Symbiotic Cells Research Articles

The jasmonate pathway promotes nodule symbiosis and suppresses host plant defense in Medicago truncatula

Root nodule symbiosis (RNS) between legumes and rhizobia is a major source of nitrogen in agricultural systems. Effective symbiosis requires precise regulation of plant defense responses. The role of the defense hormone jasmonic acid (JA) in the immune response has been extensively studied. Current research shows that JA can play either a positive or negative regulatory role in RNS depending on its concentration, but the molecular mechanisms remain to be elucidated. In this study, we found that inoculation with the rhizobia Sm1021 induces the JA pathway in Medicago truncatula, and blocking the JA pathway significantly reduces the number of infection threads. Mutations in the MtMYC2 gene, which encodes a JA signaling master transcription factor, significantly inhibited rhizobia infection, terminal differentiation, and symbiotic cell formation. Combining RNA sequencing and chromatin immunoprecipitation sequencing, we discovered that MtMYC2 regulates the expression of nodule-specific MtDNF2, MtNAD1, and MtSymCRK to suppress host defense, while it activates MtDNF1 expression to regulate the maturation of MtNCRs, which in turn promotes bacteroid formation. More importantly, MtMYC2 participates in symbiotic signal transduction by promoting the expression of MtIPD3. Notably, the MtMYC2-MtIPD3 transcriptional regulatory module is specifically present in legumes, and the Mtmyc2 mutants are susceptible to the infection by the pathogen Rhizoctonia solani. Collectively, these findings reveal the molecular mechanisms of how the JA pathway regulates RNS, broadening our understanding of the roles of JA in plant-microbe interactions.

A pathogenesis-related protein, PRP1, negatively regulates root nodule symbiosis in Lotus japonicus.

The legume-rhizobium symbiosis represents a unique model within the realm of plant-microbe interactions. Unlike typical cases of pathogenic invasion, the infection of rhizobia and their residence within symbiotic cells do not elicit a noticeable immune response in plants. Nevertheless, there is still much to uncover regarding the mechanisms through which plant immunity influences rhizobial symbiosis. In this study, we identify an important player in this intricate interplay: Lotus japonicus PRP1, which serves as a positive regulator of plant immunity but also exhibits the capacity to decrease rhizobial colonization and nitrogen fixation within nodules. The PRP1 gene encodes an uncharacterized protein and is named Pathogenesis-Related Protein1, owing to its orthologue in Arabidopsis thaliana, a pathogenesis-related family protein (At1g78780). The PRP1 gene displays high expression levels in nodules compared to other tissues. We observed an increase in rhizobium infection in the L. japonicus prp1 mutants, whereas PRP1-overexpressing plants exhibited a reduction in rhizobium infection compared to control plants. Intriguingly, L. japonicus prp1 mutants produced nodules with a pinker colour compared to wild-type controls, accompanied by elevated levels of leghaemoglobin and an increased proportion of infected cells within the prp1 nodules. The transcription factor Nodule Inception (NIN) can directly bind to the PRP1 promoter, activating PRP1 gene expression. Furthermore, we found that PRP1 is a positive mediator of innate immunity in plants. In summary, our study provides clear evidence of the intricate relationship between plant immunity and symbiosis. PRP1, acting as a positive regulator of plant immunity, simultaneously exerts suppressive effects on rhizobial infection and colonization within nodules.

The human gut fungiome: Role in physiology and detoxification

The intestinal microbiota is an ecosystem where bacteria, archaea, viruses, and protists, are entangled, but not alone. We take microbiota as the bacterial community because, in many historical papers, the probes to detect other organisms in the intestine were rarely used. But in addition, unicellular fungi or yeasts do exist in microbiota, their assembly is called the mycome or fungiome, and it can reach the size and number of our human cells. The bacteria are far more numerous. For a long time, the whole was taken for a nest of pathogens, but in fact, yeasts adapted and evolved as symbiotic cells helping not only our digestion, but also angiogenesis, tissue growth, vitamin synthesis, and our nervous, endocrine, immune systems and even our detoxification system. Much of what we swallow is or will become fermented, like bread, cheese, or wine. Since the beginning of agriculture, our food is eaten lightly by microbiota including yeasts first in plants not treated by pesticides. Natural yeasts still unknown are in hundreds of types or species at the very least, bringing in particular during their work of fermentation a multiplicity of aromas that play a role in the famous detoxification; but the usual diet only provides a few strains of these yeasts. In yeasts like in all cells, ubiquitous cytochrome P450 detoxifying enzymes form a vast family also involved in cell respiration, which is stimulated to some extent by the aromas and other signals secreted. Yeasts could attract and coordinate bacteria to eliminate in part chemical pollutants.

