Differential accumulation of beta-1,3-glucanase and chitinase isoforms in tomato roots in response to colonization by either pathogenic or non-pathogenic strains of Fusarium oxysporum

International audience

Changes in polypeptide profiles of two pea genotypes inoculated with the arbuscular mycorrhizal fungus Glomus mosseae

The symbiotic interaction between pea roots ('Pisum sativum' L.) and the endomycorrhizal fungus 'Glomus mosseae' should lead to specific gene expression of both symbionts. In order to detect symbiosis-related proteins (endomycorrhizins), we used two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) after phenolic extraction of total soluble proteins. Furthermore, to better characterize the molecular changes after fungal infection, two pea genotypes available in our laboratory were used: cv. Frisson (myc⁺) and an isogenic mycorrhiza-resistant mutant (myc⁻). The latter prevents intraradical fungal development. Several differences were observed in polypeptide patterns of silver-stained …

Protein profiling analyses in arbuscular mycorrhizal symbiosis

Proteomics, a powerful approach towards understanding functional plant root interactions with arbuscular mycorrhizal fungi

International audience

Influence of arbuscular mycorrhizal colonisation on cadmium induced Medicago truncatula root isoflavonoid accumulation.

Cadmium is a serious environmental pollution threats to the planet. Its accumulation in plants affects many cellular functions, resulting in growth and development inhibition, whose mechanisms are not fully understood. However, some fungi forming arbuscular mycorrhizal symbiosis with the majority of plant species have the capacity to buffer the deleterious effect of this heavy metal. In the present work we investigated the capacity of Rhizophagus irregularis (syn. Glomus irregularis) to alleviate cadmium stress in Medicago truncatula. In spite of a reduction in all mycorrhizal parameters, plants colonized for 21 days by R. irregularis and treated by 2 mg kg(-1) cadmium displayed less growth…

Proteome analysis and identification of symbiosis-related proteins from Medicago truncatula Gaertn. by two-dimensional electrophoresis and mass spectrometry.

Time-course analysis of root protein profiles was studied by two-dimensional gel electrophoresis and silver staining in the model plant Medicago truncatula, inoculated either with the arbuscular mycorrhizal fungus Glomus mosseae or with the nitrogen fixing bacterium Sinorhizobium meliloti. Protein modifications in relation to the development of both symbioses included down- and upregulations, as well as newly induced polypeptides. Matrix assisted laser desorption/ionization-time of flight-mass spectrometry after trypsin digestion clearly identified one polypeptide induced in nodulated roots as a M. truncatula leghemoglobin. Internal sequencing with a quadrupole time-of-flight mass spectrome…

Effets protecteurs de la symbiose mycorhizienne à arbuscule vis - à - vis du cadmium: identification des protéines potentiellement impliquées et quantification de deux transcrits

The contribution of model legumes to arbuscular mycorrhiza research

International audience

Plant defense responses induced by arbuscular mycorrhizal fungi

Plants in their environment daily face many organisms such as fungi, bacteria, mycoplasms, viruses, nematodes, etc. Many of them are potential pathogens; in fact thousands of microorganisms are known to cause plant diseases. Despite this large number of deleterious microorganisms, most of the plants are resistant to their attack since they have developed effective mechanisms to protect themselves.

Could subcellular proteomics of root plastids teach us more about mycorrhizal symbiosis?

International audience; The arbuscular mycorrhizal (AM) symbiosis is a mutualistic association between soil-borne fungi and the roots of most plant species. Involving the bilateral exchange of nutrients, the symbiosis is connected to drastic changes in plant cell organelle morphology and physiology. Root plastids, in particular, are forming extensive, network-like structures covering the main symbiotic interface, i.e., intracellular, highly branched haustorium-like fungal structures called arbuscules. These plastid networks are highly dynamic and are formed and degraded concomitantly with the formation and degradation of arbuscules. By producing basic metabolic building blocks like fatty ac…

