Gilles Comte

Exogenous glucosinolate produced by Arabidopsis thaliana has an impact on microbes in the rhizosphere and plant roots.

A specificity of Brassicaceous plants is the production of sulphur secondary metabolites called glucosinolates that can be hydrolysed into glucose and biocidal products. Among them, isothiocyanates are toxic to a wide range of microorganisms and particularly soil-borne pathogens. The aim of this study was to investigate the role of glucosinolates and their breakdown products as a factor of selection on rhizosphere microbial community associated with living Brassicaceae. We used a DNA-stable isotope probing approach to focus on the active microbial populations involved in root exudates degradation in rhizosphere. A transgenic Arabidopsis thaliana line producing an exogenous glucosinolate and the associated wild-type plant associated were grown under an enriched 13CO2 atmosphere in natural soil. DNA from the rhizospheric soil was separated by density gradient centrifugation. Bacterial (Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria and Acidobacteria), Archaea and fungal community structures were analysed by DGGE fingerprints of amplified 16S and 18S rRNA gene sequences. Specific populations were characterized by sequencing DGGE fragments. Roots of the transgenic plant line presented an altered profile of glucosinolates and other minor additional modifications. These modifications significantly influenced microbial community on roots and active populations in the rhizosphere. Alphaproteobacteria, particularly Rhizobiaceae, and fungal communities were mainly impacted by these Brassicaceous metabolites, in both structure and composition. Our results showed that even a minor modification in plant root could have important repercussions for soil microbial communities.

The flavanolignan silybin and its hemisynthetic derivatives, a novel series of potential modulators of P-glycoprotein.

A new series of potential flavonoidic modulators of P-glycoprotein activity has been prepared. The flavanolignan silybin was first oxidised to dehydrosilybin and then C-alkylated with either prenyl or geranyl bromide. The resulting isoprenoid dehydrosilybins were shown to display high in vitro affinities for direct binding to P-glycoprotein, which ranged them among the best flavonoids ever tested.

ORGANOLITHIUM MEDIATED SYNTHESIS OF PRENYLCHALCONES AS POTENTIAL INHIBITORS OF CHEMORESISTANCE

A number of substituted chalcones have been prepared by a novel LiHMDS-mediated aldol condensation, the first method consistent with the use of alkali-labile protecting groups such as tert-butyldiphenylsilyl or tert-butyldimethylsilyl. Chalcone substitution by prenylation increases their binding affinity to P-glycoprotein responsible for cancer cells chemoresistance.

Plant secondary metabolite profiling evidences strain-dependent effect in the Azospirillum-Oryza sativa association

Azospirillum is a plant growth-promoting rhizobacterium (PGPR) able to enhance growth and yield of cereals such as rice, maize and wheat. The growth-promoting ability of some Azospirillum strains appears to be highly specific to certain plant species and cultivars. In order to ascertain the specificity of the associative symbiosis between rice and Azospirillum, the physiological response of two rice cultivars, Nipponbare and Cigalon, inoculated with two rice-associated Azospirillum was analyzed at two levels: plant growth response and plant secondary metabolic response. Each strain of Azospirillum (Azospirillum lipoferum 4B isolated from Cigalon and Azospirillum sp. B510 isolated from Nipponbare) preferentially increased growth of the cultivar from which it was isolated. This specific effect is not related to a defect in colonization of host cultivar as each strain colonizes effectively both rice cultivars, either at the rhizoplane (for 4B and B510) and inside the roots (for B510). The metabolic profiling approach showed that, in response to PGPR inoculation, profiles of rice secondary metabolites were modified, with phenolic compounds such as flavonoids and hydroxycinnamic derivatives being the main metabolites affected. Moreover, plant metabolic changes differed according to Azospirillum strain×cultivar combinations; indeed, 4B induced major secondary metabolic profile modifications only on Cigalon roots, while B510, probably due to its endophytic feature, induced metabolic variations on shoots and roots of both cultivars, triggering a systemic response. Plant secondary metabolite profiling thereby evidences the specific interaction between an Azospirillum strain and its original host cultivar.

