Differential Frankia protein patterns induced by phenolic extracts from Myricaceae seeds

Two-dimensional gel electrophoresis was used to identify differentially displayed proteins expressed during the early symbiotic interactions between the bacterium Frankia and actinorhizal plants. Myricaceae, the most primitive actinorhizal family, was used as an experimental model to study specificity mechanisms because it includes species with either narrowor large specificity. Seed phenolic extracts from two Myricaceae species, Myrica gale, a narrow specificity host and Morella cerifera considered as a promiscuous host, were used to induce three Frankia strains ACN14a, M16467 and Ea112. The global protein pattern was altered by exposure to the plant extracts. The addition of 30 mg l21 of seed phenolic extracts provoked the inhibition of many protein biosynthesis whereas 1 and 10 mg l21 induced a global reprogramming of Frankia protein pattern. Changes in intensity of 115 spots in response to seed extracts were detected and analyzed by matrix-assisted laser desorption/ ionization time of flight mass spectrometry. Fifty proteins were efficiently identified with Frankia protein data banks deduced from the sequences of Frankia strains ACN14a and EaN1pec genomes. Differential proteins were involved in different metabolism pathways such as glycolysis and gluconeogenesis, transcription, fatty acids, carbohydrates, coenzymes and purines metabolisms. Chaperones biosynthesis and iron transport regulation, essential for nitrogen fixation, seem to be strain dependant. Several proteins possibly involved in the regulation of nodulation were also differentially expressed. The most obvious response was the upregulation of oxidative stress proteins such as FeSOD and Tellurium resistance proteins, suggesting a reorganization of Frankia metabolism to protect against host plant defense

Frankia alni proteome under nitrogen-fixing and nitrogen-replete conditions

Frankia alni induces root nodules on Alnus, in which the bacterium differentiates into nitrogen (N)-fixing cells called vesicles. In culture, F. alni also undergoes major morphological changes as it alternates between N-replete and N-fixing conditions. Lack of biologically available N induces the synthesis of vesicles in which nitrogenase is protected from molecular oxygen by a thick lipid hopanoid envelope. Very little is known about the molecular basis of Frankia–host interaction as well as Frankia cell differentiation. The recent determination of the complete genome sequence of F. alni strain ACN14a has permitted us to characterize its proteome, particularly in the extracellular compartment, which could be involved in Frankia–host interaction, and in the switch from N-replete to N-fixing conditions. To that end, 126 bacterial proteins were analyzed by two-dimensional protein gel electrophoresis and identified by matrix-assisted laser desorption/ionization time of flight fingerprinting using a F. alni proteome database. Interestingly, the extracellular fraction contains some glycolytic enzymes lacking secretion signals, already reported to be extracellularly localized in some streptococci, as well as some abundant stress-resistance proteins. As expected, several proteins involved in N assimilation and oxidative defense system were upregulated in F. alni grown under N-fixing vs N-replete conditions. Furthermore, two Raf kinase inhibitor protein homologs that could play a role in cellular signaling, and a hemoglobin-like protein HbN that could be involved in detoxification of nitric oxide were also upregulated. More surprising, a succinate dehydrogenase was strongly downregulated, which could be linked to the need of pyruvate for the biosynthesis of hopanoids or to reduced oxygen diffusion in vesicles

Comparative phylogenomics and phylotranscriptomics provide insights into the genetic complexity of nitrogen-fixing root-nodule symbiosis.