Plant proteome analysis

Proteome analysis is becoming a powerful tool in the functional characterization of plants. Due to the availability of vast nucleotide sequence information and based on the progress achieved in sensitive and rapid protein identification by mass spectrometry, proteome approaches open up new perspectives to analyze the complex functions of model plants and crop species at different levels. In this review, an overview is given on proteome studies performed to analyze whole plants or specific tissues with particular emphasis on important physiological processes such as germination. The chapter on subcellular proteome analysis of plants focuses on the progress achieved for plastids and mitochond…

Studying the arbuscular mycorrhizal symbiosis from axenic system to soilborne interactions by proteomics: which challenges to face?

Membrane proteomic of arbuscular mycorrhiza

Blue-copper binding proteins of Medicago truncatula: Characterization of the expression of two genes related to the arbuscular mycorrhizal symbiosis

International audience; Expression profiling of two paralogous arbuscular mycorrhizal (AM)-specific blue copper-binding gene (MtBcpla and MtBcp1b) isoforms was performed by real-time quantitative polymerase chain reaction in wild-type Medicago truncatula Jemalong 5 (J5) during the mycorrhizal development with Glomus intraradices for up to 7 weeks. Timecourse analysis in J5 showed that expression of both MtBcp1 genes increased continuously and correlated strongly with the colonization intensity and arbuscule content. MtPT4, selected as a reference gene of the functional plant-fungus association, showed a weaker correlation to mycorrhizal development. In a second experiment, a range of mycorr…

Effects of gibberellic acid (GA3) treatments on endomycorrhizas

International audience

La protéomique, un outil pour évaluer l'impact de boues de stations d'épuration sur les interactions symbiotiques entre Medicago truncatula et le champignon mycorhizogène à arbuscule Glomus mosseae ou la bactérie fixatrice d'azote Sinorhizobium meliloti

National audience

Fungal proteins in the extra-radical phase of arbuscular mycorrhiza: a shotgun proteomic picture

International audience

Membrane proteomic analyses to reveal and identify arbuscular mycorrhizal-related proteins

Proteomics: a functional genomic approach for studying mycorrhizal interactions in legumes

Comparaison des interactions plante - Fusarium oxysporum pathogènes et plante F. oxysporum non pathogènes par analyse protéique différentielle

National audience

A technical trick for studying proteomics in parallel to transcriptomics in symbiotic root-fungus interactions

We have developed a protocol in which proteins and mRNA can be analyzed from single root samples. This experimental design was validated in arbuscular mycorrhiza by comparing the proteins profiles obtained with those from a classical protein extraction process. It is a step forward to make simultaneous proteome and transcriptiome profiling possible.

Mutations in DMI3 and SUNN modify the appressorium-responsive root proteome in arbuscular mycorrhiza.

Modification of the Medicago truncatula root proteome during the early stage of arbuscular mycorrhizal symbiosis was investigated by comparing, using two-dimensional electrophoresis, the protein patterns obtained from non-inoculated roots and roots synchronized for Glomus intraradices appressorium formation. This approach was conducted in wild-type (J5), mycorrhiza-defective (TRV25, dmi3), and autoregulation-defective (TR122, sunn) M. truncatula genotypes. The groups of proteins that responded to appressorium formation were further compared between wild-type and mutant genotypes; few overlaps and major differences were recorded, demonstrating that mutations in DMI3 and SUNN modified the ap…

Effect of cadmium on growth, isoflavonoid and protein accumulation patterns in mycorrhizal roots of Medicago truncatula

International audience; Ecosystems are submitted to various abiotic stresses, among which heavy metals represent major industrial pollutants. Cadmium (Cd), that has damaging effects on plant metabolism, occurs in agricultural environments through industrial pollution and human activities, including phosphate fertiliser and sewage sludge applications. Metal availability to plants can be modulated by soil microorganisms, including arbuscular mycorrhizal (AM) fungi. In the present work, Cd effects on the model legume Medicago truncatula inoculated or not with the AM fungus Glomus intraradices have been studied at 3 levels: (1) plant biomass production together with green part chlorophyll quant…

