Country：Japan

Country：Japan

Country：Japan

Publishing type：Research paper (scientific journal)  

<p>From the 992 samples of culture extracts of microorganisms isolated from soil in Japan, we found that the extract of <i>Streptomyces</i> sp. no. 226 inhibited <i>Orobanche minor</i> seed germination without significantly affecting the seed germination of <i>Trifolium pratense</i> and the growth of <i>Aspergillus oryzae</i> and <i>Escherichia coli</i>. Using ESI-MS, ¹H-NMR, and ¹³C-NMR, we identified the active compound as cycloheximide. Cycloheximide had half-maximum inhibitory concentrations of 2.6 ng/mL for the inhibition of seed germination of <i>O. minor</i> and 2.5 µg/mL for that of the conidial germination of <i>A</i>. <i>oryzae</i>. Since cycloheximide is known to inhibit translation by interacting with ribosomal protein L28 (RPL28) in yeast, we investigated whether RPL protein of <i>O. minor</i> plays a critical role in the inhibition of <i>O. minor</i> seed germination. Our data suggested that <i>O. minor</i> RPL27A was not sensitive to cycloheximide by comparing it to the strain expressing <i>S. cerevisiae</i> RPL28. These findings suggest the presence of an unidentified mechanism by which cycloheximide hinders <i>O. minor</i> seed germination.</p>

DOI： 10.1584/jpestics.d23-038

PubMed

Publishing type：Research paper (scientific journal)  

The Ralstonia solanacearum species complex is a group of globally important plant pathogens. Bacteria in this very large and genetically diverse group all colonize the xylem elements of angiosperm plants and cause high-impact wilting diseases of many crops. Because they threaten economic and food security, several R. solanacearum species complex subgroups are strictly regulated as quarantine pests. Biologically meaningful and consistent nomenclature is essential for organisms that have major economic and regulatory importance, such as plant-pathogenic Ralstonia. There are currently three species of Ralstonia wilt pathogens: R. pseudosolanacearum (corresponding to two phylogenetic groups that are described in the literature as phylotypes I and III), R. solanacearum (phylotypes IIA, IIB, and IIC), and R. syzygii (phylotype IV, containing three subspecies: subsp. syzygii, subsp. celebensis, and subsp. indonesiensis). A recent paper proposed reclassifying phylotype I as a new species named “ Ralstonia nicotianae.” The purpose of this commentary is to register our objection to the taxon “ Ralstonia nicotianae.”

[Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY 4.0 International license .

DOI： 10.1094/phytofr-06-23-0071-le

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DOI： 10.1021/acs.joc.3c01789

PubMed

Publishing type：Research paper (scientific journal)  

DOI： 10.1080/10715762.2023.2277144

PubMed

Authorship：Lead author,　Last author,　Corresponding author   Publishing type：Research paper (scientific journal)  

Ralstonia solanacearum species complex (RSSC) strains are devastating plant pathogens distributed worldwide. The primary cell density–dependent gene expression system in RSSC strains is phc quorum sensing (QS). It regulates the expression of about 30% of all genes, including those related to cellular activity, primary and secondary metabolism, pathogenicity, and more. The phc regulatory elements encoded by the phcBSRQ operon and phcA gene play vital roles. RSSC strains use methyl 3-hydroxymyristate (3-OH MAME) or methyl 3-hydroxypalmitate (3-OH PAME) as the QS signal. Each type of RSSC strain has specificity in generating and receiving its QS signal, but their signaling pathways might not differ significantly. In this review, I describe the genetic and biochemical factors involved in QS signal input and the regulatory network and summarize control of the phc QS system, new cell–cell communications, and QS-dependent interactions with soil fungi.

DOI： 10.1146/annurev-micro-032521-030537

PubMed

Authorship：Last author,　Corresponding author   Publishing type：Research paper (scientific journal)  

Ralstonia solanacearum species complex (RSSC) strains infect both plants and fungi. The phc quorum-sensing (QS) system of RSSC is important for parasitism on plants, because it allows them to invade and proliferate within the hosts by causing appropriate activation of the system at each infection step.