Coral holobiont research needs spatial analyses at the microbial scale.

Coral holobiont research needs spatial analyses at the microbial scale.

Identifying ligand-binding specificity of the oligopeptide receptor OppA from Bifidobacterium longum KACC91563 by structure-based molecular modeling

As a ATP-binding cassette (ABC) transporter the OppA receptor plays key roles in protecting the host organism and transporting nutrients across the intestine by the oligopeptide transporter from symbiotic bacteria and directs maturation of the host immune system. Among lactic acid bacteria, Bifidobacterium longum KACC91563, isolated from fecal samples of healthy Korean neonates, has the capability to alleviate food allergy effects. Operating as a peptide importer, the extracellular OppA receptor from gram-positive B. longum KACC91563 translocates nutrients, specifically peptides, from the outside environment of the intestinal tract to the inside of symbiotic cells. In the present study we attempt to explicate the relationship between the substrate’s specificity from the OppA importer and the probiotic effects of B. longum KACC91563 in the host intestine. It was first identified in this study the specialized structure–function relationship from the OppA importer of B. longum KACC91563 with its structural and functional determinants. This could provide insights into substrate specificity of unique immunological properties and a key switch for the substrate’s metabolism to reprogramming immune responses in the host intestine by structure-based molecular modeling. The probiotic effects of oligopeptide substrate (such as a proline-rich peptide containing at least one branched residue of leucine, isoleucine, and valine) and its metabolism for the OppA from B. longum KACC91563 are attributed to enhancement of the epithelial barrier by several different strain specific pathways to prevent the strong adhesion of pathogens.

A legume kinesin controls vacuole morphogenesis for rhizobia endosymbiosis.

Symbioses between legumes and rhizobia require establishment of the plant-derived symbiosome membrane, which surrounds the rhizobia and accommodates the symbionts by providing an interface for nutrient and signal exchange. The host cytoskeleton and endomembrane trafficking systems play central roles in the formation of a functional symbiotic interface for rhizobia endosymbiosis; however, the underlying mechanisms remain largely unknown. Here we demonstrate that the nodulation-specific kinesin-like calmodulin-binding protein (nKCBP), a plant-specific microtubule-based kinesin motor, controls central vacuole morphogenesis in symbiotic cells in Medicago truncatula. Phylogenetic analysis further indicated that nKCBP duplication occurs solely in legumes of the clade that form symbiosomes. Knockout of nKCBP results in central vacuole deficiency, defective symbiosomes and abolished nitrogen fixation. nKCBP decorates linear particles along microtubules, and crosslinks microtubules with the actin cytoskeleton, to control central vacuole formation by modulating vacuolar vesicle fusion in symbiotic cells. Together, our findings reveal that rhizobia co-opted nKCBP to achieve symbiotic interface formation by regulating cytoskeletal assembly and central vacuole morphogenesis during nodule development.

Swimming and the human microbiome at the intersection of sports, clinical, and environmental sciences: A scoping review of the literature.

