Priming by Hexanoic Acid Induce Activation of Mevalonic and Linolenic Pathways and Promotes the Emission of Plant Volatiles.
ABSTRACT: Hexanoic acid (Hx) is a short natural monocarboxylic acid present in some fruits and plants. Previous studies reported that soil drench application of this acid induces effective resistance in tomato plants against Botrytis cinerea and Pseudomonas syringae and in citrus against Alternaria alternata and Xanthomonas citri. In this work, we performed an in deep study of the metabolic changes produced in citrus by the application of Hx in response to the challenge pathogen A. alternata, focusing on the response of the plant. Moreover, we used (13)C labeled hexanoic to analyze its behavior inside the plants. Finally, we studied the volatile emission of the treated plants after the challenge inoculation. Drench application of (13)C labeled hexanoic demonstrated that this molecule stays in the roots and is not mobilized to the leaves, suggesting long distance induction of resistance. Moreover, the study of the metabolic profile showed an alteration of more than 200 molecules differentially induced by the application of the compound and the inoculation with the fungus. Bioinformatics analysis of data showed that most of these altered molecules could be related with the mevalonic and linolenic pathways suggesting the implication of these pathways in the induced resistance mediated by Hx. Finally, the application of this compound showed an enhancement of the emission of 17 volatile metabolites. Taken together, this study indicates that after the application of Hx this compound remains in the roots, provoking molecular changes that may trigger the defensive response in the rest of the plant mediated by changes in the mevalonic and linolenic pathways and enhancing the emission of volatile compounds, suggesting for the first time the implication of mevalonic pathway in response to hexanoic application.
Project description:Treatment with the resistance priming inducer hexanoic acid (Hx) protects tomato plants from Botrytis cinerea by activating defence responses. To investigate the molecular mechanisms underlying hexanoic acid-induced resistance (Hx-IR), we compared the expression profiles of three different conditions: Botrytis-infected plants (Inf), Hx-treated plants (Hx) and Hx-treated + infected plants (Hx+Inf). The microarray analysis at 24?h post-inoculation showed that Hx and Hx+Inf plants exhibited the differential expression and priming of many Botrytis-induced genes. Interestingly, we found that the activation by Hx of other genes was not altered by the fungus at this time point. These genes may be considered to be specific targets of the Hx priming effect and may help to elucidate its mechanisms of action. It is noteworthy that, in Hx and Hx+Inf plants, there was up-regulation of proteinase inhibitor genes, DNA-binding factors, enzymes involved in plant hormone signalling and synthesis, and, remarkably, the genes involved in oxidative stress. Given the relevance of the oxidative burst occurring in plant-pathogen interactions, the effect of Hx on this process was studied in depth. We showed by specific staining that reactive oxygen species (ROS) accumulation in Hx+Inf plants was reduced and more restricted around infection sites. In addition, these plants showed higher ratios of reduced to oxidized glutathione and ascorbate, and normal levels of antioxidant activities. The results obtained indicate that Hx protects tomato plants from B.?cinerea by regulating and priming Botrytis-specific and non-specific genes, preventing the harmful effects of oxidative stress produced by infection.
Project description:Hexanoic acid-induced resistance (Hx-IR) is effective against several pathogens in tomato plants. Our study of the mechanisms implicated in Hx-IR against Pseudomonas syringae pv. tomato?DC3000 suggests that hexanoic acid (Hx) treatment counteracts the negative effect of coronatine (COR) and jasmonyl-isoleucine (JA-Ile) on the salicylic acid (SA) pathway. In Hx-treated plants, an increase in the expression of jasmonic acid carboxyl methyltransferase (JMT) and the SA marker genes PR1 and PR5 indicates a boost in this signalling pathway at the expense of a decrease in JA-Ile. Moreover, Hx treatment potentiates 12-oxo-phytodienoic acid accumulation, which suggests that this molecule might play a role per se in Hx-IR. These results support a positive relationship between the SA and JA pathways in Hx-primed plants. Furthermore, one of the mechanisms of virulence mediated by COR is stomatal re-opening on infection with P.?syringae. In this work, we observed that Hx seems to inhibit stomatal opening in?planta in the presence of COR, which suggests that, on infection in tomato, this treatment suppresses effector action to prevent bacterial entry into the mesophyll.
