Metabolic cross-feeding between the competent degrader Rhodococcus sp. strain p52 and an incompetent partner during catabolism of dibenzofuran: Understanding the leading and supporting roles.

Microbial interactions, particularly metabolic cross-feeding, play important roles in removing recalcitrant environmental pollutants; however, the underlying mechanisms involved in this process remain unclear. Thus, this study aimed to elucidate the mechanism by which metabolic cross-feeding occurs during synergistic dibenzofuran degradation between a highly efficient degrader, Rhodococcus sp. strain p52, and a partner incapable of utilizing dibenzofuran. A bottom-up approach combined with pairwise coculturing was used to examine metabolic cross-feeding between strain p52 and Arthrobacter sp. W06 or Achromobacter sp. D10. Pairwise coculture not only promoted bacterial pair growth but also facilitated dibenzofuran degradation. Specifically, strain p52, acting as a donor, released dibenzofuran metabolic intermediates, including salicylic acid and gentisic acid, for utilization and growth, respectively, by the partner strains W06 and D10. Both salicylic acid and gentisic acid exhibited biotoxicity, and their accumulation inhibited dibenzofuran degradation. The transcriptional activity of the genes responsible for the catabolism of dibenzofuran and its metabolic intermediates was coordinately regulated in strain p52 and its cocultivated partners, thus achieving synergistic dibenzofuran degradation. This study provides insights into microbial metabolic cross-feeding during recalcitrant environmental pollutant removal.

Modified chitosan with different phenolic acids: Characterization, physicochemical properties, and biological activity.

This study synthesized five phenolic acid-chitosan copolymers utilizing the carbodiimide-mediated chemical crosslinking reaction. Comprehensive evaluations were conducted on their structural attributes, physicochemical properties, and biological activities. Fourier transform infrared confirmed successful grafting of phenolic acids onto chitosan via amide linkages. Additionally, ultraviolet-visible absorption spectroscopy and proton nuclear magnetic resonance analyses revealed novel absorption peaks between 200 and 400 nm and 6.0-8.0 ppm, respectively, attributable to the incorporated phenolic acids. Notably, the chitosan-gentisate acid copolymer exhibited significantly enhanced biological activity (p < 0.05) compared to pure chitosan and the other four conjugates, attributed to its highest grafting degree of approximately 295.93 mg/g. These modified chitosan derivatives effectively preserved the quality of sea bass (Lateolabrax japonicus) during refrigerated storage, extending its shelf-life by up to 9 days, 7 days, and 4 days relative to control, chitosan, and gentisate acid groups.

Bringing up to date the toolkit for the catabolism of aromatic compounds in fungi: The unexpected 1,2,3,5-tetrahydroxybenzene central pathway.

Saprophytic fungi are able to catabolize many plant-derived aromatics, including, for example, gallate. The catabolism of gallate in fungi is assumed to depend on the five main central pathways, i.e., of the central intermediates' catechol, protocatechuate, hydroxyquinol, homogentisate and gentisate, but a definitive demonstration is lacking. To shed light on this process, we analysed the transcriptional reprogramming of the growth of Aspergillus terreus on gallate compared with acetate as the control condition. Surprisingly, the results revealed that the five main central pathways did not exhibit significant positive regulation. Instead, an in-depth analysis identified four highly expressed and upregulated genes that are part of a conserved gene cluster found in numerous species of fungi, though not in Aspergilli. The cluster comprises a monooxygenase gene and a fumarylacetoacetate hydrolase-like gene, which are recognized as key components of catabolic pathways responsible for aromatic compound degradation. The other two genes encode proteins with no reported enzymatic activities. Through functional analyses of gene deletion mutants in Aspergillus nidulans, the conserved short protein with no known domains could be linked to the conversion of the novel metabolite 5-hydroxydienelatone, whereas the DUF3500 gene likely encodes a ring-cleavage enzyme for 1,2,3,5-tetrahydroxybenzene. These significant findings establish the existence of a new 1,2,3,5-tetrahydroxybenzene central pathway for the catabolism of gallate and related compounds (e.g. 2,4,6-trihydroxybenzoate) in numerous fungi where this catabolic gene cluster was observed.

User-friendly platform for analysis of high mass intact proteins and glycopeptides by laser desorption/ionization-mass spectrometry based on copper oxide particles.