Differential Effects of Rare Specific Flavonoids on Compatible and Incompatible Strains in the Myrica gale-Frankia Actinorhizal Symbiosis

ABSTRACT Plant secondary metabolites, and specifically phenolics, play important roles when plants interact with their environment and can act as weapons or positive signals during biotic interactions. One such interaction, the establishment of mutualistic nitrogen-fixing symbioses, typically involves phenolic-based recognition mechanisms between host plants and bacterial symbionts during the early stages of interaction. While these mechanisms are well studied in the rhizobia-legume symbiosis, little is known about the role of plant phenolics in the symbiosis between actinorhizal plants and Frankia genus strains. In this study, the responsiveness of Frankia strains to plant phenolics was correlated with their symbiotic compatibility. We used Myrica gale, a host species with narrow symbiont specificity, and a set of compatible and noncompatible Frankia strains. M. gale fruit exudate phenolics were extracted, and 8 dominant molecules were purified and identified as flavonoids by high-resolution spectroscopic techniques. Total fruit exudates, along with two purified dihydrochalcone molecules, induced modifications of bacterial growth and nitrogen fixation according to the symbiotic specificity of strains, enhancing compatible strains and inhibiting incompatible ones. Candidate genes involved in these effects were identified by a global transcriptomic approach using ACN14a strain whole-genome microarrays. Fruit exudates induced differential expression of 22 genes involved mostly in oxidative stress response and drug resistance, along with the overexpression of a whiB transcriptional regulator. This work provides evidence for the involvement of plant secondary metabolites in determining symbiotic specificity and expands our understanding of the mechanisms, leading to the establishment of actinorhizal symbioses.

Evidence for biological denitrification inhibition (BDI) by plant secondary metabolites.

Previous studies on the effect of secondary metabolites on the functioning of rhizosphere microbial communities have often focused on aspects of the nitrogen (N) cycle but have overlooked biological denitrification inhibition (BDI), which can affect plant N-nutrition. Here, we investigated the BDI by the compounds of Fallopia spp., an invasive weed shown to be associated with a low potential denitrification of the soil. Fallopia spp. extracts were characterized by chromatographic analysis and were used to test the BDI effects on the metabolic and respiratory activities of denitrifying bacteria, under aerobic and anaerobic (denitrification) conditions. The BDI of Fallopia spp. extracts was tested on a complex soil community by measuring denitrification enzyme activity (DEA), substrate induced respiration (SIR), as well as abundances of denitrifiers and total bacteria. In 15 strains of denitrifying bacteria, extracts led to a greater BDI (92%) than respiration inhibition (50%). Anaerobic metabolic activity reduction was correlated with catechin concentrations and the BDI was dose dependent. In soil, extracts reduced the DEA/SIR ratio without affecting the denitrifiers: total bacteria ratio. We show that secondary metabolite(s) from Fallopia spp. inhibit denitrification. This provides new insight into plant-soil interactions and improves our understanding of a plants ability to shape microbial soil functioning.

High-Affinity Binding of Silybin Derivatives to the Nucleotide-Binding Domain of a Leishmania tropica P-Glycoprotein-Like Transporter and Chemosensitization of a Multidrug-Resistant Parasite to Daunomycin

ABSTRACT In order to overcome the multidrug resistance mediated by P-glycoprotein-like transporters in Leishmania spp., we have studied the effects produced by derivatives of the flavanolignan silybin and related compounds lacking the monolignol unit on (i) the affinity of binding to a recombinant C-terminal nucleotide-binding domain of the L. tropica P-glycoprotein-like transporter and (ii) the sensitization to daunomycin on promastigote forms of a multidrug-resistant L. tropica line overexpressing the transporter. Oxidation of the flavanonol silybin to the corresponding flavonol dehydrosilybin, the presence of the monolignol unit, and the addition of a hydrophobic substituent such as dimethylallyl, especially at position 8 of ring A, considerably increased the binding affinity. The in vitro binding affinity of these compounds for the recombinant cytosolic domain correlated with their modulation of drug resistance phenotype. In particular, 8-(3,3-dimethylallyl)-dehydrosilybin effectively sensitized multidrug-resistant Leishmania spp. to daunomycin. The cytosolic domains are therefore attractive targets for the rational design of inhibitors against P-glycoprotein-like transporters.