Project description:The efficacy of hexanoic acid (Hx) as an inducer of resistance in tomato plants against Pseudomonas syringae pv. tomato DC3000 was previously demonstrated, and the plant response was characterized. Because little is known about the reaction of the pathogen to this effect, the goal of the present work was to determine whether the changes in the plant defence system affect the pathogen behaviour. This work provides the first demonstration of the response of the pathogen to the changes observed in plants after Hx application in terms of not only the population size but also the transcriptional levels of genes involved in quorum sensing establishment and pathogenesis. Therefore, it is possible that Hx treatment attenuates the virulence and survival of bacteria by preventing or diminishing the appearance of symptoms and controlling the growth of the bacteria in the mesophyll. It is interesting to note that the gene transcriptional changes in the bacteria from the treated plants occur at the same time as the changes in the plants. Hx is able to alter bacteria pathogenesis and survival only when it is applied as a resistance inducer because the changes that it promotes in plants affect the bacteria.
Project description:Alternaria alternata is one of the most studied fungi to date because of its impact on human life - from plant pathogenicity to allergenicity. However, its sesquiterpene emissions have not been systematically explored. Alternaria regularly co-occurs with Fusarium fungi, which are common plant pathogens, on withering plants. We analyzed the diversity and determined the absolute quantities of volatile organic compounds (VOCs) in the headspace above mycelial cultures of A. alternata and Fusarium oxysporum under different conditions (nutrient rich and poor, single cultures and co-cultivation) and at different mycelial ages. Using stir bar sorptive extraction and gas chromatography-mass spectrometry, we observed A. alternata to strongly emit sesquiterpenes, particularly during the early growth stages, while emissions from F. oxysporum consistently remained comparatively low. The emission profile characterizing A. alternata comprised over 20 sesquiterpenes with few effects from nutrient quality and age on the overall emission profile. Co-cultivation with F. oxysporum resulted in reduced amounts of VOCs emitted from A. alternata although its profile remained similar. Both fungi showed distinct emission profiles, rendering them suitable biomarkers for growth-detection of their phylotype in ambient air. The study highlights the importance of thorough and quantitative evaluations of fungal emissions of volatile infochemicals such as sesquiterpenes.
Project description:Unlike fungal and bacterial diseases, no direct method is available to control viral diseases. The use of resistance-inducing compounds can be an alternative strategy for plant viruses. Here we studied the basal response of melon to <i>Melon necrotic spot virus</i> (MNSV) and demonstrated the efficacy of hexanoic acid (Hx) priming, which prevents the virus from systemically spreading. We analysed callose deposition and the hormonal profile and gene expression at the whole plant level. This allowed us to determine hormonal homeostasis in the melon roots, cotyledons, hypocotyls, stems and leaves involved in basal and hexanoic acid-induced resistance (Hx-IR) to MNSV. Our data indicate important roles of salicylic acid (SA), 12-oxo-phytodienoic acid (OPDA), jasmonic-isoleucine, and ferulic acid in both responses to MNSV. The hormonal and metabolites balance, depending on the time and location associated with basal and Hx-IR, demonstrated the reprogramming of plant metabolism in MNSV-inoculated plants. The treatment with both SA and OPDA prior to virus infection significantly reduced MNSV systemic movement by inducing callose deposition. This demonstrates their relevance in Hx-IR against MNSV and a high correlation with callose deposition. Our data also provide valuable evidence to unravel priming mechanisms by natural compounds.