Matrix-assisted laser desorption/ionization-mass spectrometry (MALDI-MS) based on micro/nanostructured materials with different natures has received increasing attention for the analysis of a wide variety of analytes. However, up to now, only a few studies have shown the application of simple platforms in MALDI-MS for the identification of intact proteins. The present work reports on the application of copper oxide particles (Cu2O PS), obtained by a greener route, in combination with low amounts of 2,5-dihydroxybenzoic acid (DHB) as a novel hybrid platform. The combined Cu2O PS@DHB matrix, containing only 2.5 mg mL-1 of particles and 10 mg mL-1 of DHB, was easily applicable in MALDI-MS without surface modification of target plates. Under optimal conditions, the analysis of intact proteins up to 150,000 Da was possible, including immunoglobulin G, bovine serum albumin, and cytochrome C with adequate spot-to-spot signal reproducibility (RSD < 10%). In addition, the analysis of glycopeptides from IgG digests was carried out to prove the multipurpose application of the Cu2O PS@DHB platform in the low m/z range (2500-3000 Da). From the obtained results, it can be concluded that the optical and surface properties of as-synthesized Cu2O PS are likely to be responsible for the superior performance of Cu2O PS@DHB in comparison with conventional matrices. In this sense, the proposed user-friendly methodology opens up the prospect for possible implementation in bioanalysis and diagnostic research.

Sex differences in associations of plasma metabolites with blood pressure and heart rate variability: The HELIUS study.

BACKGROUND AND AIMS: Since plasma metabolites can modulate blood pressure (BP) and vary between men and women, we examined sex differences in plasma metabolite profiles associated with BP and sympathicovagal balance. Our secondary aim was to investigate associations between gut microbiota composition and plasma metabolites predictive of BP and heart rate variability (HRV). METHODS: From the HELIUS cohort, we included 196 women and 173 men. Office systolic BP and diastolic BP were recorded, and heart rate variability (HRV) and baroreceptor sensitivity (BRS) were calculated using finger photoplethysmography. Plasma metabolomics was measured using untargeted LC-MS/MS. Gut microbiota composition was determined using 16S sequencing. We used machine learning models to predict BP and HRV from metabolite profiles, and to predict metabolite levels from gut microbiota composition. RESULTS: In women, best predicting metabolites for systolic BP included dihomo-lineoylcarnitine, 4-hydroxyphenylacetateglutamine and vanillactate. In men, top predictors included sphingomyelins, N-formylmethionine and conjugated bile acids. Best predictors for HRV in men included phenylacetate and gentisate, which were associated with lower HRV in men but not in women. Several of these metabolites were associated with gut microbiota composition, including phenylacetate, multiple sphingomyelins and gentisate. CONCLUSIONS: Plasma metabolite profiles are associated with BP in a sex-specific manner. Catecholamine derivatives were more important predictors for BP in women, while sphingomyelins were more important in men. Several metabolites were associated with gut microbiota composition, providing potential targets for intervention.

Mechanism of Gentisic Acid on Rheumatoid Arthritis Based on miR-19b-3p/RAF1 Axis.

OBJECTIVE: To investigate the therapeutic effect of gentisic acid (GA) on rheumatoid arthritis (RA) based on the miR-19b-3p/RAF1 axis. METHODS: The cell counting kit-8 method was used to detect the growth inhibitory effect of different concentrations of GA on MH7A cells, and the drug concentration of GA was determined in the experiment. The quantificational real-time polymerase chain reaction (qRT-PCR) was used to detect the expression of miR-19b-3p and RAF1. RAF1, extracellular regulated protein kinases1/2 (ERK1/2) and phospho-ERK1/2 (p-ERK1/2) were examined by Western blotting. Three methods (dual-luciferase assay, qRT-PCR and Western blot analysis) were used to verify miR-19b-3p targeting RAF1. Flow cytometry was performed to detect MH7A cell apoptosis. Transwell and wound healing assays were used to determine the invasion and migration capacities of MH7A cells. RESULTS: The growth of MH7A cells was gradually inhibited with increasing GA concentration. When the GA concentration exceeded 80 mmol/L, GA was significantly cytotoxic to MH7A cells, so the half maximal inhibitory concentration of GA for MH7A cells was calculated as 67.019 mmol/L. GA upregulated miR-19b-3p expression, downregulated RAF1 expression, inhibited ERK1/2 phosphorylation, induced MH7A cell apoptosis and suppressed MH7A cell invasion and migration (P<0.05 or P<0.01). RAF1 was identified as the target of miR-19b-3p and reversed inhibitory effects on miR-19b-3p expression (P<0.05 or P<0.01). The miR-19b-3p inhibitor upregulated RAF1 expression and ERK1/2 phosphorylation, suppressed MH7A cell apoptosis and induced MH7A cell invasion and migration (P<0.01). CONCLUSION: GA regulated miR-19b-3p/RAF1 axis to mediate ERK pathway and inhibit the development of RA.