DOI： 10.1128/spectrum.00036-23

PubMed

Publishing type：Research paper (scientific journal)  

Abstract

The gram‐negative plant‐pathogenic β‐proteobacterium Ralstonia pseudosolanacearum strain OE1‐1 produces methyl 3‐hydroxymyristate as a quorum sensing (QS) signal through methyltransferase PhcB and senses the chemical via the sensor histidine kinase PhcS. This leads to activation of the LysR family transcription regulator PhcA, which regulates the genes (QS‐dependent genes) responsible for QS‐dependent phenotypes, including virulence. The transcription regulator ChpA, which possesses a response regulator receiver domain and also a hybrid sensor histidine kinase/response regulator phosphore‐acceptor domain but lacks a DNA‐binding domain, is reportedly involved in QS‐dependent biofilm formation and virulence of R. pseudosolanacearum strain GMI1000. To explore the function of ChpA in QS of OE1‐1, we generated a chpA‐deletion mutant (ΔchpA) and revealed that the chpA deletion leads to significantly altered QS‐dependent phenotypes. Furthermore, ΔchpA exhibited a loss in its infectivity in xylem vessels of tomato plant roots, losing virulence on tomato plants, similar to the phcA‐deletion mutant (ΔphcA). Transcriptome analysis showed that the transcript levels of phcB, phcQ, phcR, and phcA in ΔchpA were comparable to those in OE1‐1. However, the transcript levels of 89.9% and 88.9% of positively and negatively QS‐dependent genes, respectively, were significantly altered in ΔchpA compared with OE1‐1. Furthermore, the transcript levels of these genes in ΔchpA were positively correlated with those in ΔphcA. Together, our results suggest that ChpA is involved in the regulation of these QS‐dependent genes, thereby contributing to the behaviour in host plant roots and virulence of OE1‐1.

DOI： 10.1111/mpp.13374

PubMed

Publishing type：Research paper (scientific journal)  

DOI： 10.1016/j.pmpp.2023.101995

Publishing type：Research paper (scientific journal)  

DOI： 10.1124/molpharm.122.000601

PubMed

Authorship：Last author,　Corresponding author   Publishing type：Research paper (scientific journal)  

Ralstonia solanacearum species complex (RSSC) strains are plant pathogens that produce lipopeptides (ralstonins and ralstoamides) by the polyketide synthase-nonribosomal peptide synthetase (PKS-NRPS) enzyme hybrid. Recently, ralstonins were found to be key molecules in the parasitism of RSSC to other hosts, Aspergillus and Fusarium fungi. The PKS-NRPS genes of RSSC strains in the GenBank database suggest the production of additional lipopeptides, although it has not been confirmed to date. Here, we report the genome-driven and mass-spectrometry-guided discovery, isolation, and structural elucidation of ralstopeptins A and B from strain MAFF 211519. Ralstopeptins were found to be cyclic lipopeptides with two amino acid residues less than ralstonins. The partial deletion of the gene encoding PKS-NRPS obliterated the production of ralstopeptins in MAFF 211519. Bioinformatic analyses suggested possible evolutionary events of the biosynthetic genes of RSSC lipopeptides, where intragenomic recombination may have occurred within the PKS-NRPS genes, reducing the gene size. The chlamydospore-inducing activities of ralstopeptins A and B, ralstonins A and B, and ralstoamide A in the fungus Fusarium oxysporum indicated a structural preference for ralstonins. Altogether, we propose a model for the evolutionary processes that contribute to the chemical diversity of RSSC lipopeptides and its relation to the endoparasitism of RSSC in fungi.