Analyse protéique de différentes souches de Fusarium oxysporum

Erratum to “Hydrolytic enzyme activity of Paenibacillus sp. strain B2 and effects of the antagonistic bacterium on cell integrity of two soil-borne pathogenic fungi”

Ce texte contient des corrections apportées à l'article " Hydrolytic enzyme activity of Paenibacillus sp. strain B2 and effects of the antagonistic bacterium on cell integrity of two soil-borne pathogenic fungi" sorti dans le numéro 15 du journal "Applied Soil Ecology" sorti en 2000.

Germination and storage protein mobilisation of Trifolium subterraneum Var 45C seeds under hypoxic treatment

International audience; The rate, speed and latency period of germination were defined for seeds of Trifolium subterraneum variety 45 C. The latency period was 12 hours on distilled water. The hypoxic stress was applied at the beginning (T12) and the end of latency period (T24). It consists on the addition of nitrogen (N2) under 1 bar pressure in hermetically closed boxes during 2 hours. The germination of treated seeds was then assessed on distilled water. The germination test during hypoxic treatment showed that the hypoxic stress delayed germination, slowing down its speed and decreasing the final percentage of germinated seeds. In order to better understand this depressive effect observ…

The membrane proteome of Medicago truncatula roots displays qualitative and quantitative changes in response to arbuscular mycorrhizal symbiosis

International audience; Arbuscular mycorrhizal (AM) symbiosis that associates roots of most land plants with soil-borne fungi (Glomeromycota), is characterized by reciprocal nutritional benefits. Fungal colonization of plant roots induces massive changes in cortical cells where the fungus differentiates an arbuscule, which drives proliferation of the plasma membrane. Despite the recognized importance of membrane proteins in sustaining AM symbiosis, the root microsomal proteome elicited upon mycorrhiza still remains to be explored. In this study, we first examined the qualitative composition of the root membrane proteome of Medicago truncatula after microsome enrichment and subsequent in dep…

Impact of sewage sludges on Medicago truncatula symbiotic proteome

The effects of sewage sludges were investigated on the symbiotic interactions between the model plant Medicago truncatula and the arbuscular mycorrhizal fungus Glomus mosseae or the rhizobial bacteria Sinorhizobium meliloti. By comparison to a control sludge showing positive effects on plant growth and root symbioses, sludges enriched with polycylic aromatic hydrocarbons or heavy metals were deleterious. Symbiosis-related proteins were detected and identified by two-dimensional electrophoresis and matrix-assisted laser desorption ionization mass spectrometry, and image analysis was used to study the effects of sewage sludges on M. truncatula symbiotic proteome.

Approche protéomique pour l'identification des protéines du champignon mycorhizogène à arbuscules Glomus intraradices N.C. Schenk et G.S. Sm (DAOM 181602) produit in vitro sur racines de carottes transformées Ri T-DNA (Daucus carota L.)

National audience

Sub-protéomique de la symbiose mycorhizienne de Medicago truncatula avec le champignon mycorhizogène à arbuscules Glomus mosseae

National audience

Symbiosis-related gene expression during early interactions between AM fungi and roots

International audience

Specificity and compatibility in arbuscular mycorrhizal symbiosis

International audience

Protéome membranaire en réponse à la symbiose mycorhizienne à arbuscules par GeLC-MS/MS

Proteomics as a tool to monitor plant-microbe endosymbioses in the rhizosphere

In recent years, outstanding molecular approaches have been used to investigate genes and functions involved in plant-microbe endosymbioses. In this review, we outline the use of proteomic analysis, based on two-dimensional electrophoresis and mass spectrometry, to characterize symbiosis-related proteins. During the last decade, proteomics succeeded in identifying about 400 proteins associated with the development and functioning of both mycorrhizal and rhizobial symbioses. Further progress in prefractionation procedures is expected to allow the detection of symbiotic proteins showing low abundance or being present in certain cell compartments.