Mechanism of Gentisic Acid on Rheumatoid Arthritis Based on miR-19b-3p/RAF1 Axis / 中国结合医学杂志

OBJECTIVE@#To investigate the therapeutic effect of gentisic acid (GA) on rheumatoid arthritis (RA) based on the miR-19b-3p/RAF1 axis.@*METHODS@#The cell counting kit-8 method was used to detect the growth inhibitory effect of different concentrations of GA on MH7A cells, and the drug concentration of GA was determined in the experiment. The quantificational real-time polymerase chain reaction (qRT-PCR) was used to detect the expression of miR-19b-3p and RAF1. RAF1, extracellular regulated protein kinases1/2 (ERK1/2) and phospho-ERK1/2 (p-ERK1/2) were examined by Western blotting. Three methods (dual-luciferase assay, qRT-PCR and Western blot analysis) were used to verify miR-19b-3p targeting RAF1. Flow cytometry was performed to detect MH7A cell apoptosis. Transwell and wound healing assays were used to determine the invasion and migration capacities of MH7A cells.@*RESULTS@#The growth of MH7A cells was gradually inhibited with increasing GA concentration. When the GA concentration exceeded 80 mmol/L, GA was significantly cytotoxic to MH7A cells, so the half maximal inhibitory concentration of GA for MH7A cells was calculated as 67.019 mmol/L. GA upregulated miR-19b-3p expression, downregulated RAF1 expression, inhibited ERK1/2 phosphorylation, induced MH7A cell apoptosis and suppressed MH7A cell invasion and migration (P<0.05 or P<0.01). RAF1 was identified as the target of miR-19b-3p and reversed inhibitory effects on miR-19b-3p expression (P<0.05 or P<0.01). The miR-19b-3p inhibitor upregulated RAF1 expression and ERK1/2 phosphorylation, suppressed MH7A cell apoptosis and induced MH7A cell invasion and migration (P<0.01).@*CONCLUSION@#GA regulated miR-19b-3p/RAF1 axis to mediate ERK pathway and inhibit the development of RA.

SlS5H silencing reveals specific pathogen-triggered salicylic acid metabolism in tomato.

BACKGROUND: Salicylic acid (SA) is a major plant hormone that mediates the defence pathway against pathogens. SA accumulates in highly variable amounts depending on the plant-pathogen system, and several enzyme activities participate in the restoration of its levels. Gentisic acid (GA) is the product of the 5-hydroxylation of SA, which is catalysed by S5H, an enzyme activity regarded as a major player in SA homeostasis. GA accumulates at high levels in tomato plants infected by Citrus Exocortis Viroid (CEVd), and to a lesser extend upon Pseudomonas syringae DC3000 pv. tomato (Pst) infection. RESULTS: We have studied the induction of tomato SlS5H gene by different pathogens, and its expression correlates with the accumulation of GA. Transient over-expression of SlS5H in Nicotiana benthamiana confirmed that SA is processed by SlS5H in vivo. SlS5H-silenced tomato plants were generated, displaying a smaller size and early senescence, together with hypersusceptibility to the necrotrophic fungus Botrytis cinerea. In contrast, these transgenic lines exhibited an increased defence response and resistance to both CEVd and Pst infections. Alternative SA processing appears to occur for each specific pathogenic interaction to cope with SA levels. In SlS5H-silenced plants infected with CEVd, glycosylated SA was the most discriminant metabolite found. Instead, in Pst-infected transgenic plants, SA appeared to be rerouted to other phenolics such as feruloyldopamine, feruloylquinic acid, feruloylgalactarate and 2-hydroxyglutarate. CONCLUSION: Using SlS5H-silenced plants as a tool to unbalance SA levels, we have studied the re-routing of SA upon CEVd and Pst infections and found that, despite the common origin and role for SA in plant pathogenesis, there appear to be different pathogen-specific, alternate homeostasis pathways.