The human microbiota is comprised of more than 10–100 trillion microbial taxa and symbiotic cells. Two major human sites that are host to microbial communities are the gut and the skin. Physical exercise has favorable effects on the structure of human microbiota and metabolite production in sedentary subjects. Recently, the concept of “athletic microbiome” has been introduced. To the best of our knowledge, there exists no review specifically addressing the potential role of microbiomics for swimmers, since each sports discipline requires a specific set of techniques, training protocols, and interactions with the athletic infrastructure/facility. Therefore, to fill in this gap, the present scoping review was undertaken. Four studies were included, three focusing on the gut microbiome, and one addressing the skin microbiome. It was found that several exercise-related variables, such as training volume/intensity, impact the athlete’s microbiome, and specifically the non-core/peripheral microbiome, in terms of its architecture/composition, richness, and diversity. Swimming-related power-/sprint- and endurance-oriented activities, acute bouts and chronic exercise, anaerobic/aerobic energy systems have a differential impact on the athlete’s microbiome. Therefore, their microbiome can be utilized for different purposes, including talent identification, monitoring the effects of training methodologies, and devising ad hoc conditioning protocols, including dietary supplementation. Microbiomics can be exploited also for clinical purposes, assessing the effects of exposure to swimming pools and developing potential pharmacological strategies to counteract the insurgence of skin infections/inflammation, including acne. In conclusion, microbiomics appears to be a promising tool, even though current research is still limited, warranting, as such, further studies.

Legume Plant Peptides as Sources of Novel Antimicrobial Molecules Against Human Pathogens.

Antimicrobial peptides are prominent components of the plant immune system acting against a wide variety of pathogens. Legume plants from the inverted repeat lacking clade (IRLC) have evolved a unique gene family encoding nodule-specific cysteine-rich NCR peptides acting in the symbiotic cells of root nodules, where they convert their bacterial endosymbionts into non-cultivable, polyploid nitrogen-fixing cells. NCRs are usually 30–50 amino acids long peptides having a characteristic pattern of 4 or 6 cysteines and highly divergent amino acid composition. While the function of NCRs is largely unknown, antimicrobial activity has been demonstrated for a few cationic Medicago truncatula NCR peptides against bacterial and fungal pathogens. The advantages of these plant peptides are their broad antimicrobial spectrum, fast killing modes of actions, multiple bacterial targets, and low propensity to develop resistance to them and no or low cytotoxicity to human cells. In the IRLC legumes, the number of NCR genes varies from a few to several hundred and it is possible that altogether hundreds of thousands of different NCR peptides exist. Due to the need for new antimicrobial agents, we investigated the antimicrobial potential of 104 synthetic NCR peptides from M. truncatula, M. sativa, Pisum sativum, Galega orientalis and Cicer arietinum against eight human pathogens, including ESKAPE bacteria. 50 NCRs showed antimicrobial activity with differences in the antimicrobial spectrum and effectivity. The most active peptides eliminated bacteria at concentrations from 0.8 to 3.1 μM. High isoelectric point and positive net charge were important but not the only determinants of their antimicrobial activity. Testing the activity of shorter peptide derivatives against Acinetobacter baumannii and Candida albicans led to identification of regions responsible for the antimicrobial activity and provided insight into their potential modes of action. This work provides highly potent lead molecules without hemolytic activity on human blood cells for novel antimicrobial drugs to fight against pathogens.

Ploidy dynamics in aphid host cells harboring bacterial symbionts

Aphids have evolved bacteriocytes or symbiotic host cells that harbor the obligate mutualistic bacterium Buchnera aphidicola. Because of the large cell size (approximately 100 μm in diameter) of bacteriocytes and their pivotal role in nutritional symbiosis, researchers have considered that these cells are highly polyploid and assumed that bacteriocyte polyploidy may be essential for the symbiotic relationship between the aphid and the bacterium. However, little is known about the ploidy levels and dynamics of aphid bacteriocytes. Here, we quantitatively analyzed the ploidy levels in the bacteriocytes of the pea-aphid Acyrthosiphon pisum. Image-based fluorometry revealed the hyper polyploidy of the bacteriocytes ranging from 16- to 256-ploidy throughout the lifecycle. Bacteriocytes of adult parthenogenetic viviparous females were ranged between 64 and 128C DNA levels, while those of sexual morphs (oviparous females and males) were comprised of 64C, and 32–64C cells, respectively. During post-embryonic development of viviparous females, the ploidy level of bacteriocytes increased substantially, from 16 to 32C at birth to 128–256C in actively reproducing adults. These results suggest that the ploidy levels are dynamically regulated among phenotypes and during development. Our comprehensive and quantitative data provides a foundation for future studies to understand the functional roles and biological significance of the polyploidy of insect bacteriocytes.