DOI： 10.1021/acschembio.2c00907

PubMed

Publishing type：Research paper (scientific journal)  

Abstract

After infecting roots of tomato plants, the gram‐negative bacterium Ralstonia pseudosolanacearum strain OE1‐1 activates quorum sensing (QS) to induce production of plant cell wall‐degrading enzymes, such as β‐1,4‐endoglucanase (Egl) and β‐1,4‐cellobiohydrolase (CbhA), via the LysR family transcriptional regulator PhcA and then invades xylem vessels to exhibit virulence. The phcA‐deletion mutant (ΔphcA) exhibits neither the ability to infect xylem vessels nor virulence. Compared with strain OE1‐1, the egl‐deletion mutant (Δegl) exhibits lower cellulose degradation activity, lower infectivity in xylem vessels, and reduced virulence. In this study, we analysed functions of CbhA other than cell wall degradation activity that are involved in the virulence of strain OE1‐1. The cbhA‐deletion mutant (ΔcbhA) lacked the ability to infect xylem vessels and displayed loss of virulence, similar to ΔphcA, but exhibited less reduced cellulose degradation activity compared with Δegl. Transcriptome analysis revealed that the phcA expression levels in ΔcbhA were significantly lower than in OE1‐1, with significantly altered expression of more than 50% of PhcA‐regulated genes. Deletion of cbhA led to a significant change in QS‐dependent phenotypes, similar to the effects of phcA deletion. Complementation of ΔcbhA with native cbhA or transformation of this mutant with phcA controlled by a constitutive promoter recovered its QS‐dependent phenotypes. The expression level of phcA in ΔcbhA‐inoculated tomato plants was significantly lower than in strain OE1‐1‐inoculated plants. Our results collectively suggest that CbhA is involved in the full expression of phcA, thereby contributing to the QS feedback loop and virulence of strain OE1‐1.

DOI： 10.1111/mpp.13322

PubMed

Other URL： https://onlinelibrary.wiley.com/doi/full-xml/10.1111/mpp.13322

Publishing type：Research paper (scientific journal)  

DOI： 10.1021/acs.jafc.2c06791

PubMed

Publishing type：Research paper (scientific journal)  

DOI： 10.1007/s10265-022-01427-3

PubMed

Other URL： https://link.springer.com/article/10.1007/s10265-022-01427-3/fulltext.html

DOI： 10.1584/jpestics.w22-29

Publishing type：Research paper (scientific journal)  

DOI： 10.1016/j.pestbp.2022.105074

Publishing type：Research paper (scientific journal)  

DOI： 10.1111/mpp.13124

Other URL： https://onlinelibrary.wiley.com/doi/full-xml/10.1111/mpp.13124

Publishing type：Research paper (scientific journal)  

DOI： 10.1080/10715762.2021.1937620

PubMed

Authorship：Last author,　Corresponding author   Publishing type：Research paper (scientific journal)  

DOI： 10.1021/acschembio.1c00276

PubMed

Authorship：Last author,　Corresponding author   Publishing type：Research paper (scientific journal)  

First, we revisited the reported NMR data of bradyoxetin, a putative cell density factor of Bradyrhizobium japonicum, and found some inconsistencies in the proposed structure. To elucidate the correct structure, we synthesized model oxetane compounds and confirmed that the NMR data of the synthetic compounds did not match those of the reported bradyoxetin. After reinterpreting the reported NMR data, we concluded that bradyoxetin must be chloramphenicol. Next, some derivatives of 2-hydroxy-4-((methylamino)(phenyl)methyl)cyclopentanone (HMCP), which is a putative quorum-sensing molecule of Ralstonia solanacearum, were synthesized. The NMR spectra of the synthesized compounds were completely different from those of the reported natural products. Based on theoretical studies, including the estimation of 1H and 13C NMR chemical shifts using density functional theory calculations, we confirmed the correctness of the structure of the synthesized compound. These results strongly suggest that the proposed structure of HMCP could be incorrect.

DOI： 10.1021/acs.jnatprod.0c01369

PubMed

Publishing type：Research paper (scientific journal)  

