Extracellular Vesicle-Driven Crosstalk between Legume Plants and Rhizobia: The Peribacteroid Space of Symbiosomes as a Protein Trafficking Interface.

Prokaryotes and eukaryotes secrete extracellular vesicles (EVs) into the surrounding milieu to preserve and transport elevated concentrations of biomolecules across long distances. EVs encapsulate metabolites, DNA, RNA, and proteins, whose abundance and composition fluctuate depending on environmental cues. EVs are involved in eukaryote-to-prokaryote communication owing to their ability to navigate different ecological niches and exchange molecular cargo between the two domains.

Exopolysaccharide is detrimental for the symbiotic performance of Sinorhizobium fredii HH103 mutants with a truncated lipopolysaccharide core.

The nitrogen-fixing rhizobia-legume symbiosis relies on a complex interchange of molecular signals between the two partners during the whole interaction. On the bacterial side, different surface polysaccharides, such as lipopolysaccharide (LPS) and exopolysaccharide (EPS), might play important roles for the success of the interaction. In a previous work we studied two Sinorhizobium fredii HH103 mutants affected in the rkpK and lpsL genes, which are responsible for the production of glucuronic acid and galacturonic acid, respectively.

Microscope Subcellular Localization of Plant-Interacting Bacterial Effectors in Animal Cell Cultures.

Eukaryote-interacting bacteria have developed along the evolution of an arsenal of tools to interact with potential hosts and to evade their defensive responses. Among these tools, the effector proteins are gaining a special importance due to the high diversity of molecular actions that they play in the host cell, with the final aim of taking the control over the cell. Bacteria inject these effectors into the cytosol of the host cells through distinct ways, as the type III secretion system.

A complex regulatory network governs the expression of symbiotic genes in HH103.

Introduction: The establishment of the rhizobium-legume nitrogen-fixing symbiosis relies on the interchange of molecular signals between the two symbionts. We have previously studied by RNA-seq the effect of the symbiotic regulators NodD1, SyrM, and TtsI on the expression of the symbiotic genes (the regulon) of HH103 upon treatment with the isoflavone genistein. In this work we have further investigated this regulatory network by incorporating new RNA-seq data of HH103 mutants in two other regulatory genes, and .

Membrane vesicle engineering with "à la carte" bacterial-immunogenic molecules for organism-free plant vaccination.

The United Nations heralds a world population exponential increase exceeding 9.7 billion by 2050. This poses the challenge of covering the nutritional needs of an overpopulated world by the hand of preserving the environment.

Multitask Approach to Localize Rhizobial Type Three Secretion System Effector Proteins Inside Eukaryotic Cells.

Rhizobia can establish mutually beneficial interactions with legume plants by colonizing their roots to induce the formation of a specialized structure known as a nodule, inside of which the bacteria are able to fix atmospheric nitrogen. It is well established that the compatibility of such interactions is mainly determined by the bacterial recognition of flavonoids secreted by the plants, which in response to these flavonoids trigger the synthesis of the bacterial Nod factors that drive the nodulation process. Additionally, other bacterial signals are involved in the recognition and the efficiency of this interaction, such as extracellular polysaccharides or some secreted proteins.

The Rhizobial Type 3 Secretion System: The Dr. Jekyll and Mr. Hyde in the Rhizobium-Legume Symbiosis.

Rhizobia are soil bacteria that can establish a symbiotic association with legumes. As a result, plant nodules are formed on the roots of the host plants where rhizobia differentiate to bacteroids capable of fixing atmospheric nitrogen into ammonia. This ammonia is transferred to the plant in exchange of a carbon source and an appropriate environment for bacterial survival.

The Rhizobium tropici CIAT 899 NodD2 protein promotes symbiosis and extends rhizobial nodulation range by constitutive nodulation factor synthesis.

In the symbiotic associations between rhizobia and legumes, the NodD regulators orchestrate the transcription of the specific nodulation genes. This set of genes is involved in the synthesis of nodulation factors, which are responsible for initiating the nodulation process. Rhizobium tropici CIAT 899 is the most successful symbiont of Phaseolus vulgaris and can nodulate a variety of legumes.

The Gene of HH103 Restores Nodulation Capacity on Bean in a CIAT 899 / Mutant, but the Secondary Symbiotic Regulators , or Prevent HH103 to Nodulate with This Legume.

Rhizobial NodD proteins and appropriate flavonoids induce rhizobial nodulation gene expression. In this study, we show that the gene of HH103, but not the gene, can restore the nodulation capacity of a double / mutant of CIAT 899 in bean plants (). HH103 only induces pseudonodules in beans.