Enhancing the Photo and Thermal Stability of Nicotine through Crystal Engineering with Gentisic Acid.

The use of crystal engineering to convert liquids into crystalline solids remains a powerful method for inhibiting undesired degradation pathways. When nicotine, a liquid sensitive to both light and air, is combined with the GRAS-listed compound, gentisic acid, the resulting crystalline solid, exhibits enhanced photo and thermal stability. Despite a modest ΔTm of 42.7 °C, the melting point of 155.9 °C for the nicotinium gentisate salt is the highest reported for nicotine-containing crystalline solids. An analysis of the crystal packing and thermodynamic properties provides context for the observed properties.

Submicron 3,4-dihydroxybenzoic acid-TiO2 composite particles for enhanced MALDI MS imaging of secondary metabolites in the root of differently aged baical skullcap.

Matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) has become an attractive technique for the localization and visualization of small molecules in various biological tissue sections. In this work, submicron 3,4-dihydroxybenzoic acid-TiO2 composite particles (3,4-DHB-TiO2 CPs) were synthesized for enhanced MALDI MSI of secondary metabolites in the root of Scutellaria baicalensis Georgi (baical skullcap). Submicron TiO2 particles were synthesized as starting materials by using a facile sol-gel method and chemically modified with six analogs of dihydroxybenzoic acids (DHB) (2,3-DHB, 2,4-DHB, 2,5-DHB, 2,6-DHB, 3,4-DHB, and 3,5-DHB). Among them, 3,4-DHB-TiO2 CPs provided superior performance in MALDI MSI of small molecules. Compared with conventional matrices, such as 2,5-dihydroxybenzoic acid (2,5-DHB) and α-cyano-4-hydroxycinnamic acid (CHCA), 3,4-DHB-TiO2 CPs exhibited low background noise and high detection sensitivity for the visualization of spatial distribution patterns of secondary metabolites in the roots of differently aged S. baicalensis by using MALDI MSI. The age-related spatial and content changes of flavonoids in S. baicalensis roots were demonstrated and further validated by liquid chromatography-mass spectrometry (LC-MS). This work provides a potential organic-inorganic hybrid matrix for MALDI MSI of secondary metabolites in plant tissues.

Transcriptome and proteome profiling reveals complex adaptations of Candida parapsilosis cells assimilating hydroxyaromatic carbon sources.

Many fungal species utilize hydroxyderivatives of benzene and benzoic acid as carbon sources. The yeast Candida parapsilosis metabolizes these compounds via the 3-oxoadipate and gentisate pathways, whose components are encoded by two metabolic gene clusters. In this study, we determine the chromosome level assembly of the C. parapsilosis strain CLIB214 and use it for transcriptomic and proteomic investigation of cells cultivated on hydroxyaromatic substrates. We demonstrate that the genes coding for enzymes and plasma membrane transporters involved in the 3-oxoadipate and gentisate pathways are highly upregulated and their expression is controlled in a substrate-specific manner. However, regulatory proteins involved in this process are not known. Using the knockout mutants, we show that putative transcriptional factors encoded by the genes OTF1 and GTF1 located within these gene clusters function as transcriptional activators of the 3-oxoadipate and gentisate pathway, respectively. We also show that the activation of both pathways is accompanied by upregulation of genes for the enzymes involved in ß-oxidation of fatty acids, glyoxylate cycle, amino acid metabolism, and peroxisome biogenesis. Transcriptome and proteome profiles of the cells grown on 4-hydroxybenzoate and 3-hydroxybenzoate, which are metabolized via the 3-oxoadipate and gentisate pathway, respectively, reflect their different connection to central metabolism. Yet we find that the expression profiles differ also in the cells assimilating 4-hydroxybenzoate and hydroquinone, which are both metabolized in the same pathway. This finding is consistent with the phenotype of the Otf1p-lacking mutant, which exhibits impaired growth on hydroxybenzoates, but still utilizes hydroxybenzenes, thus indicating that additional, yet unidentified transcription factor could be involved in the 3-oxoadipate pathway regulation. Moreover, we propose that bicarbonate ions resulting from decarboxylation of hydroxybenzoates also contribute to differences in the cell responses to hydroxybenzoates and hydroxybenzenes. Finally, our phylogenetic analysis highlights evolutionary paths leading to metabolic adaptations of yeast cells assimilating hydroxyaromatic substrates.

