16th March)
IR BioInfo position_INRA-IAM_Nancy (Eng)
JOB TITLE
Research engineer in bioinformatics and data analysis
Environment
The position is open in the “Trees / Microorganisms Interactions” research Unit (INRA / Lorraine University) (http://mycor.nancy.inra.fr/IAM/?page_id=2000).
The recruited person will work in collaboration with scientists of the unit and with a permanent bioinformatics engineer from the technical platform in bioinformatics. This person will develop an expertise in microbial metagenomics, and particularly in fungal metagenomics / metatranscriptomics, and he / she will be involved in national (National Research Agency, Meta-omics and Microbial Ecosystems (MEM) meta-program) and international research programs (US Department of Energy Joint Genome Institute; e.g. Metatranscriptomics of Forest Soil Ecosystems).
CONTEXT AND ACTIVITIES
The huge microbial richness (bacteria and fungi) in forest soils remains little explored in spite of fundamental or applied research opportunities (understanding of the functioning of ecosystems, analysis of functional diversity, extended phenotype of host plants, forestry management, ecosystem services, bioenergy). The study of this microbial diversity and their trophic and functional interactions requires integrative approaches (“meta-omics”) and powerful analysis and data mining. The use and development of bioinformatics tools for New Generation Sequencing (NGS) data analysis from microbial (prokaryotic and eukaryotic) metagenomic and metatranscriptomic researches will require:
(i) to develop and use appropriate bioinformatics procedures
(ii) to store, search, and compare meta-omic data sets
(iii) to identify the gene networks and associated key functions (indicators) controlling these complex systems.
Finally, in interaction with colleagues of the technical platform, the recruited person will also lead and deploy the hardware and software architecture and its application (installation, assistance, training, evaluation).
SKILLS AND Personal Qualities
– PhD in biology, bioinformatics, computer science or Engineer degree from graduate schools or equivalent.
– Professional experience in biology and/or in genomics / metagenomics, ideally applied to microbiology, molecular ecology or plant / microbe physiology. Good knowledge of programming languages and tools for building / managing databases adapted to the analysis of recommended metagenomes.
– One or more proven experiments in (meta) comparative genomics of gene repertoires; Computer processing of sequencing data (HiSeq, MiSeq, PacBio …); Quantification of fungal gene expression (trimming, mapping, normalization); Differential expression and analysis of co-expression networks.
Cora Miquel-Guennoc défendra publiquement sa thèse Lundi 6 mars à 14h30 (amphi 7 FST).
“Etude de l’interaction physique entre le champignon ectomycorhizien Laccaria bicolor S238N et la bactérie auxiliaire de la mycorhization Pseudomonas fluorescens BBc6R8”
Résumé
Dans les sols, les champignons ectomycorhiziens (ECM) forment une symbiose très répandue avec les racines des arbres et contribuent ainsi à leur croissance et à leur santé. Des études antérieures ont montré que certaines bactéries pouvaient influencer positivement la symbiose entre les ECM et les arbres, appelées BAM pour Bactéries Auxiliaires de la Mycorhization. Les mécanismes de l’effet auxiliaire des BAM sont encore peu connus. En amont de cette thèse, il avait été montré in vitro que la BAM Pseudomonas fluorescens BBc6 formait des structures similaires à des biofilms sur les hyphes de l’ECM Laccaria bicolor. Dans ce contexte, afin d’enrichir les connaissances concernant les interactions entre les ECM et les BAM, cette thèse a porté sur l’interaction physique entre ces deux organismes. L’étude a en partie été réalisée via une méthode d’analyse par microscopie confocale, développée durant cette thèse. Les résultats obtenus ont montré que cette bactérie formait des biofilms localisés préférentiellement sur la région apicale des colonies de l’ECM ce qui pourrait indiquer une interaction trophique. L’existence d’une telle interaction a d’ailleurs par la suite été confirmée. Les résultats ont également montré que l’interaction physique entre L. bicolor et BBc6 n’est pas spécifique puisque l’ensemble des treize autres souches bactériennes testées a formé des biofilms sur les hyphes de L. bicolor. En revanche, BBc6 s’est montrée incapable de former des biofilms sur certains champignons appartenant aux Ascomycètes, suggérant des mécanismes d’inhibition. De plus, l’étude de la matrice des biofilms formés par BBc6R8 a révélé la présence de réseaux de filaments constitués d’ADN qui semblent structurer ces biofilms et qui ont aussi été observés chez l’ensemble des souches bactériennes testées. Ces résultats révèlent un rôle structural de la molécule d’ADN qui, bien qu’il semble répandu, n’a que peu été reporté jusqu’à présent. Enfin, il a été montré que des mutants de BBc6 qui ont perdu leur effet auxiliaire forment des biofilms différents de la souche sauvage sur une surface abiotique suggérant un lien potentiel entre l’effet auxiliaire et la formation de biofilms.