Membrane proteomics of arbuscular mycorrhiza

International audience

A proteomic approach to studying plant response to crenate broomrape (Orobanche crenata) in pea (Pisum sativum)

Abstract Crenate broomrape ( Orobanche crenata ) is a parasitic plant that threatens legume production in Mediterranean areas. Pea ( Pisum sativum ) is severely affected, and only moderate levels of genetic resistance have so far been identified. In the present work we selected the most resistant accession available (Ps 624) and compared it with a susceptible (Messire) cultivar. Experiments were performed by using pot and Petri dish bioassays, showing little differences in the percentage of broomrape seed germination induced by both genotypes, but a significant hamper in the number of successfully installed tubercles and their developmental stage in the Ps 624 compared to Messire. The prote…

Proteomic approaches to learn more on legume-micobe interactions

Bioprotection du pois par le champignon mycorhizogène à arbuscules Glomus mosseae vis-à-vis du champignon pathogène Aphanomyces euteiches

National audience

Mycorrhiza-related chitinase and chitosanase activity isoforms in Medicago truncatula Gaertn

International audience

La protéomique comme moyen d'identification de protéines induites par le cadmium et différenciellement régulées dans les racines de pois colonisées par un champignon mycorhizogène à arbuscules

National audience

Localized versus systemic effect of arbuscular mycorrhizal fungi on defence responses to Phytophthora infection in tomato plants

Development of biological control for plant diseases is accepted as a durable and environmentally friendly alternative for agrochemicals. Arbuscular mycorrhizal fungi (AMF), which form symbiotic associations with root systems of most agricultural, horticultural and hardwood crop species, have been suggested as widespread potential bioprotective agents. In the present study the ability of two AMF (Glomus mosseae and Glomus intraradices) to induce local or systemic resistance to Phytophthora parasitica in tomato roots have been compared using a split root experimental system. Glomus mosseae was effective in reducing disease symptoms produced by P. parasitica infection, and evidence points to …

The arbuscular mycorrhizal symbiosis a modulator of cadmium stress

International audience; Ecosystems are submitted to various abiotic stresses, among which heavy metals represent major industrial pollutants. Cadmium (Cd), that has damaging effects on plant metabolism, occurs in agricultural environments through industrial pollution and human activities, including phosphate fertiliser and sewage sludge applications. Metal availability to plants can be modulated by soil microorganisms, such as arbuscular mycorrhizal (AM) fungi. In the present work, Cd effects on the model legume Medicago truncatula inoculated or not with the AM fungus Glomus intraradices have been studied at 3 levels: (1) plant biomass production together with green part chlorophyll quantif…

Getting to the symbiotic proteome of Medicago truncatula

Purification and partial amino acid sequencing of a mycorrhiza-related chitinase isoform from Glomus mosseae-inoculated roots of Pisum sativum L

International audience

A proteomic approach to study the effects of two polycyclic aromatic hydrocarbons on arbuscular mycorrhiza of Cichorium intybus roots in gnotobiotic conditions

Proteomics for the functional analysis of microorganisms

International audience; Dans l’ère post-génomique, la protéomique est une approche en pleine expansion qui bénéficie des avancées de méthodologies de pointe pour la séparation et l’identification des produits des gènes et de leurs fonctions. Dans ce contexte, elle apparaît un outil de choix pour étudier les réponses des micro-organismes aux perturbations de leur environnement.