The soil-borne, plant-pathogenic Ralstonia solanacearum strain OE1-1 produces and secretes methyl 3-hydroxymyristate (3-OH MAME) as a quorum sensing (QS) signal, which contributes to its virulence. A global virulence regulator, PhcA, functioning through the QS system, positively regulates the expression of ralA, which encodes furanone synthase, to produce aryl-furanone secondary metabolites, ralfuranones. A ralfuranone-deficient mutant (ΔralA) is weakly virulent when directly inoculated into tomato xylem vessels. To investigate the functions of ralfuranones, we analysed R. solanacearum transcriptome data generated by RNA sequencing technology. ΔralA expressed phcB, which is associated with 3-OH MAME production, and phcA at levels similar to those in strain OE1-1. In addition, ΔralA exhibited down-regulated expression of more than 90% of the QS positively regulated genes, and up-regulated expression of more than 75% of the QS negatively regulated genes. These results suggest that ralfuranones affect the QS feedback loop. Ralfuranone supplementation restored the ability of ΔralA cells to aggregate. In addition, ralfuranones A and B restored the swimming motility of ΔralA to wild-type levels. However, the application of exogenous ralfuranones did not affect the production of the major exopolysaccharide, EPS I, in ΔralA. Quantitative real-time polymerase chain reaction assays revealed that the deletion of ralA results in the down-regulated expression of vsrAD and vsrBC, which encode a sensor kinase and a response regulator, respectively, in the two-component regulatory systems that influence EPS I production. The application of ralfuranone B restored the expression of these two genes. Overall, our findings indicate that integrated signalling via ralfuranones influences the QS and virulence of R. solanacearum.

DOI： 10.1111/mpp.12537

PubMed

DOI： 10.1038/s41598-020-64687-w

Publishing type：Research paper (scientific journal)  

A gram-negative plant-pathogenic bacterium Ralstonia solanacearum strain OE1-1 produces and extracellularly secretes methyl 3-hydroxymyristate (3-OH MAME), and senses the chemical as a quorum-sensing (QS) signal, activating QS. During QS a functional global transcriptional regulator PhcA, through the 3-OH MAME-dependent two-component system, induces the production of virulence factors including a major extracellular polysaccharide EPS I and ralfuranone. To elucidate the mechanisms of phcA regulation underlying the QS system, among Tn5-mutants from the strain OE1-1, we identified a mutant of RSc1351 gene (phcK), encoding a putative sensor histidine kinase, that exhibited significantly decreased QS-dependent cell aggregation. We generated a phcK-deletion mutant (ΔphcK) that produced significantly less EPS I and ralfuranone than the wild-type strain OE1-1. Quantitative reverse transcription PCR assays showed that the phcA expression level was significantly down-regulated in the ΔphcK mutant but not in other QS mutants. The transcriptome data generated with RNA sequencing technology revealed that the expression levels of 88.2% of the PhcA-positively regulated genes were down-regulated in the ΔphcK mutant, whereas the expression levels of 85.9% of the PhcA-negatively regulated genes were up-regulated. Additionally, the native phcK-expressing complemented ΔphcK strain and the ΔphcK mutant transformed with phcA controlled by a constitutive promoter recovered their cell aggregation phenotypes. Considered together, the results of this study indicate that phcK is required for full phcA expression, thereby driving the QS circuit of R. solanacearum strain OE1-1. This is the first report of the phcA transcriptional regulation of R. solanacearum.

DOI： 10.1111/mpp.12998

PubMed

Publishing type：Research paper (scientific journal)  

To characterize the molecular mechanisms underlying life-stage transitions in Phytophthora infestans, we initiated a chemical genetics approach by screening for a stage-specific inhibitor of morphological development from microbial culture extracts prepared mostly from actinomycetes from soil in Japan. Of the more than 700 extracts, one consistently inhibited Ph. infestans cyst germination. Purification and identification of the active compound by ESI–MS, 1H-NMR, and 13C-NMR identified β-rubromycin as the inhibitor of cyst germination (IC50 = 19.8 μg/L); β-rubromycin did not inhibit growth on rye media, sporangium formation, zoospore release, cyst formation, or appressorium formation in Ph. infestans. Further analyses revealed that β-rubromycin inhibited the germination of cysts and oospores in Pythium aphanidermatum. A chemical genetic approach revealed that β-rubromycin stimulated the expression of RIO kinase-like gene (PITG_04584) by 60-fold in Ph. infestans. Genetic analyses revealed that PITG_04584, which lacks close non-oomycete relatives, was involved in zoosporogenesis, cyst germination, and appressorium formation in Ph. infestans. These data imply that further functional analyses of PITG_04584 may contribute to new methods to suppress diseases caused by oomycetes.