Phytochemical analysis of mature tree root exudates in situ and their role in shaping soil microbial communities in relation to tree N-acquisition strategy

Eperua falcata (Aublet), a late-successional species in tropical rainforest and one of the most abundant tree in French Guiana, has developed an original strategy concerning N-acquisition by largely preferring nitrate, rather than ammonium (H. Schimann, S. Ponton, S. Hättenschwiler, B. Ferry, R. Lensi, A.M. Domenach, J.C. Roggy, Differing nitrogen use strategies of two tropical rainforest tree species in French Guiana: evidence from (15)N natural abundance and microbial activities, Soil Biol. Biochem. 40 (2008) 487-494). Given the preference of this species for nitrate, we hypothesized that root exudates would promote nitrate availability by (a) enhancing nitrate production by stimulating ammonium oxidation or (b) minimizing nitrate losses by inhibiting denitrification. Root exudates were collected in situ in monospecific planted plots. The phytochemical analysis of these exudates and of several of their corresponding root extracts was achieved using UHPLC/DAD/ESI-QTOF and allowed the identification of diverse secondary metabolites belonging to the flavonoid family. Our results show that (i) the distinct exudation patterns observed are related to distinct root morphologies, and this was associated with a shift in the root flavonoid content, (ii) a root extract representative of the diverse compounds detected in roots showed a significant and selective metabolic inhibition of isolated denitrifiers in vitro, and (iii) in soil plots the abundance of nirK-type denitrifiers was negatively affected in rhizosphere soil compared to bulk. Altogether this led us to formulate hypothesis concerning the ecological role of the identified compounds in relation to N-acquisition strategy of this species.

Phenylpropanoids from leaves of Juniperus phoenicea

Abstract Two new compounds, junipetriolosides A (3-methoxy-4-hydroxy-phenylpropane-7.8-(2′,1′- O -β- d -glucopyranosyl)-7.8,9-triol) and B (3-methoxy-4- O -β- d -glucopyranosyl-phenylpropane-7,8,9-triol) have been isolated from a methanolic extract of the aerial parts of Juniperus phoenicea , along with the rare compound, guaiacylglycerol (3-methoxy-4-hydroxy-phenylpropane-7,8,9,-triol). Structural elucidation of these natural products was achieved mainly by spectroscopic methods.

Analysis of Hydroxycinnamic Acid Degradation in Agrobacterium fabrum Reveals a Coenzyme A-Dependent, Beta-Oxidative Deacetylation Pathway

ABSTRACT The soil- and rhizosphere-inhabiting bacterium Agrobacterium fabrum (genomospecies G8 of the Agrobacterium tumefaciens species complex) is known to have species-specific genes involved in ferulic acid degradation. Here, we characterized, by genetic and analytical means, intermediates of degradation as feruloyl coenzyme A (feruloyl-CoA), 4-hydroxy-3-methoxyphenyl-β-hydroxypropionyl–CoA, 4-hydroxy-3-methoxyphenyl-β-ketopropionyl–CoA, vanillic acid, and protocatechuic acid. The genes atu1416, atu1417, and atu1420 have been experimentally shown to be necessary for the degradation of ferulic acid. Moreover, the genes atu1415 and atu1421 have been experimentally demonstrated to be essential for this degradation and are proposed to encode a phenylhydroxypropionyl-CoA dehydrogenase and a 4-hydroxy-3-methoxyphenyl-β-ketopropionic acid (HMPKP)–CoA β-keto-thiolase, respectively. We thus demonstrated that the A. fabrum hydroxycinnamic degradation pathway is an original coenzyme A-dependent β-oxidative deacetylation that could also transform p-coumaric and caffeic acids. Finally, we showed that this pathway enables the metabolism of toxic compounds from plants and their use for growth, likely providing the species an ecological advantage in hydroxycinnamic-rich environments, such as plant roots or decaying plant materials.