Cadmium stress alleviation by arbuscular mycorrhizal symbiosis: proteomic studies of the mechanisms involved in shoots of M. truncatula

Proteomes of arbuscular mycorrhizal symbiosis : past and present achievements

Exploring the Genome of glomeromycotan fungi

Chapitre 1; International audience; All fungi forming a mutualistic symbiosis with plant roots called arbuscular mycorrhiza were formerly grouped together in one order, the Glomales, placed in the Zygomycota (Morton 1993). Based on molecular analyses suggestingthat arbuscular mycorrhizal fungi should be separated from other fungal taxa, they were transferred a decade ago to the Glomeromy-cota , a new phylum created specifically for them (Schu¨ ssler et al. 2001). Whilst members of this monophyletic group originated from the same common ancestor as the Ascomycota and Basidiomycota, they have no obvious affinity to other major extant phylogenetic groups in the kingdom Fungi (James et al. 2006) …

Plant mycorrhizal interaction: What can we learn from a proteomic approach?

Transcription of two blue copper-binding protein isogenes is highly correlated with arbuscular mycorrhizal development in Medicago truncatula.

International audience; Expression profiling of two paralogous arbuscular mycorrhizal (AM)-specific blue copper-binding gene (MtBcp1a and MtBcp1b) isoforms was performed by real-time quantitative polymerase chain reaction in wild-type Medicago truncatula Jemalong 5 (J5) during the mycorrhizal development with Glomus intraradices for up to 7 weeks. Time-course analysis in J5 showed that expression of both MtBcp1 genes increased continuously and correlated strongly with the colonization intensity and arbuscule content. MtPT4, selected as a reference gene of the functional plant-fungus association, showed a weaker correlation to mycorrhizal development. In a second experiment, a range of mycor…

Differential expression proteomics to investigate responses and resistance to Orobanche crenata in legumes

International audience; In order to study the plant response to parasitic plants and the molecular basis of the resistance we have used a proteomic approach. The root proteome of two accessions of the model legume Medicago truncatula and crop pea displaying differences in their resistance phenotype, in control as well as in infected plants have been compared. We report quantitative as well as qualitative differences in the two-dimensional gel electrophoresis (2-DE) maps between plant genotypes and treatments (control, non-infected, and infected). Differential spots spots were successfully identified by peptide mass fingerprinting (PMF) following MALDI-TOF mass spectrometry. Many of the prot…

Participation au « Model Legumes Congress 2007 » (MLC2007)- Compte-rendu de mission

La microdissection laser: une technique d’analyse du profil d’expression génique et protéique des types cellulaires spécifiques de la symbiose mycorhizienne à arbuscules

Wanted : pathogenesis-related marker molecules for Fusarium oxysporum

Summary Although Fusarium oxysporum pathogens cause severe wilts in about 80 botanical species, the mechanisms of pathogenicity and symptom induction are poorly understood. Knowledge about the genetic and biochemical pathways involved in the pathogenesis of F. oxysporum would be invaluable in getting targets for both fungicide development and search for biocontrol agents. In this respect, we described the main approaches that have been developed to identify some mechanisms underlying the pathogenesis of F. oxysporum . During the last decades, the potential functions triggering of F. oysporum pathogenicity have mainly been investigated by comparing soilborne pathogenic strains with nonpathog…

Protein actors sustaining arbuscular mycorrhizal symbiosis: underground artists break the silence

'Summary' 26 I. 'Casting for a scenario' 26 II. 'Nominees for a preliminary role' 27 III. 'Nominees for a leading role' 32 IV. 'Future artists' 37   'Acknowledgements' 38   References 38 Summary The roots of most land plants can enter a relationship with soil-borne fungi belonging to the phylum Glomeromycota. This symbiosis with arbuscular mycorrhizal (AM) fungi belongs to the so-called biotrophic interactions, involving the intracellular accommodation of a microorganism by a living plant cell without causing the death of the host. Although profiling technologies have generated an increasing depository of plant and fungal proteins eligible for sustaining AM accommodation and functioning, a …

Un repertoire GeLC - MS/MS des proteins extra - radiculaires de Glomus intraradices

Arbuscular mycorhizal proteomes: what news at the nearby and distant horizon?