DOI： 10.1038/s41598-020-79326-7

PubMed

Authorship：Last author,　Corresponding author   Publishing type：Research paper (scientific journal)  

Strains of Ralstonia solanacearum species complex (RSSC) are devastating plant pathogens distributed globally with a wide host range and genetic diversity. Many RSSC strains harbor the polyketide synthase-nonribosomal peptide synthetase (PKS-NRPS) hybrid gene rmyA/rmyB for ralstonin production. We report that ralstoamides A (1) and B (2), which are ralstonin-like but shorter lipopeptides, were discovered from the Japanese strains using accumulated RSSC genome data and LC/MS-based metabolite analysis. Their structures, including absolute configurations, were elucidated by spectroscopic analysis and chemical techniques. ramA, a PKS-NRPS gene for ralstoamide production, was identified from the producer strains by genome sequencing and gene-deletion experiments. Based on the analysis of biosynthetic genes of ralstoamides and ralstonins, we suggest the occurrence of NRPS-module reduction of rmyA/rmyB genes in some RSSC strains. This possible molecular evolution changed not only the structures, but also the biological activity of RSSC lipopeptides.

DOI： 10.1021/acschembio.0c00605

PubMed

Authorship：Last author,　Corresponding author   Publishing type：Research paper (scientific journal)  

Strains of Ralstonia solanacearum species complex (RSSC) cause “bacterial wilt” on a wide range of plant species and thus lead to marked economic losses in agriculture. Quorum sensing (QS), a bacterial cell-cell communication mechanism, controls the virulence of RSSC strains by regulating the production of extracellular polysaccharide (EPS) and secondary metabolites, biofilm formation, and cellular motility. R. solanacearum strain OE1-1 employs (R)-methyl 3-hydroxymyristate (3-OH MAME) as a QS signal, which is synthesized by the PhcB methyltransferase and sensed by the PhcS/ PhcRQ two-component system. We describe the design, synthesis, and biological evaluation of inhibitors of the phc QS system. Initial screening of a small set of QS signal analogues revealed that methyl 3-hydroxy-8-phenyl-octanoate, named, PQI-1 (phc quorum sensing inhibitor-1), inhibited biofilm formation by strain OE1-1. To improve its inhibitory activity, the derivatives of PQI-1 were synthesized, and their QS inhibition activities were evaluated. PQIs-2-5 evolved from PQI-1 more strongly inhibited not only biofilm formation but also the production of ralfuranone and EPS. Furthermore, RNA-Seq analysis revealed that the PQIs effectively inhibited QS-dependent gene expression and repression in strain OE1-1. On the other hand, the PQIs did not affect the canonical QS systems of the representative reporter bacteria. These antagonists, especially PQI-5, reduced wilting symptoms of the tomato plants infected with strain OE1-1. Taken together, we suggest that targeting the phc QS system has potential for the development of chemicals that protect agricultural crops from bacterial wilt disease.

DOI： 10.1021/acschembio.0c00752

PubMed

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Cupriavidus taiwanensis LMG19424, a β-rhizobial symbiont of Mimosa pudica, harbors phc and tqs quorum sensing (QS), which are the homologous cell-cell communication systems previously identified from the plant pathogen Ralstonia solanacearum and the human pathogen Vibrio cholerae, respectively. However, there has been no experimental evidence reported that these QS systems function in C. taiwanensis LMG19424. We identified (R)-methyl 3-hydroxymyristate (3-OH MAME) and (S)-3-hydroxypentadecan-4-one (C15-AHK) as phc and tqs QS signals, respectively, and characterized these QS systems. The expression of the signal synthase gene phcB and tqsA in E. coli BL21(DE3) resulted in the high production of 3-OH MAME and C15-AHK, respectively. Their structures were elucidated by comparison of EI-MS data and GC/chiral LC retention times with synthetic standards. The deletion of phcB reduced cell motility and increased biofilm formation, and the double deletion of phcB/tqsA caused the accumulation of the metal chelator coproporphyrin III in its mutant culture. Although the deletion of phcB and tqsA slightly reduced its ability to nodulate on aseptically grown seedlings of M. pudica, there was no significant difference in nodule formation between LMG19424 and its QS mutants when commercial soils were used. Taken together, this is the first example of the simultaneous production of 3-OH MAME/C15-AHK as QS signals in a bacterial species, and the importance of the phc/tqs QS systems in the saprophytic stage of C. taiwanensis LMG19424 is suggested.