Plant root-nodule symbiosis (RNS) with mutualistic nitrogen-fixing bacteria is restricted to a single clade of angiosperms, the Nitrogen-Fixing Nodulation Clade (NFNC), and is best understood in the legume family. Nodulating species share many commonalities, explained either by divergence from a common ancestor over 100 million years ago or by convergence following independent origins over that same time period. Regardless, comparative analyses of diverse nodulation syndromes can provide insights into constraints on nodulation-what must be acquired or cannot be lost for a functional symbiosis-and the latitude for variation in the symbiosis. However, much remains to be learned about nodulation, especially outside of legumes. Here, we employed a large-scale phylogenomic analysis across 88 species, complemented by 151 RNA-seq libraries, to elucidate the evolution of RNS. Our phylogenomic analyses further emphasize the uniqueness of the transcription factor NIN as a master regulator of nodulation and identify key mutations that affect its function across the NFNC. Comparative transcriptomic assessment revealed nodule-specific upregulated genes across diverse nodulating plants, while also identifying nodule-specific and nitrogen-response genes. Approximately 70% of symbiosis-related genes are highly conserved in the four representative species, whereas defense-related and host-range restriction genes tend to be lineage specific. Our study also identified over 900 000 conserved non-coding elements (CNEs), over 300 000 of which are unique to sampled NFNC species. NFNC-specific CNEs are enriched with the active H3K9ac mark and are correlated with accessible chromatin regions, thus representing a pool of candidate regulatory elements for genes involved in RNS. Collectively, our results provide novel insights into the evolution of nodulation and lay a foundation for engineering of RNS traits in agriculturally important crops

Comparative phylogenomics and phylotranscriptomics provide insights into the genetic complexity of nitrogen-fixing root-nodule symbiosis

International audiencePlant root-nodule symbiosis (RNS) with mutualistic nitrogen-fixing bacteria is restricted to a single clade of angiosperms, the Nitrogen-Fixing Nodulation Clade (NFNC), and is best understood in the legume family. Nodulating species share many commonalities, explained either by divergence from a common ancestor over 100 million years ago or by convergence following independent origins over that same time period. Regardless, comparative analyses of diverse nodulation syndromes can provide insights into constraints on nodulation—what must be acquired or cannot be lost for a functional symbiosis—and the latitude for variation in the symbiosis. However, much remains to be learned about nodulation, especially outside of legumes. Here, we employed a large-scale phylogenomic analysis across 88 species, complemented by 151 RNA-seq libraries, to elucidate the evolution of RNS. Our phylogenomic analyses further emphasize the uniqueness of the transcription factor NIN as a master regulator of nodulation and identify key mutations that affect its function across the NFNC. Comparative transcriptomic assessment revealed nodule-specific upregulated genes across diverse nodulating plants, while also identifying nodule-specific and nitrogen-response genes. Approximately 70% of symbiosis-related genes are highly conserved in the four representative species, whereas defense-related and host-range restriction genes tend to be lineage specific. Our study also identified over 900 000 conserved non-coding elements (CNEs), over 300 000 of which are unique to sampled NFNC species. NFNC-specific CNEs are enriched with the active H3K9ac mark and are correlated with accessible chromatin regions, thus representing a pool of candidate regulatory elements for genes involved in RNS. Collectively, our results provide novel insights into the evolution of nodulation and lay a foundation for engineering of RNS traits in agriculturally important crop

Genome characteristics of facultatively symbiotic Frankia sp. strains reflect host range and host plant biogeography

Soil bacteria that also form mutualistic symbioses in plants encounter two major levels of selection. One occurs during adaptation to and survival in soil, and the other occurs in concert with host plant speciation and adaptation. Actinobacteria from the genus Frankia are facultative symbionts that form N(2)-fixing root nodules on diverse and globally distributed angiosperms in the “actinorhizal” symbioses. Three closely related clades of Frankia sp. strains are recognized; members of each clade infect a subset of plants from among eight angiosperm families. We sequenced the genomes from three strains; their sizes varied from 5.43 Mbp for a narrow host range strain (Frankia sp. strain HFPCcI3) to 7.50 Mbp for a medium host range strain (Frankia alni strain ACN14a) to 9.04 Mbp for a broad host range strain (Frankia sp. strain EAN1pec.) This size divergence is the largest yet reported for such closely related soil bacteria (97.8%–98.9% identity of 16S rRNA genes). The extent of gene deletion, duplication, and acquisition is in concert with the biogeographic history of the symbioses and host plant speciation. Host plant isolation favored genome contraction, whereas host plant diversification favored genome expansion. The results support the idea that major genome expansions as well as reductions can occur in facultative symbiotic soil bacteria as they respond to new environments in the context of their symbioses