OnfD, an AraC-Type Transcriptional Regulator Encoded by CIAT 899 and Involved in Nod Factor Synthesis and Symbiosis.

CIAT 899 is a broad-host-range rhizobial strain that establishes symbiotic interactions with legumes and tolerates different environmental stresses such as heat, acidity, or salinity. This rhizobial strain produces a wide variety of symbiotically active nodulation factors (NF) induced not only by the presence of plant-released flavonoids but also under osmotic stress conditions through the LysR-type transcriptional regulators NodD1 (flavonoids) and NodD2 (osmotic stress). However, the activation of NodD2 under high-osmotic-stress conditions remains elusive.

The Sinorhizobium fredii HH103 type III secretion system effector NopC blocks nodulation with Lotus japonicus Gifu.

The broad-host-range bacterium Sinorhizobium fredii HH103 cannot nodulate the model legume Lotus japonicus Gifu. This bacterium possesses a type III secretion system (T3SS), a specialized secretion apparatus used to deliver effector proteins (T3Es) into the host cell cytosol to alter host signaling and/or suppress host defence responses to promote infection. However, some of these T3Es are recognized by specific plant receptors and hence trigger a strong defence response to block infection.

Regulation of Protein Secretion Systems Mediated by Cyclic Diguanylate in Plant-Interacting Bacteria.

The ubiquitous second messenger cyclic diguanylate (c-di-GMP) is involved in the regulation of different processes in bacteria. In phytopathogens, intracellular fluctuations in the concentration of this molecule contribute to the lifestyle switching from a motile and virulent stage to a sessile and biofilm-forming phase. Among the virulence mechanisms used by bacterial pathogens, different specific type secretion systems (TSSs) and the effector proteins that they translocate are included.

Osmotic stress activates nif and fix genes and induces the Rhizobium tropici CIAT 899 Nod factor production via NodD2 by up-regulation of the nodA2 operon and the nodA3 gene.

The symbiosis between rhizobia and legumes is characterized by a complex molecular dialogue in which the bacterial NodD protein plays a major role due to its capacity to activate the expression of the nodulation genes in the presence of appropiate flavonoids. These genes are involved in the synthesis of molecules, the nodulation factors (NF), responsible for launching the nodulation process. Rhizobium tropici CIAT 899, a rhizobial strain that nodulates Phaseolus vulgaris, is characterized by its tolerance to multiple environmental stresses such as high temperatures, acidity or elevated osmolarity.

Sinorhizobium fredii HH103 nolR and nodD2 mutants gain capacity for infection thread invasion of Lotus japonicus Gifu and Lotus burttii.

Sinorhizobium fredii HH103 Rif , a broad-host-range rhizobial strain, forms ineffective nodules with Lotus japonicus but induces nitrogen-fixing nodules in Lotus burttii roots that are infected by intercellular entry. Here we show that HH103 Rif nolR or nodD2 mutants gain the ability to induce infection thread formation and to form nitrogen-fixing nodules in L. japonicus Gifu.

Strains HH103 and NGR234 Form Nitrogen Fixing Nodules With Diverse Wild Soybeans () From Central China but Are Ineffective on Northern China Accessions.

indigenous populations are prevalent in provinces of Central China whereas species (, , , and others) are more abundant in northern and southern provinces. The symbiotic properties of different soybean rhizobia have been investigated with 40 different wild soybean () accessions from China, Japan, Russia, and South Korea. Bradyrhizobial strains nodulated all the wild soybeans tested, albeit efficiency of nitrogen fixation varied considerably among accessions.

Transcriptomic Studies of the Effect of nod Gene-Inducing Molecules in Rhizobia: Different Weapons, One Purpose.

Simultaneous quantification of transcripts of the whole bacterial genome allows the analysis of the global transcriptional response under changing conditions. RNA-seq and microarrays are the most used techniques to measure these transcriptomic changes, and both complement each other in transcriptome profiling. In this review, we exhaustively compiled the symbiosis-related transcriptomic reports (microarrays and RNA sequencing) carried out hitherto in rhizobia.

The Rhizobium tropici CIAT 899 NodD2 protein regulates the production of Nod factors under salt stress in a flavonoid-independent manner.

In the symbiotic associations between rhizobia and legumes, NodD promotes the expression of the nodulation genes in the presence of appropriate flavonoids. This set of genes is implied in the synthesis of Nodulation factors, which are responsible for launching the nodulation process. Rhizobium tropici CIAT 899 is the most successful symbiont of Phaseolus vulgaris and can nodulate a variety of legumes.

Studies of rhizobial competitiveness for nodulation in soybean using a non-destructive split-root system.