DOI： 10.1021/acs.jnatprod.0c00054

PubMed

Publishing type：Research paper (scientific journal)  

Skeletal muscle atrophy causes decreased physical activity and increased risk of metabolic diseases. We investigated the effects of oleamide (cis-9,10-octadecanamide) treatment on skeletal muscle health. The plasma concentration of endogenous oleamide was approximately 30 nM in the male ddY mice under normal physiological conditions. When the stable isotope-labelled oleamide was orally administered to male ddY mice (50 mg/kg), the plasma concentration of exogenous oleamide reached approximately 170 nM after 1 h. Male ddY mice were housed in small cages (one-sixth of normal size) to enforce sedentary behaviour and orally administered oleamide (50 mg/kg/day) for 4 weeks. Housing in small cages decreased tibialis anterior (TA) muscle mass and the cross-sectional area (CSA) of the myofibres in TA muscle. Dietary oleamide alleviated the decreases in TA muscle and resulted in plasma oleamide concentration of approximately 120 nM in mice housed in small cages. Housing in small cages had no influence on the phosphorylation levels of Akt, mTOR, and p70S6K in TA muscle; nevertheless, oleamide increased the phosphorylation levels of the proteins. Housing in small cages increased the expression of LC3-II and p62, but not LC3-I, in TA muscle, and oleamide reduced LC3-I, LC3-II, and p62 expression levels. In C2C12 myotubes, oleamide increased myotube diameter at ≥ 100 nM. Furthermore, the mTOR inhibitor, Torin 1, suppressed oleamide-induced increases in myotube diameter and protein synthesis. These results indicate that dietary oleamide rescued TA muscle atrophy in mice housed in small cages, possibly by activating the PI3K/Akt/mTOR signalling pathway and restoring autophagy flux.

DOI： 10.1017/S0007114520004304

PubMed

Authorship：Last author,　Corresponding author   Publishing type：Research paper (scientific journal)  

Pyoverdines, a group of peptide siderophores produced by Pseudomonas species, function not only in iron acquisition, but also in their virulence in hosts. Thus, chemical inhibition of pyoverdine production may be an effective strategy to control Pseudomonas virulence. In the plant pathogen Pseudomonas cichorii SPC9018 (SPC9018), pyoverdine production is required for virulence on eggplant. We screened microbial culture extracts in a pyoverdine-production inhibition assay of SPC9018 and found Streptomyces sp. RM-32 as a candidate-producer. We isolated two active compounds from RM-32 cultures, and elucidated their structures to be actinomycins X2 and D. Actinomycins X2 and D inhibited pyoverdine production by SPC9018 with IC50 values of 17.6 and 29.6 μM, respectively. Furthermore, pyoverdine production in other Pseudomonas bacteria, such as the mushroom pathogen P. tolaasii, was inhibited by the actinomycins. Therefore, these actinomycins may be useful as chemical tools to examine pyoverdine functions and as seed compounds for anti-Pseudomonas virulence agents.

DOI： 10.1080/09168451.2020.1785839

PubMed

Publishing type：Research paper (scientific journal)  

