Coronatine Research Articles

Bottleneck removal of paclitaxel biosynthetic pathway by overexpression of DBTNBT gene under methyl-β-cyclodextrin and coronatine elicitation in Taxus baccata L.

Paclitaxel is a highly functionalized diterpenoid that is broadly used for the treatment of several cancer types. This valuable specialized metabolite naturally exists in the inner bark of Taxus species in low amounts. The limited-scale production of paclitaxel in Taxus cell cultures has necessitated the use of several elicitors. Recently, methyl-β-cyclodextrin (CD) and coronatine (COR) have been considered to be highly effective elicitors in producing plant specialized metabolites. Given the limited production of paclitaxel due to the rate limiting enzymes' function, bottleneck removal is conducive to the production of more significant amounts of paclitaxel. In the present study, the full length of DBTNBT coding sequence (CDS), as one of the paclitaxel pathway bottlenecks, was integrated downstream of the CaMV 35S promoter (pCAMBIA1304-DBTNBT) and transiently expressed in Taxus baccata leaves via Agrobacterium tumefaciens and vacuum infiltration method. Paclitaxel production and the expression level of several involved genes were evaluated through different treatments. The transient overexpression of the DBTNBT gene, associated with dual elicitation, resulted in 7.4-fold more paclitaxel production compared with the no-inoculation/no-elicitation control. These ratios were 2.1 and 1.8 in the CD + COR and pCAM treatments, respectively. Among T13αH, T14βH, DBAT, BAPT, DBTNBT, and ABC genes, the most increased expression level belonged to the DBTNBT gene, followed by ABC and BAPT genes. It seems as though in the near future, bottleneck removal could be used on a large scale in Taxus metabolic engineering, resulting in the relative removal of some other bottlenecks and an increase in the final paclitaxel production.

ZmCOI2a and ZmCOI2b redundantly regulate anther dehiscence and gametophytic male fertility in maize.

In higher plants, the generation and release of viable pollen from anthers is vital for double fertilization and the initiation of seed development. Thus, the characterization of genes related to pollen development and anther dehiscence in plants is of great significance. The F-box protein COI1 plays a crucial role in the jasmonate (JA) signaling pathway and interacts with many JAZ family proteins in the presence of jasmonoyl-isoleucine (JA-Ile) or coronatine (COR). The mutation of AtCOI1 in Arabidopsis leads to defective anther dehiscence and male sterility (MS), although COI has not been shown to affect fertility in Zea mays (maize). Here we identified two genes, ZmCOI2a and ZmCOI2b, that redundantly regulate gametophytic male fertility. Both ZmCOI2a and ZmCOI2b are highly homologous and constitutively expressed in all tissues tested. Subcellular localization revealed that ZmCOI2a and ZmCOI2b were located in the nucleus. The coi2a coi2b double mutant, generated by CRISPR/Cas9, had non-dehiscent anthers, delayed anther development and MS. In addition, coi2a coi2b male gametes could not be transmitted to the next generation because of severe defects in pollen germination. The JA content of coi2a coi2b anthers was unaltered compared with those of the wild type, and the exogenous application of JA could not rescue the fertility defects of coi2a coi2b. Transcriptome analysis showed that the expression of genes involving the JA signaling transduction pathway, including ZmJAZ3, ZmJAZ4, ZmJAZ5 and ZmJAZ15, was affected in coi2a coi2b. However, yeast two-hybrid assays showed that ZmJAZs interacted with ZmCOI1s, but not with ZmCOI2s. In conclusion, ZmCOI2a and ZmCOI2b redundantly regulate anther dehiscence and gametophytic male fertility in maize.

Arabidopsi s Spliceosome Factor SmD3 Modulates Immunity to Pseudomonas syringae Infection.

SmD3 is a core component of the small nuclear ribonucleoprotein (snRNP) that is essential for pre-mRNA splicing. The role of Arabidopsis SmD3 in plant immunity was assessed by testing sensitivity of smd3a and smd3b mutants to Pseudomonas syringae pv. tomato (Pst) DC3000 infection and its pathogenesis effectors flagellin (flg22), EF-Tu (elf18) and coronatine (COR). Both smd3 mutants exhibited enhanced susceptibility to Pst accompanied by marked changes in the expression of key pathogenesis markers. mRNA levels of major biotic stress response factors were also altered upon treatment with Pseudomonas effectors. Our genome-wide transcriptome analysis of the smd3b-1 mutant infected with Pst, verified by northern and RT-qPCR, showed that lack of SmD3-b protein deregulates defense against Pst infection at the transcriptional and posttranscriptional levels including defects in splicing and an altered pattern of alternative splicing. Importantly, we show that SmD3-b dysfunction impairs mainly stomatal immunity as a result of defects in stomatal development. We propose that it is the malfunction of the stomata that is the primary cause of an altered mutant response to the pathogen. Other changes in the smd3b-1 mutant involved enhanced elf18- and flg22-induced callose deposition, reduction of flg22-triggered production of early ROS and boost of secondary ROS caused by Pst infection. Together, our data indicate that SmD3 contributes to the plant immune response possibly via regulation of mRNA splicing of key pathogenesis factors.