Ascidian-associated photosymbionts from Manado, Indonesia: secondary metabolites, bioactivity simulation, and biosynthetic insight

This research work aimed at investigating the chemistry and biosynthetic potential of ascidian-associated symbionts that originated from Manado Bay, North Sulawesi. We initially enriched the symbiotic cells associated with the Manadonese ascidian Lissoclinum patella. Subsequently we identified the presence of Prochloron didemni in both unenriched and salt-enriched samples by examining the 16S rRNA gene and the chlorophyll A oxygenase (CAO) gene. Investigation of the secondary metabolites by HPLC, LCMS/MS, and NMR showed the presence of ulithiacyclamide along with patellamide E in the unenriched symbiotic cells. Interestingly, ulithiacyclamide was detected in the enriched cells from the same Ascidian specimen. Molecular docking showed the high binding affinity of both compounds to estrogen receptor beta (ER-β) protein. This is a subtype of the nuclear receptor superfamily expressed abnormally in ovarian cancer cells. Furthermore, we isolated a patE gene variant encoding for a precursor with patellamide E (VTVCITFC) and ulithiacyclamide (CTLCCTLC) core peptides from the enriched cells. This represents a new core peptide combination. The outcome of this work provides a basis for producing useful cyclic peptides, in sustainable way, through symbiont cultivation. This could become a platform for bioengineering to generate diverse compound analogues.

Single symbiotic cell transcriptome sequencing of coral

Zooxanthellae and coral can form an intracellular symbiotic system. Yet, little is known about the molecular mechanism underlying this symbiosis. In this study, we characterized the symbiosis based on analyses of gene expression at the single-cell level. Among 9110 single coral cells, we identified 4871 symbiotic cells based on the detection of both coral and zooxanthellae gene transcripts within a single cell. Using the bioinformatics tool Seurat, symbiotic cells were further clustered into five groups, 52 genes exhibited differential expression between groups. We proposed an index called the “symbiosis index”, to indicate the degree of gene expression of both species in a single symbiotic cell. Interestingly, the index differed distinctly among the five groups. The symbiosis index was highly correlated with the expression of the coral gene gfas1.m1.6761 (ANKRD40), which encodes ankyrin repeat domain-containing protein 40 and is involved in DNA replication (r = 0.76). Two metabolism-related genes, DAGLA and betaGlu, were highly expressed in cells with a high symbiosis index. Four zooxanthellae genes, PRPF19, ATRN, aAA-ATPases and AK812-SmicGene44833, exhibited substantial changes in expression levels when zooxanthellae lived within coral. A trajectory analysis suggested that cells with a higher symbiosis index may be derived from those with a lower index during coral colony development. Taken together, our results provide evidence for zooxanthellae residing within coral, forming a symbiotic system. The symbiosis index is an effective indicator of different cell groups, with lineage relationships among groups. Additionally, we identified specific genes that exhibit expression changes in the symbiotic system.

Blkn, a novel Medicago truncatula mutant achieving black nodule phenotype

blkn is a Medicago truncatula mutant that is achieving null function-black nodule phenotype. blkn is a Tnt1-retrotransposon mutant, Tnt1 is Nicotiana tabacum retro-transposon which is replicated via RNA copy and integrated in plant genome. Interestingly, blkn exhibited double contents of phenolic compounds comparing to R108 wild type. The mutated black nodule is displaying cells abnormality in both infection and nitrogen fixation zones. Transverse section of blkn nodule doesn’t display clearly characteristic shape like the control and the symbiotic cells don't totally filled with bacteroids along with high lignification at the cell wall periphery. Our goal was blkn mutant; phenotype, physiological, and molecular characterizations. AFLP-based PCR method was used to detect the mutated gene(s) in this mutant line. About 25 Tnt1-tagged fragments ranging from ~100 to ~500 bp were isolated, sequenced and submitted to Genbank. The Tnt1 insertion was precisely located next to the base number 303 post ATG start codon of M. truncatula L-type lectin-domain receptor kinase VII.2 gene encodes Lectin_LegB Receptor Like Kinase (MtLectinRLK). MtLectinRLK contains Lectin_legB domain, two transmembrane helix (TMhilex) and an extracellular Receptor Protein kinase (Pkinase). MtLectinRLK is an ancestry related to probable L-type lectin-domain containing receptor kinase Cicer arietinum, Trifolium pretense, Phaseolus vulgaris, Vigna radiate and Glycine soja.