The Gram-negative soil-borne bacterium Ralstonia solanacearum first infects roots of host plants and then invades xylem vessels. In xylem vessels, the bacteria grow vigorously and produce exopolysaccharides (EPSs) to cause a wilt symptom on host plants. The EPSs are thus the main virulence factors of R. solanacearum. The strain OE1-1 of R. solanacearum produces methyl 3-hydroxymyristate as a quorum-sensing (QS) signal, and senses this QS signal, activating QS. The QS-activated LysR-type transcriptional regulator PhcA induces the production of virulence-related metabolites including ralfuranone and the major EPS, EPS I. To elucidate the function of EPS I, the transcriptomes of R. solanacearum strains were analysed using RNA sequencing technology. The expression of 97.2% of the positively QS-regulated genes was down-regulated in the epsB-deleted mutant ΔepsB, which lost its EPS I productivity. Furthermore, expression of 98.0% of the negatively QS-regulated genes was up-regulated in ΔepsB. The deficiency to produce EPS I led to a significantly suppressed ralfuranone productivity and significantly enhanced swimming motility, which are suppressed by QS, but did not affect the expression levels of phcA and phcB, which encode a methyltransferase required for methyl 3-hydroxymyristate production. Overall, QS-dependently produced EPS I may be associated with the feedback loop of QS.

DOI： 10.1111/mpp.12870

PubMed

Authorship：Last author,　Corresponding author   Publishing type：Research paper (scientific journal)  

Ralstonia solanacearum strains are devastating plant pathogens with global distribution, a wide host range, and genetic diversity, and they are now also referred to as the R. solanacearum species complex (RSSC). RSSC strains employ the quorum sensing (QS) system composed of the phcBSR operon to regulate their virulence on plants. The RSSC strains previously examined produce either (R)-methyl 3-hydroxymyristate (3-OH MAME) or (R)-methyl 3-hydroxypalmitate (3-OH PAME) as their QS signals. Analogously, the phylogenetic analyses of the signal synthase PhcB and the signal receptor PhcS from 15 RSSC strains revealed that these proteins have two clades dependent on their QS signal types. However, the biochemical mechanism underlying this selectivity of QS signal production remains to be elucidated. We demonstrated that the PhcB methyltransferases synthesize QS signals from the cognate fatty acids (R)-3-hydroxymyristic acid or (R)-3-hydroxypalmitic acid. The RSSC strains used here produced both fatty acids, and thus the selectivity of QS signal production depends on the activity of PhcB enzymes. On the other hand, the enantioselective supply of the precursors functioned in the production of enantiopure QS signals. The opposite QS signals weakly induced the production of virulence factors in the RSSC strains. Furthermore, the complementation of the phcB gene encoding the 3-OH PAME-type synthase to the phcB-deletion mutant of the 3-OH MAME-producing strain did not rescue its virulence on tomato plants. Taken together, we propose that the specific production of 3-OH MAME/3-OH PAME ensures full virulence of the RSSC strains.

DOI： 10.1021/acschembio.9b00553

PubMed

Kind of work：Joint Work  

Authorship：Last author,　Corresponding author   Publishing type：Research paper (scientific journal)  

Plant pathogenic bacteria possess sophisticated mechanisms to detect the presence of host plants by sensing host-derived compounds. Ralstonia solanacearum, the causative agent of bacterial wilt on solanaceous plants, employs quorum sensing to control the production of the secondary metabolite ralfuranones/ralstonins, which have been suggested to be involved in virulence. Here, we report that d-galactose and d-glucose, plant sugars, activate the production of ralfuranones/ralstonins in R. solanacearum. As a result, two new derivatives, ralfuranone M (1) and ralstonin C (2), were found in the culture extracts, and their structures were elucidated by spectroscopic and chemical methods. Ralstonin C (2) is a cyclic lipopeptide containing a unique fatty acid, (2S,3S,Z)-3-amino-2-hydroxyicos-13-enoic acid, whereas ralfuranone M (1) has a common aryl-furanone structure with other ralfuranones. d-Galactose and d-glucose activated the expression of the biosynthetic ralfuranone/ralstonin genes and in part became the biosynthetic source of ralfuranones/ralstonins. Ralfuranones and ralstonins were detected from the xylem fluid of the infected tomato plants, and their production-deficient mutants exhibited reduced virulence on tomato and tobacco plants. Taken together, these results suggest that activation of ralfuranone/ralstonin production by host sugars functions in R. solanacearum virulence.