The protective effect of gentisic acid on rheumatoid arthritis via the RAF/ERK signaling pathway.

BACKGROUND: RAF and ERK pathways are known to be activated in human rheumatoid arthritis (RA) fibroblast-like synoviocytes (FLS), which play an important role in the pathogenesis and destruction of RA. Gentisic acid (GA) was a natural product derived from plants, which has been reported can attenuate pressure overload-induced cardiac hypertrophy and fibrosis in mice through inhibition of the ERK1/2 pathway. Whether GA can inhibit the occurrence and development of RA through RAF/ERK signaling pathway has not been reported. The purpose of this study is to determine whether GA may have a certain therapeutic effect on RA-FLS. METHOD: Bovine type II collagen was used to establish a rat model of rheumatism. Enzyme-linked immunosorbent assay was used to detect inflammatory factors, anti-inflammatory mediators, and rheumatoid factor. Hematoxylin and eosin and TUNEL staining were used to detect the effect of GA on histochemical with rheumatoid arthritis. RAF, ERK, and p-ERK expressions in synovial tissue were measured by western blot and immunohistochemical. Besides, human rheumatoid arthritis fibroblast-like synoviocytes cell line MH7A was used to investigate the biological behavior influenced by GA. Apoptosis assay was performed to detect apoptosis of GA on MH7A cells. Transwell invasion assay was performed to detect the ability of cell migration. RESULT: The result showed that GA could reduce joint swelling and inflammation. At the same time, it can also promote the apoptosis of synovial cells and down-regulate the RAF/ERK pathway. CONCLUSION: GA may ameliorate inflammatory factors' abnormality, synovial hyperplasia, and apoptosis of synovium via inhibiting the RAF/ERK signaling pathway.

Spectral and time domain fluorescence spectroscopy of gentisic acid molecule in protic and aprotic polymer matrix.

In the present work, the effect of polymer microenvironment on the photophysics of gentisic acid molecule [2,5-dihydroxybenzoic acid] (GA), steady-state and time-domain fluorescence measurements at different pH conditions were carried out in protic [polyvinyl alcohol PVA] and aprotic [polymethyl methacrylate (PMMA)] polymer matrices. Change in the proticity of the microenvironment of the polymer traps different ionic species along with the neutral form of rotamer P and R conformers of GA molecule, are found to be responsible for the change in the spectral, multi-exponential decay behaviour. In protic polymer, the appearance of a single emission band indicates, dissociation of the GA molecule is very high, and it present as a monoanion along with hydrogen-bonded P and R rotamers. However, in the basic polymer film, most of the conformers of R converted to the anion. In contrast, protonation slows down the dissociation of both P and R forms in the acidic film. Unlike PVA matrix, in PMMA, dual emission band appears due to slow dissociation of GA molecule and hydrogen-bonded rotamer P, and R form exists with monoanion species. The magnitude of large stokes shifted red emission due to excited-state intramolecular proton transfers (ESIPT) found grater in rotamer P compared to its anionic species (green emission) and a blue emission corresponds to rotamer R.

Design, synthesis and characterization of lysozyme-gentisic acid dual-functional conjugates with antibacterial/antioxidant activities.

Both microbiological and chemical food spoilages remain to be the major challenges in the food industry's efforts to combat food waste and loss because of the lack of high efficacy food preservatives. In this study, dual-functional conjugates that simultaneously suppress both lipid oxidation and microorganism growth are fabricated by covalently conjugating natural antioxidant gentisic acid (GA) on native antibacterial lysozyme (Lys). The mixing ratio of Lys and GA determines the particle size, morphology, antioxidant activity, and antimicrobial performance of the ensuing conjugates. With more of GA being grafted, a drastic decrease in the net surface charge with the concomitant occurrence of aggregations are observed in the conjugates. The maximum antioxidant activity and antibacterial performance of the conjugates is achieved when Lys:GA molar ratio is 1:112. The findings could guide the rational design of future functional food ingredients that combine multiple natural bioactive compounds to effectively intervene food waste and loss.

Two LysR Family Transcriptional Regulators, McbH and McbN, Activate the Operons Responsible for the Midstream and Downstream Pathways, Respectively, of Carbaryl Degradation in Pseudomonas sp. Strain XWY-1.