Unexplored Arsenals of Legume Peptides With Potential for Their Applications in Medicine and Agriculture.

During endosymbiosis, bacteria live intracellularly in the symbiotic organ of their host. The host controls the proliferation of endosymbionts and prevents their spread to other tissues and organs. In Rhizobium-legume symbiosis the major host effectors are secreted nodule-specific cysteine-rich (NCR) peptides, produced exclusively in the symbiotic cells. NCRs have evolved in the Inverted Repeat Lacking Clade (IRLC) of the Leguminosae family. They are secreted peptides that mediate terminal differentiation of the endosymbionts, forming polyploid, non-cultivable cells with increased membrane permeability. NCRs form an extremely large family of peptides, which have four or six conserved cysteines but otherwise highly diverse amino acid sequences, resulting in a wide variety of anionic, neutral and cationic peptides. In vitro, many synthetic NCRs have strong antimicrobial activities against both Gram-negative and Gram-positive bacteria, including the ESKAPE strains and pathogenic fungi. The spectra and minimal bactericidal and anti-fungal concentrations of NCRs differ, indicating that, in addition to their charge, the amino acid composition and sequence also play important roles in their antimicrobial activity. NCRs attack the bacteria and fungi at the cell envelope and membrane as well as intracellularly, forming interactions with multiple essential cellular machineries. NCR-like peptides with similar symbiotic functions as the NCRs also exist in other branches of the Leguminosae family. Thus, legumes provide countless and so far unexplored sources of symbiotic peptides representing an enormous resource of pharmacologically interesting molecules.

A New Method for Collecting Large Amounts of Symbiotic Gastrodermal Cells from Octocorals.

The study of cnidarian-dinoflagellate endosymbiosis in octocorals is becoming increasingly important. As symbiotic gastrodermal cells (SGCs) are the key cells in a symbiotic relationship, obtaining SGCs and studying their functions represent an urgent need. The majority of the cells dissociated from octocoral tissues consist of host cells and algal cells, and very few intact SGCs can be observed. To solve this problem, we developed a new method to collect large amounts of SGCs from octocorals. We incubated the tissue of Sinularia flexibilis in high-salinity (60‰) filtered seawater for 6 h and were able to collect more than 18 times the number of SGCs from the control group. To test the quality of the dissociated cells, we performed three assays to evaluate their cell viability. All three assays demonstrated that cell viability was good after incubating in a high-salinity solution. We also used two other octocorals, Paralemnalia thyrsoides and Sinularia compressa, to perform the same experiment, and the results were similar to those for Sinularia flexibilis. Therefore, a high-salinity-induced increase in the SGC ratio is a common phenomenon among octocorals. This method allows researchers to collect large amounts of SGCs from octocorals and helps us to better understand the complex molecular interactions in cnidarian-dinoflagellate endosymbiosis.

Stages of the symbiotic zooxanthellae–host cell division and the dynamic role of coral nucleus in the partitioning process: a novel observation elucidated by electron microscopy