DOI： 10.1021/acschembio.9b00301

PubMed

Publishing type：Research paper (scientific journal)  

The soil-borne bacterium Ralstonia solanacearum invades the roots and colonizes the intercellular spaces and then the xylem. The expression of lecM, encoding a lectin LecM, is induced by an OmpR family response regulator HrpG in R. solanacearum strain OE1-1. LecM contributes to the attachment of strain OE1-1 to the host cells of intercellular spaces. OE1-1 produces methyl 3-hydroxymyristate (3-OH MAME) through a methyltransferase (PhcB) and extracellularly secretes the chemical as a quorum sensing (QS) signal, which activates QS. The expression of lecM is also induced by the PhcA virulence regulator functioning through QS, and the resulting LecM is implicated in the QS-dependent production of major exopolysaccharide EPS I and the aggregation of OE1-1 cells. To investigate the function of LecM in QS, we analysed the transcriptome of R. solanacearum strains generated by RNA sequencing technology. In the lecM mutant, the expression of positively QS-regulated genes and negatively QS-regulated genes was down-regulated (by >90%) and up-regulated (by ~60%), respectively. However, phcB and phcA in the lecM mutant were expressed at levels similar to those in strain OE1-1. The lecM mutant produced significantly less ralfuranone and exhibited a significantly greater swimming motility, which were positively and negatively regulated by QS, respectively. In addition, the extracellular 3-OH MAME content of the lecM mutant was significantly lower than that of OE1-1. The application of 3-OH MAME more strongly increased EPS I production in the phcB-deleted mutant and strain OE1-1 than in the lecM mutant. Thus, the QS-dependent production of LecM contributes to the QS signalling pathway.

DOI： 10.1111/mpp.12757

PubMed

Publishing type：Research paper (scientific journal)  

Imidazole-containing dipeptides (IDPs), such as carnosine and anserine, are found exclusively in various animal tissues, especially in the skeletal muscles and nerves. IDPs have antioxidant activity because of their metal-chelating and free radical-scavenging properties. However, the underlying mechanisms that would fully explain IDP antioxidant effects remain obscure. Here, using HPLC- electrospray ionization-tandem MS analyses, we comprehensively investigated carnosine and its related small peptides in the soluble fractions of mouse tissue homogenates and ubiquitously detected 2-oxo-histidine-containing dipeptides (2-oxo-IDPs) in all examined tissues. We noted enhanced production of the 2-oxo-IDPs in the brain of a mouse model of sepsis-associated encephalopathy. Moreover, in SH-SY5Y human neuroblastoma cells stably expressing carnosine synthase, H2O2 exposure resulted in the intracellular production of 2-oxo-carnosine, which was associated with significant inhibition of the H2O2 cytotoxicity. Notably, 2-oxo-carnosine showed a better antioxidant activity than endogenous antioxidants such as GSH and ascorbate. Mechanistic studies indicated that carnosine monooxygenation is mediated through the formation of a histidyl-imidazole radical, followed by the addition of molecular oxygen. Our findings reveal that 2-oxo-IDPs are metal-catalyzed oxidation products present in vivo and provide a revised paradigm for understanding the antioxidant effects of the IDPs.

DOI： 10.1074/jbc.RA118.006111

PubMed

Authorship：Last author,　Corresponding author   Publishing type：Research paper (scientific journal)  

Microorganisms produce and secrete a variety of secondary metabolites including fatty acids, polyketides, terpenoids, alkaloids, and peptides. Among them, many molecules act as chemical signals that play important roles in inter-/intra-species microbial communication or the interaction with host organisms. In this review, I focus on our recent reports of the microbial signaling molecules involved in bacterium-fungus, bacterium-plant, and fungus-plant interactions. Their potential contribution to pest management is also discussed.

DOI： 10.1584/jpestics.J19-02

Presentation type：Oral presentation (invited, special)  

Presentation type：Oral presentation (invited, special)  

Presentation type：Oral presentation (invited, special)  

Presentation type：Oral presentation (invited, special)  

Presentation type：Oral presentation (invited, special)  

Presentation type：Oral presentation (invited, special)  

Presentation type：Oral presentation (invited, special)  

Presentation type：Oral presentation (invited, special)