Project description:Interaction between insect herbivores and host plants can be modulated by endogenous and exogenous compounds present in the source of food and might be successfully exploited in Colorado potato beetle (CPB) pest management. Feeding tests with CPB larvae reared on three solanaceous plants (potato, eggplant and tomato) resulted in variable larval growth rates and differential susceptibility to Bacillus thuringiensis Cry3Aa toxin as a function of the host plant. An inverse correlation with toxicity was observed in Cry3Aa proteolytic patterns generated by CPB midgut brush-border membrane vesicles (BBMV) from Solanaceae-fed larvae, being the toxin most extensively proteolyzed on potato, followed by eggplant and tomato. We found that CPB cysteine proteases intestains may interact with Cry3Aa toxin and, in CPB BBMV from larvae fed all three Solanaceae, the toxin was able to compete for the hydrolysis of a papain substrate. In response to treatment with the JA-dependent plant inducer Hexanoic acid (Hx), we showed that eggplant reduced OPDA basal levels and both, potato and eggplant induced JA-Ile. CPB larvae feeding on Hx-induced plants exhibited enhanced Cry3Aa toxicity, which correlated with altered papain activity. Results indicated host-mediated effects on B. thuringiensis efficacy against CPB that can be enhanced in combination with Hx plant induction.
Project description:Plant-derived protein hydrolysates (PHs) are an important category of biostimulants able to increase plant growth and crop yield especially under environmental stress conditions. PHs can be applied as foliar spray or soil drench. Foliar spray is generally applied to achieve a relatively short-term response, whereas soil drench is used when a long-term effect is desired. The aim of the study was to elucidate the biostimulant action of PH application method (foliar spray or substrate drench) on morpho-physiological traits and metabolic profile of tomato grown under limited water availability. An untreated control was also included. A high-throughput image-based phenotyping (HTP) approach was used to non-destructively monitor the crop response under limited water availability (40% of container capacity) in a controlled environment. Moreover, metabolic profile of leaves was determined at the end of the trial. Dry biomass of shoots at the end of the trial was significantly correlated with number of green pixels (R 2 = 0.90) and projected shoot area, respectively. Both drench and foliar treatments had a positive impact on the digital biomass compared to control while the photosynthetic performance of the plants was slightly influenced by treatments. Overall drench application under limited water availability more positively influenced biomass accumulation and metabolic profile than foliar application. Significantly higher transpiration use efficiency was observed with PH-drench applications indicating better stomatal conductance. The mass-spectrometry based metabolomic analysis allowed the identification of distinct biochemical signatures in PH-treated plants. Metabolomic changes involved a wide and organized range of biochemical processes that included, among others, phytohormones (notably a decrease in cytokinins and an accumulation of salicylates) and lipids (including membrane lipids, sterols, and terpenes). From a general perspective, treated tomato plants exhibited an improved tolerance to reactive oxygen species (ROS)-mediated oxidative imbalance. Such capability to cope with oxidative stress might have resulted from a coordinated action of signaling compounds (salicylic acid and hydroxycinnamic amides), radical scavengers such as carotenoids and prenyl quinones, as well as a reduced biosynthesis of tetrapyrrole coproporphyrins.
Project description:Terpenoids are a diverse class of metabolites that impact plant metabolism in response to environmental cues. They are synthesized either via a predominantly cytosolic (MVA) pathway or a plastidic pathway (MEP). In Arabidopsis, several enzymes from the MVA and MEP pathways are encoded by gene families, excluding MVK and DXR, which are single-copy genes. In this study, we assess the diversity, evolution and expression of DXR and MVK genes in selected angiosperms and Coffea arabica in particular. Evolutionary analysis revealed that DXR and MVK underwent purifying selection, but the selection effect for DXR was stronger than it was for MVK. Digital gene expression (DGE) profile analysis of six species revealed that expression levels of MVK in flowers and roots were high, whereas for DXR peak values were observed in leaves. In C. arabica, both genes were highly expressed in flowers, and CaDXR was upregulated in response to methyl jasmonate. C. arabica DGE data were validated by assessing gene expression in selected organs, and by plants treated with hexanoic acid (Hx) using RT-qPCR. MVK expression was upregulated in roots treated with Hx. CaDXR was downregulated in leaves by Hx treatment in a genotype-specific manner, indicating a differential response to priming.