International audience; Proteomics has soon emerged as a powerful tool to point out protein modifications in roots interacting with arbuscular mycorrhiza (AM) fungi. Depending on the developmental mycorrhizal stage and on the available amount of mycorrhizal material, untargeted and/or sub-cellular proteomic approaches were applied to reveal and identify proteins whose accumulation was modified during the AM colonisation of Medicago truncatula roots. For the early stage of symbiosis, the protein patterns obtained from noninoculated roots and roots synchronized for appressorium formation in wild-type (Jemalong J5), penetration-defective (TRV25, dmi3) and autoregulation-defective (TR122, sunn)…

Fungal protein accumulation in the extra - radical phase of arbuscular mycorrhiza: a shotgun proteomic picture

Hydrolytic enzyme activity of Paenibacillus sp. strain B2 and effects of the antagonistic bacterium on cell integrity of two soil-borne pathogenic fungi

Paenibacillus sp. strain B2, isolated from the mycorrhizosphere of Sorghum bicolor and having an antagonistic activity towards soil-borne fungal pathogens, possessed extracellular cellulolytic, proteolytic, chitinolytic and pectinolytic enzyme activities. The eventual role of these lytic enzymes in cellular interactions between Paenibacillus sp. strain B2 and Phytophthora parasitica and Fusariumoxysporum was investigated by electron microscopy and molecular cytology. Electron microscopic observations showed that the presence of Paenibacillus sp. strain B2 resulted in disorganisation of cell walls and/or cell contents of P. parasitica and F. oxysporum. However, when P. parasitica was treated…

Un répertoire GeLC - MS/MS des protéines extra - radiculaires de Glomus intraradices

Contribution of proteomics to arbuscular mycorrhiza in Medicago truncatula

International audience; Because proteins are key effectors of plant responses to environmental cues including recognition, signalling, transport and defence reactions, main interest has been paid to characterize those involved in the establishment and functioning of arbuscular mycorrhizal (AM) symbiosis. In our group, the setting up of high throughput proteomic techniques on the model species, Medicago truncatula, is providing step-by-step a large-scale analysis of AM symbiosis-related proteins. Depending on the symbiotic stage targeted and on the abundance of mycorrhizal material, different proteomic strategies that can be combined with other large-scale approaches (transcriptomic and meta…

Proteomics as a way to identify extra-radicular fungal proteins from Glomus intraradices - RiT-DNA carrot root mycorrhizas

To identify fungal proteins involved in the arbuscular mycorrhizal symbiosis, root-inducing transferred-DNA transformed roots of carrot (Daucus carota L.) were in vitro inoculated with Glomus intraradices. Proteins extracted from the extra-radical fungus were analysed by two-dimensional gel electrophoresis. A fungal reference map displaying 438 spots was set up. Four proteins, among the 14 selected for tandem mass spectrometry analysis, were identified including a NmrA-like protein, an oxido-reductase, a heat-shock protein and an ATP synthase beta mitochondrial precursor. The possible fungal origin of a MYK15-like protein found in mycorrhizal roots was further discussed. This is the first r…

Vers l'étude quantitative et fonctionnelle des protéomes membranaires des racines mis en jeu au cours de la symbiose mycorhizienne à arbuscules de Medicago truncatula

Targeted proteomics to identify cadmium-induced protein modifications in Glomus mosseae-inoculated pea roots

International audience

Label-free 1-DE-LC-MS/MS to identify arbuscular mycorrhiza-related membrane proteins

Deep changes in the shape and number of organelles, together with profound modifications in various membrane compartments, are induced within arbuscular mycorrhizal (AM) symbiosis. In this context, to investigate the membrane-associated proteins that are regulated in the model interaction Medicago truncatula – Rhizophagus irregularis, label-free 1DE-LC-MS/MS approach has been employed as alternative to two-dimensional gel electrophoresis. The existence of a correlation between protein abundance and peak areas or number of MS/MS spectra has widened the choice of label-free quantitative proteomics. The results highlighted microsomal protein candidates that could be involved in the symbiotic e…