Abstract
In soil ecosystems, ectomycorrhizal fungi (ECM) form a widespread symbiosis with roots of trees, contributing to tree growth and health. It has been shown that some bacteria, called mycorrhization helper bacteria (MHB), stimulate mycorrhizal symbiosis. The mechanisms of this helper effect are poorly understood. Previous studies have shown that the MHB Pseudomonas fluorescens BBc6 formed biofilm-like structures around the hyphae of the ECM Laccaria bicolor during their in vitro interaction. In this context, in order to increase knowledge concerning MHB / ECM interactions, the work presented here focuses on the physical interaction between these two organisms. To this purpose, a method of analysis based on confocal microscopy was developed. The results showed that the bacteria formed biofilms preferentially localized on the apical region of the ECM colonies, which could indicate a trophic interaction. Such an interaction has been subsequently confirmed. The results also showed that the physical interaction between L. bicolor and BBc6 is not specific since all thirteen other bacterial strains tested formed biofilms on the hyphae of L. bicolor. On the other hand, BBc6 was unable to form biofilms on some fungi belonging to Ascomycetes, suggesting the existence of inhibition mechanisms. Moreover, the study of the BBc6 biofilm matrix revealed networks of DNA-containing filaments which seem to structure these biofilms and which have also been observed in all the bacterial strains tested. These results reveal a structural role of the DNA molecule, a role that has been rarely reported so far despite its probable high occurence. Finally, it has been shown that BBc6R8 mutants having lost their helper effect presented a modified phenotype concerning their biofilm formation on abiotic surface, suggesting a potential link between the helper effect and the biofilms formation.
The defense was held the 2th March 2017 at 13H30 in Amphitheater 7, Faculté des Sciences, Boulevard des Aiguillettes, Vandoeuvre.
Congratulations Claire for your brilliant defense!
Post-translational modifications of Medicago truncatula glutathione peroxidase 1 induced by nitric oxide C Castella, I Mirtziou, A Seassau, A Boscari, F Montrichard, … 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 Gpx1 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.
Functional outcomes of fungal community shifts driven by tree genotype and spatial‐temporal factors in Mediterranean pine forests L Pérez‐Izquierdo, M Zabal‐Aguirre, D Flores‐Rentería, … Environmental Microbiology
Fungi provide relevant ecosystem services contributing to primary productivity and the cycling of nutrients in forests. These fungal inputs can be decisive for the resilience of Mediterranean forests under global change scenarios, making necessary an in-deep knowledge about how fungal communities operate in these ecosystems. By using high-throughput sequencing and enzymatic approaches, we studied the fungal communities associated with three genotypic variants of Pinus pinaster trees, in 45-yr-old common garden plantations. We aimed to determine the impact of biotic (i.e. tree genotype) and abiotic (i.e. season, site) factors on the fungal community structure, and to explore whether structural shifts triggered functional responses affecting relevant ecosystem processes. Tree genotype and spatial-temporal factors were pivotal structuring fungal communities, mainly by influencing their assemblage and selecting certain fungi. Diversity variations of total fungal community and of that of specific fungal guilds, together with edaphic properties and tree’s productivity, explained relevant ecosystem services such as processes involved in carbon turnover and phosphorous mobilization. A mechanistic model integrating relations of these variables and ecosystem functional outcomes is provided. Our results highlight the importance of structural shifts in fungal communities because they may have functional consequences for key ecosystem processes in Mediterranean forests. This article is protected by copyright. All rights reserved.