Project description:Plant-derived protein hydrolysates (PHs) have received increased attention in the last decade because of their potential to improve yield, nutritional quality as well as tolerance to abiotic stressors. The current study investigated the effects and the molecular mechanisms of a legume-derived PH under optimal and sub-optimal nitrogen (N) concentrations (112 and 7 mg L<sup>-1</sup>, respectively) in tomato (<i>Solanum lycopersicum</i> L.). Growth and mineral composition of tomato plants treated with PHs by foliar spray or substrate drench were compared to untreated plants. In addition, the expression was determined of genes encoding ammonium and nitrate transporters and seven enzymes involved in N metabolism: nitrate reductase (<i>NR</i>), nitrite reductase (<i>NiR</i>), glutamine synthetase 1 (<i>GS1</i>), glutamine synthetase 2 (<i>GS2</i>), ferredoxin-dependent glutamate synthase (<i>GLT</i>), NADH-dependent glutamate synthase (<i>GLS</i>), and glutamate dehydrogenase (<i>GDH</i>). The root and total plant dry weight, SPAD index and leaf nitrogen content were higher by 21, 17, 7, and 6%, respectively, in plants treated by a substrate drench in comparison to untreated tomato plants, whereas foliar application of PH gave intermediate values. PH-treated plants grown with lower N availability showed reduced expression of <i>NR</i> and <i>NiR</i> as well as of nitrate and ammonium transporter transcripts in both leaf and root tissues in comparison with untreated plants; this was especially pronounced after application of PH by substrate drench. Conversely, the transcript level of an amino acid transporter gene was up-regulated in comparison with untreated plants. At high N regime, the transcript levels of the ammonium and amino acid transporters and also <i>NR</i>, <i>NiR</i>, and <i>GLT</i> were significantly up-regulated in root after PH foliar and substrate drench applications compared with untreated plants. An up-regulation was also observed for <i>GS1</i>, <i>GS2</i>, and <i>GDH</i> transcripts in leaf after substrate drench. These results highlighted the potential benefits of using legume PH in vegetable production systems to increase growth and N-nutritional status of plants.
Project description:Wintersweet (Chimonanthus praecox L.) is an ornamental and economically significant shrub known for its unique flowering characteristics, especially the emission of abundant floral volatile organic compounds. Thus, an understanding of the molecular mechanism of the production of these compounds is necessary to create new breeds with high volatile production. In this study, two bHLH transcription factors (CpMYC2 and CpbHLH13) of Wintersweet H29 were functionally characterized to illustrate their possible role in the production of volatile compounds. The qRT-PCR results showed that the expression of CpMYC2 and CpbHLH13 increased from the flower budding to full bloom stage, indicating that these two genes may play an essential role in blooming and aroma production in wintersweet. Gas chromatography-mass spectroscopy (GC-MS) analysis revealed that the overexpression of CpMYC2 in arabidopsis (Arabidopsis thaliana) AtMYC2-2 mutant (Salk_083483) and tobacco (Nicotiana tabaccum) genotype Petit Havana SR1 significantly increased floral volatile monoterpene, especially linalool, while the overexpression of CpbHLH13 in Arabidopsis thaliana ecotype Columbia-0 (Col-0) and tobacco genotype SR1 increased floral sesquiterpene ?-caryophyllene production in both types of transgenic plants respectively. High expression of terpene synthase (TPS) genes in transgenic A. thaliana along with high expression of CpMYC2 and CpbHLH13 in transgenic plants was also observed. The application of a combination of methyl jasmonic acid (MeJA) and gibberellic acid (GA3) showed an increment in linalool production in CpMYC2-overexpressing arabidopsis plants, and the high transcript level of TPS genes also suggested the involvement of CpMYC2 in the jasmonic acid (JA) signaling pathway. These results indicate that both the CpMYC2 and CpbHLH13 transcription factors of wintersweet are possibly involved in the positive regulation and biosynthesis of monoterpene (linalool) and sesquiterpene (?-caryophyllene) in transgenic plants. This study also indicates the potential application of wintersweet as a valuable genomic material for the genetic modification of floral scent in other flowering plants that produce less volatile compounds.