Split-root systems (SRS) constitute an appropriate methodology for studying the relevance of both local and systemic mechanisms that participate in the control of rhizobia-legume symbioses. In fact, this kind of approach allowed to demonstrate the autoregulation of nodulation (AON) systemic response in soybean in the 1980s. In SRS, the plant main root is cut and two lateral roots that emerge from the seedlings after root-tip removal are confined into separate compartments.

The Sinorhizobium (Ensifer) fredii HH103 Nodulation Outer Protein NopI Is a Determinant for Efficient Nodulation of Soybean and Cowpea Plants.

The type III secretion system (T3SS) is a specialized secretion apparatus that is commonly used by many plant and animal pathogenic bacteria to deliver proteins, termed effectors, to the interior of the host cells. These effectors suppress host defenses and interfere with signal transduction pathways to promote infection. Some rhizobial strains possess a functional T3SS, which is involved in the suppression of host defense responses, host range determination, and symbiotic efficiency.

RNA-seq analysis of the Rhizobium tropici CIAT 899 transcriptome shows similarities in the activation patterns of symbiotic genes in the presence of apigenin and salt.

Background: Rhizobium tropici strain CIAT 899 establishes effective symbioses with several legume species, including Phaseolus vulgaris and Leucaena leucocephala. This bacterium synthesizes a large variety of nodulation factors in response to nod-gene inducing flavonoids and, surprisingly, also under salt stress conditions. The aim of this study was to identify differentially expressed genes in the presence of both inducer molecules, and analyze the promoter regions located upstream of these genes.

NopC Is a Rhizobium-Specific Type 3 Secretion System Effector Secreted by Sinorhizobium (Ensifer) fredii HH103.

Sinorhizobium (Ensifer) fredii HH103 is a broad host-range nitrogen-fixing bacterium able to nodulate many legumes, including soybean. In several rhizobia, root nodulation is influenced by proteins secreted through the type 3 secretion system (T3SS). This specialized secretion apparatus is a common virulence mechanism of many plant and animal pathogenic bacteria that delivers proteins, called effectors, directly into the eukaryotic host cells where they interfere with signal transduction pathways and promote infection by suppressing host defenses.

The Sinorhizobium (Ensifer) fredii HH103 Type 3 Secretion System Suppresses Early Defense Responses to Effectively Nodulate Soybean.

Plants that interact with pathogenic bacteria in their natural environments have developed barriers to block or contain the infection. Phytopathogenic bacteria have evolved mechanisms to subvert these defenses and promote infection. Thus, the type 3 secretion system (T3SS) delivers bacterial effectors directly into the plant cells to alter host signaling and suppress defenses, providing an appropriate environment for bacterial multiplication.

The symbiotic biofilm of Sinorhizobium fredii SMH12, necessary for successful colonization and symbiosis of Glycine max cv Osumi, is regulated by Quorum Sensing systems and inducing flavonoids via NodD1.

Bacterial surface components, especially exopolysaccharides, in combination with bacterial Quorum Sensing signals are crucial for the formation of biofilms in most species studied so far. Biofilm formation allows soil bacteria to colonize their surrounding habitat and survive common environmental stresses such as desiccation and nutrient limitation. This mode of life is often essential for survival in bacteria of the genera Mesorhizobium, Sinorhizobium, Bradyrhizobium, and Rhizobium.

Rice and bean AHL-mimic quorum-sensing signals specifically interfere with the capacity to form biofilms by plant-associated bacteria.

Many bacteria regulate their gene expression in response to changes in their population density in a process called quorum sensing (QS), which involves communication between cells mediated by small diffusible signal molecules termed autoinducers. n-acyl-homoserine-lactones (AHLs) are the most common autoinducers in proteobacteria. QS-regulated genes are involved in complex interactions between bacteria of the same or different species and even with some eukaryotic organisms.

Nodulation-gene-inducing flavonoids increase overall production of autoinducers and expression of N-acyl homoserine lactone synthesis genes in rhizobia.

Legume-nodulating rhizobia use N-acyl homoserine lactones (AHLs) to regulate several physiological traits related to the symbiotic plant-microbe interaction. In this work, we show that Sinorhizobium fredii SMH12, Rhizobium etli ISP42 and Rhizobium sullae IS123, three rhizobial strains with different nodulation ranges, produced a similar pattern of AHL molecules, sharing, in all cases, production of N-octanoyl homoserine lactone and its 3-oxo and/or 3-hydroxy derivatives. Interestingly, production of AHLs was enhanced when these three rhizobia were grown in the presence of their respective nod-gene-inducing flavonoid, while a new molecule, C14-HSL, was produced by S.