Previously, a LysR family transcriptional regulator, McbG, that activates the mcbBCDEF gene cluster involved in the upstream pathway (from carbaryl to salicylate) of carbaryl degradation in Pseudomonas sp. strain XWY-1 was identified by us (Z. Ke, Y. Zhou, W. Jiang, M. Zhang, et al., Appl Environ Microbiol 87:e02970-20, 2021, https://doi.org/10.1128/AEM.02970-20). In this study, we identified McbH and McbN, which activate the mcbIJKLM cluster (responsible for the midstream pathway, from salicylate to gentisate) and the mcbOPQ cluster (responsible for the downstream pathway, from gentisate to pyruvate and fumarate), respectively. They both belong to the LysR family of transcriptional regulators. Gene disruption and complementation study reveal that McbH is essential for transcription of the mcbIJKLM cluster in response to salicylate and McbN is indispensable for the transcription of the mcbOPQ cluster in response to gentisate. The results of electrophoretic mobility shift assay (EMSA) and DNase I footprinting showed that McbH binds to the 52-bp motif in the mcbIJKLM promoter area and McbN binds to the 58-bp motif in the mcbOPQ promoter area. The key sequence of McbH binding to the mcbIJKLM promoter is a 13-bp motif that conforms to the typical characteristics of the LysR family. However, the 12-bp motif that is different from the typical characteristics of the LysR family regulator binding site sequence is identified as the key sequence for McbN to bind to the mcbOPQ promoter. This study revealed the regulatory mechanisms for the midstream and downstream pathways of carbaryl degradation in strain XWY-1 and further our knowledge of (and the size of) the LysR transcription regulator family. IMPORTANCE The enzyme-encoding genes involved in the complete degradation pathway of carbaryl in Pseudomonas sp. strain XWY-1 include mcbABCDEF, mcbIJKLM, and mcbOPQ. Previous studies demonstrated that the mcbA gene, responsible for hydrolysis of carbaryl to 1-naphthol, is constitutively expressed and that the transcription of mcbBCDEF was regulated by McbG. However, the transcription regulation mechanisms of mcbIJKLM and mcbOPQ have not been investigated yet. In this study, we identified two LysR-type transcriptional regulators, McbH and McbN, which activate the mcbIJKLM cluster (responsible for the degradation of salicylate to gentisate) and the mcbOPQ cluster (responsible for the degradation of gentisate to pyruvate and fumarate), respectively. The 13-bp motif is critical for McbH to bind to the promoter of mcbIJKLM, and 12-bp motif different from the typical characteristics of the LysR-type transcriptional regulator (LTTR) binding sequence affects the binding of McbN to the promoter. These findings help to expand the understanding of the regulatory mechanism of microbial degradation of carbaryl.

Modulation of Inflammatory Pathways and Adipogenesis by the Action of Gentisic Acid in RAW 264.7 and 3T3-L1 Cell Lines.

Gentisic acid (GA), a benzoic acid derivative present in various food ingredients, has been shown to have diverse pharmaceutical activities such as anti-carcinogenic, antioxidant, and hepatoprotective effects. In this study, we used a co-culture system to investigate the mechanisms of the anti-inflammatory and anti-adipogenic effects of GA on macrophages and adipocytes, respectively, as well as its effect on obesity-related chronic inflammation. We found that GA effectively suppressed lipopolysaccharide-stimulated inflammatory responses by controlling the production of nitric oxide and pro-inflammatory cytokines and modulating inflammation-related protein pathways. GA treatment also inhibited lipid accumulation in adipocytes by modulating the expression of major adipogenic transcription factors and their upstream protein pathways. Furthermore, in the macrophage-adipocyte co-culture system, GA decreased the production of obesity-related cytokines. These results indicate that GA possesses effective anti-inflammatory and anti-adipogenic activities and may be used in developing treatments for the management of obesity-related chronic inflammatory diseases.

Loss of function of a DMR6 ortholog in tomato confers broad-spectrum disease resistance.