The relationship of the symbiotic algae or commonly referred as zooxanthellae (zo) and its host invertebrate species is uniquely observed among cnidarians where these tiny single-celled endosymbionts are used to reside densely within the gastrodermal tissue. However, at the ultrastructural level, little has been known on how zo are maintained through proliferation within its host cells. In the present study, the stages of division of the zo that occurred in the coral cells were investigated by electron microscopy. Scleractinian Pocillopora damicornis was fixed by freeze-substitution methods using a high-pressure freezer and embedded in resin. Ultrathin sections were observed using transmission electron microscope. Symbiotic cell at interphase showed the cortical fine structures of the non-dividing zo enclosed by the outermost plasma membrane (zpm). Overlying them, two distinct layers of intermediate membranes (inm) and a symbiosome membrane (sm) were found. When cytokinesis occurred, the zpm at the opposite division regions formed ‘U-shaped’ furrows which led amphiesmal vesicle (av) to protrude inwardly. As they invaginates, new avs and series of cortical microtubules (mi) are used to develop concomitant with the symbiont’ nuclear division. After the cytokinesis, the overlying membranes successively invaginate; thus, the daughter zooxanthellae were transformed into coccoid cells. These cells, however, remain contiguous to each other until when the coral nucleus (cnu) migrated toward one of the cleavage sites to intervene and dissociate them. Upon occupying the interzooxanthellar space, host nuclear division occurred prior to cytokinesis. These events involving the role of cnu in the partitioning process are first reported in this study, in which its dynamic intervening behavior had been elucidated as a novel finding from the cnidarian species.

Potent Chimeric Antimicrobial Derivatives of the Medicago truncatula NCR247 Symbiotic Peptide.

In Rhizobium-legume symbiosis, the bacteria are converted into nitrogen-fixing bacteroids. In many legume species, differentiation of the endosymbiotic bacteria is irreversible, culminating in definitive loss of their cell division ability. This terminal differentiation is mediated by plant peptides produced in the symbiotic cells. In Medicago truncatula more than ∼700 nodule-specific cysteine-rich (NCR) peptides are involved in this process. We have shown previously that NCR247 and NCR335 have strong antimicrobial activity on various pathogenic bacteria and identified interaction of NCR247 with many bacterial proteins, including FtsZ and several ribosomal proteins, which prevent bacterial cell division and protein synthesis. In this study we designed and synthetized various derivatives of NCR247, including shorter fragments and various chimeric derivatives. The antimicrobial activity of these peptides was tested on the ESKAPE bacteria; Enterococcus faecalis, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Escherichia coli as a member of Enterobacteriaceae and in addition Listeria monocytogenes and Salmonella enterica. The 12 amino acid long C-terminal half of NCR247, NCR247C partially retained the antimicrobial activity and preserved the multitarget interactions with partners of NCR247. Nevertheless NCR247C became ineffective on S. aureus, P. aeruginosa, and L. monocytogenes. The chimeric derivatives obtained by fusion of NCR247C with other peptide fragments and particularly with a truncated mastoparan sequence significantly increased bactericidal activity and altered the antimicrobial spectrum. The minimal bactericidal concentration of the most potent derivatives was 1.6 μM, which is remarkably lower than that of most classical antibiotics. The killing activity of the NCR247-based chimeric peptides was practically instant. Importantly, these peptides had no hemolytic activity or cytotoxicity on human cells. The properties of these NCR derivatives make them promising antimicrobials for clinical use.

Gene Expression in Nitrogen-Fixing Symbiotic Nodule Cells in Medicago truncatula and Other Nodulating Plants.

Root nodules formed by plants of the nitrogen-fixing clade (NFC) are symbiotic organs that function in the maintenance and metabolic integration of large populations of nitrogen-fixing bacteria. These organs feature unique characteristics and processes, including their tissue organization, the presence of specific infection structures called infection threads, endocytotic uptake of bacteria, symbiotic cells carrying thousands of intracellular bacteria without signs of immune responses, and the integration of symbiont and host metabolism. The early stages of nodulation are governed by a few well-defined functions, which together constitute the common symbiosis-signaling pathway (CSSP). The CSSP activates a set of transcription factors (TFs) that orchestrate nodule organogenesis and infection. The later stages of nodule development require the activation of hundreds to thousands of genes, mostly expressed in symbiotic cells. Many of these genes are only active in symbiotic cells, reflecting the unique nature of nodules as plant structures. Although how the nodule-specific transcriptome is activated and connected to early CSSP-signaling is poorly understood, candidate TFs have been identified using transcriptomic approaches, and the importance of epigenetic and chromatin-based regulation has been demonstrated. We discuss how gene regulation analyses have advanced our understanding of nodule organogenesis, the functioning of symbiotic cells, and the evolution of symbiosis in the NFC.