Phylogenomics reveals multiple losses of nitrogen-fixing root nodule symbiosis

The root nodule symbiosis of plants with nitrogen-fixing bacteria affects global nitrogen cycles and food production but is restricted to a subset of genera within a single clade of flowering plants. To explore the genetic basis for this scattered occurrence, we sequenced the genomes of 10 plant species covering the diversity of nodule morphotypes, bacterial symbionts, and infection strategies. In a genome-wide comparative analysis of a total of 37 plant species, we discovered signatures of multiple independent loss-of-function events in the indispensable symbiotic regulator NODULE INCEPTION in 10 of 13 genomes of nonnodulating species within this clade. The discovery that multiple independent losses shaped the present-day distribution of nitrogen-fixing root nodule symbiosis in plants reveals a phylogenetically wider distribution in evolutionary history and a so-far-underestimated selection pressure against this symbiosis.Fil: Griesmann, Maximilian. Ludwig Maximilians Universitat; AlemaniaFil: Chang, Yue. No especifíca;Fil: Liu, Xin. No especifíca;Fil: Song, Yue. No especifíca;Fil: Haberer, Georg. Helmholtz Center Munich; AlemaniaFil: Crook, Matthew B.. Weber State University; Estados UnidosFil: Billault-Penneteau, Benjamin. Ludwig Maximilians Universitat; AlemaniaFil: Lauressergues, Dominique. Université de Toulouse; FranciaFil: Keller, Jean. Université de Toulouse; FranciaFil: Imanishi, Leandro Ezequiel. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Roswanjaya, Yuda Purwana. University of Agriculture Wageningen; Países BajosFil: Kohlen, Wouter. University of Agriculture Wageningen; Países BajosFil: Pujic, Petar. Université de Lyon; FranciaFil: Battenberg, Kai. University of California at Davis; Estados UnidosFil: Alloisio, Nicole. No especifíca;Fil: Liang, Yuhu. No especifíca;Fil: Hilhorst, Henk. No especifíca;Fil: Salgado, Marco G.. Stockholms Universitet; SueciaFil: Hocher, Valerie. Université Montpellier II; FranciaFil: Gherbi, Hassen. Université Montpellier II; FranciaFil: Svistoonoff, Sergio. Université Montpellier II; FranciaFil: Doyle, Jeff J.. Cornell University; Estados UnidosFil: He, Shixu. No especifíca;Fil: Xu, Yan. China National Genebank; ChinaFil: Xu, Shanyun. China National Genebank; ChinaFil: Qu, Jing. China National Genebank; ChinaFil: Gao, Qiang. No especifíca;Fil: Fang, Xiaodong. No especifíca;Fil: Fu, Yuan. China National Genebank; ChinaFil: Normand, Philippe. Universite Lyon 2; FranciaFil: Berry, Alison M.. University of California at Davis; Estados UnidosFil: Wall, Luis Gabriel. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Ané, Jean Michel. University of Wisconsin; Estados UnidosFil: Pawlowski, Katharina. Stockholms Universitet; SueciaFil: Xu, Xun. China National Genebank; ChinaFil: Yang, Huanming. James D. Watson Institute Of Genome Sciences; ChinaFil: Spannagl, Manuel. Helmholtz Center Munich German Research Center For Environmental Health; AlemaniaFil: Mayer, Klaus F.X.. Helmholtz Center Munich German Research Center For Environmental Health; AlemaniaFil: Wong, Gane Ka-Shu. University of Alberta; CanadáFil: Parniske, Martin. Ludwig Maximilians Universitat; AlemaniaFil: Delaux, Pierre Marc. No especifíca;Fil: Cheng, Shifeng. China National Genebank; Chin