Plant diseases are among the major causes of crop yield losses around the world. To confer disease resistance, conventional breeding relies on the deployment of single resistance (R) genes. However, this strategy has been easily overcome by constantly evolving pathogens. Disabling susceptibility (S) genes is a promising alternative to R genes in breeding programs, as it usually offers durable and broad-spectrum disease resistance. In Arabidopsis, the S gene DMR6 (AtDMR6) encodes an enzyme identified as a susceptibility factor to bacterial and oomycete pathogens. Here, we present a model-to-crop translational work in which we characterize two AtDMR6 orthologs in tomato, SlDMR6-1 and SlDMR6-2. We show that SlDMR6-1, but not SlDMR6-2, is up-regulated by pathogen infection. In agreement, Sldmr6-1 mutants display enhanced resistance against different classes of pathogens, such as bacteria, oomycete, and fungi. Notably, disease resistance correlates with increased salicylic acid (SA) levels and transcriptional activation of immune responses. Furthermore, we demonstrate that SlDMR6-1 and SlDMR6-2 display SA-5 hydroxylase activity, thus contributing to the elucidation of the enzymatic function of DMR6. We then propose that SlDMR6 duplication in tomato resulted in subsequent subfunctionalization, in which SlDMR6-2 specialized in balancing SA levels in flowers/fruits, while SlDMR6-1 conserved the ability to fine-tune SA levels during pathogen infection of the plant vegetative tissues. Overall, this work not only corroborates a mechanism underlying SA homeostasis in plants, but also presents a promising strategy for engineering broad-spectrum and durable disease resistance in crops.

Unveiling the CoA mediated salicylate catabolic mechanism in Rhizobium sp. X9.

Salicylate is a typical aromatic compound widely distributed in nature. Microbial degradation of salicylate has been well studied and salicylate hydroxylases play essential roles in linking the peripheral and ring-cleavage catabolic pathways. The direct hydroxylation of salicylate catalyzed by salicylate-1-hydroxylase or salicylate-5-hydroxylase has been well studied. However, the CoA mediated salicylate 5-hydroxylation pathway has not been characterized in detail. Here, we elucidate the molecular mechanism of the reaction in the conversion of salicylate to gentisate in the carbaryl-degrading strain Rhizobium sp. X9. Three enzymes (salicylyl-CoA ligase CehG, salicylyl-CoA hydroxylase CehH and gentisyl-CoA thioesterase CehI) catalyzed the conversion of salicylate to gentisate via a route, including CoA thioester formation, hydroxylation and thioester hydrolysis. Further analysis indicated that genes cehGHI are also distributed in other bacteria from terrestrial environment and marine sediments. These genomic evidences highlight the role of this salicylate degradation pathway in the carbon cycle of soil organic compounds and marine sediments. Our findings of this three-step strategy enhanced the current understanding of CoA mediated degradation of salicylate.

Heat-killed endophytic bacterium induces robust plant defense responses against important pathogens.

Stress caused by pathogens strongly damages plants. Developing products to control plant disease is an important challenge in sustainable agriculture. In this study, a heat-killed endophytic bacterium (HKEB), Bacillus aryabhattai, is used to induce plant defense against fungal and bacterial pathogens, and the main defense pathways used by the HKEB to activate plant defense are revealed. The HKEB induced high protection against different pathogens through the salicylic and jasmonic acid pathways. We report the presence of gentisic acid in the HKEB for the first time. These results show that HKEBs may be a useful tool for the management of plant diseases.

Influence of calcium channel modulators on the production of serotonin, gentisic acid, and a few other biosynthetically related phenolic metabolites in seedling leaves of salt tolerant rice variety Nonabokra.

Rice, a most salt-sensitive cereal plant, adopts diverse pathways to withstand sodium chloride-induced salinity-related adversities. During the present study, attempt was made to understand the role of calcium on metabolite profile of the leaves of salt tolerant rice seedlings of variety of Nonabokra under sodium chloride induced salinity, by Gas Chromatography-Mass Spectrometry-based metabolomics approach. Calcium availability in the seedlings was reduced or enhanced applying inhibitors (vanadyl sulfate, lanthanum chloride, and verapamil) or promoters of calcium influx (calcimycin also known as calcium ionophore A23187) in the sodium chloride (100 mM) supplemented growth medium. Growth medium of ten-day-old seedlings was replaced by sodium chloride supplemented hydroponic solution with promotor or inhibitors of calcium channel. Fifteen days old seedlings were harvested. It was observed that depletion of calcium availability increased the level of serotonin and gentisic acid whereas increased calcium level decreased these metabolites. It was concluded from the results that production of the signaling molecules serotonin and gentisic acids was elevated in calcium-deficient seedlings under salt stress the condition that was considered as control during the experiment. The two signaling molecules probably help this tolerant rice variety Nonabokra to withstand the salt-induced adversities.