Overexpression of the Chromosome Partitioning Gene parA in Azorhizobium caulinodans ORS571 Alters the Bacteroid Morphotype in Sesbania rostrata Stem Nodules.

Azorhizobium caulinodans ORS571 is a diazotroph that forms N2-fixing nodules on the roots and stems of the tropical legume Sesbania rostrata. Deletion of the parA gene of this bacterium results in cell cycle defects, pleiomorphic cell shape, and formation of immature stem nodules on its host plant. In this study, we constructed a parA overexpression mutant (PnptII-parA) to complement a previous study and provide new insights into bacteroid formation. We found that overproduction of ParA did not affect growth, cell morphology, chromosome partitioning, or vegetative nitrogen fixation in the free-living state. Under symbiosis, however, distinctive features, such as a single swollen bacteroid in one symbiosome, relatively narrow symbiosome space, and polyploid cells were observed. The morphotype of the PnptII-parA bacteroid is reminiscent of terminal differentiation in some IRLC indeterminate nodules, but S. rostrata is not thought to produce the NCR peptides that induce terminal differentiation in rhizobia. In addition, the transcript patterns of many symbiosis-related genes elicited by PnptII-parA were different from those elicited by the wild type. Accordingly, we propose that the particular symbiosome formation in PnptII-parA stem-nodules is due to cell cycle disruption caused by excess ParA protein in the symbiotic cells during nodulation.

Repeated evolution of bacteriocytes in lygaeoid stinkbugs.

Host-microbe symbioses often evolved highly complex developmental processes and colonization mechanisms for establishment of stable associations. It has long been recognized that many insects harbour beneficial bacteria inside specific symbiotic cells (bacteriocytes) or organs (bacteriomes). However, the evolutionary origin and mechanisms underlying bacterial colonization in bacteriocyte/bacteriome formation have been poorly understood. In order to uncover the origin of such evolutionary novelties, we studied the development of symbiotic organs in five stinkbug species representing the superfamily Lygaeoidea in which diverse bacteriocyte/bacteriome systems have evolved. We tracked the symbiont movement within the eggs during the embryonic development and determined crucial stages at which symbiont infection and bacteriocyte formation occur, using whole-mount fluorescence in situ hybridization. In summary, three distinct developmental patterns were observed: two different modes of symbiont transfer from initial symbiont cluster (symbiont ball) to presumptive bacteriocytes in the embryonic abdomen, and direct incorporation of the symbiont ball without translocation of bacterial cells. Across the host taxa, only closely related species seemed to have evolved relatively conserved types of bacteriome development, suggesting repeated evolution of host symbiotic cells and organs from multiple independent origins.

The Ca2+-ATPase in Legume Root Nodule Peribacteroid Membrane as a Potential Key Determinant of Ca-Dependent Regulation of Symbiosome Functioning

Certain aspects of Ca-dependent regulation of symbiosomes functioning mediated by the operation of Ca2+-translocating ATPase on the symbiosome membrane (SM) were considered. As follows from the results recently obtained by us, the mechanism underlying the action of this calcium pump exhibits a certain functional feature caused by that transfer by it of calcium ion through the SM is closely coupled with transmembrane translocation through the same membrane but in opposite direction of H+ ions. The data here outlined show that such the cation-exchanged mechanism of the Ca2+-ATPase operation can putatively lead to corresponding modulation or maintenance of both ionic and redox homeostasis of the symbiosome space (SS) around bacteroids at a proper level and thereby to the regulation of symbiosomes functioning. It is suggested that based on the same mechanism inherent function of the enzyme in question is associated with its involvement in symbiotic partner cells signaling as well. All these potential activities of the Ca2+-ATPase lead to the important question whether it is the factor determining substantial functions of symbiosomes. Although it is evident that research an answer to this question is a complex enough experimental problem, its solution may give significant contribution to elucidating mechanisms of Ca-dependent regulation of the processes occurring in mature symbiosomes.