Multiple virulence factors regulated by AlgU contribute to the pathogenicity of Pseudomonas savastanoi pv. glycinea in soybean.

Pseudomonas savastanoi pv. glycinea (Psg) causes bacterial blight of soybean. To identify candidate virulence factors, transposon-mediated mutational analysis of Psg was carried out. We syringe-inoculated soybean leaves with Psg transposon mutants and identified 28 mutants which showed reduced virulence from 1,000 mutants screened. Next, we spray-inoculated soybean leaves with these mutants and demonstrated that the algU mutant showed significantly reduced virulence together with reduced bacterial populations in planta. Expression profiles comparison between the Psg wild-type (WT) and algU mutant in HSC broth revealed that expression of coronatine (COR)-related genes (including cmaA and corR) were down-regulated in the algU mutant compared with Psg WT. Moreover, we also showed that COR production were reduced in the algU mutant compared with WT. We also demonstrated that algD, which is related to alginate biosynthesis, showed reduced expression and biofilm formation was significantly suppressed in the algU mutant. Furthermore, hrpL also showed less expression in the algU mutant. These results indicate that AlgU plays a critical role in promoting Psg pathogenesis by regulating multiple virulence factors.

Physio-biochemical and proteomic mechanisms of coronatine induced potassium stress tolerance in xylem sap of cotton

Potassium (K) is a major plant nutrient, and its deficiency can limit plant growth and development. Coronatine (COR) could increase cotton seedling tolerance to K deficiency, which was hypothesized to be in relation to improving the physiological and proteomic profile of xylem sap. To test this hypothesis, cotton seedlings growth, physio-biochemical and proteomic profile of xylem sap treated with (0 and 10 nM COR) under deficient K solution (0.05 mM KCl) were explored. Compared with control, COR treatment significantly increased lateral root number and root diameter and decreased contents of malondialdehyde (MDA), some cations like potassium and calcium in xylem sap and its volume. These morphological and physiological presentations were well evidenced by differentially expressed proteins (DEPs) in the xylem sap. For instance, increasing the average root diameter and lateral roots were related to up-regulation of cobalamin-independent methionine synthase family proteins, auxin-responsive protein, and cell wall remodeling proteins such as dirigent-like protein, laccase, and the pectin lyase; lessening xylem sap volume and some cations contents was in connections with down-regulation of uclacyanin 1 and up-regulation of calmodulin-domain protein kinase 7; the MDA content reducing was associated with many PCD-related proteins’ down-regulation or loss. Furthermore, COR potentially weakened plant defense, owing to lessening or disappearing lipid-transfer proteins, signaling proteins, and other proteins positively involved in plant defense.

Coronatine Modulated the Generation of Reactive Oxygen Species for Regulating the Water Loss Rate in the Detaching Maize Seedlings

Coronatine (COR), a structural and functional mimic of jasmonates, is involved in a wide array of effects on plant development and defense response. The present study aims to investigate the role of COR, in counteracting drought stress by modulating reactive oxygen species (ROS) homeostasis, water balance, and antioxidant regulation in detached maize plants. Our results showed that COR can markedly decrease the water loss rate, but the antioxidants diphenyleneiodonium chloride (DPI) and dimethylthiourea (DMTU) eliminate the effect of water loss induced by COR. Using the dye 2′,7′-dichlorofluorescein diacetate (H2DCF-DA) loaded in the maize epidermis guard cells, it is observed that COR could increase ROS production, and then antioxidants DPI and DMTU decreased ROS production induced by COR. In addition, the expression of ZmRBOHs genes, which were associated with ROS generation was increased by COR in levels and ZmRBOHC was highly expressed in the epidermis guard cells. Moreover, COR-treated plants increased H2O2 and O2−· accumulation, antioxidant enzyme activities in control plants, while COR relieved the ROS accumulation and antioxidant enzyme activities under PEG treatment. These results indicated that COR could improve maize performance under drought stress by modulating ROS homeostasis to maintain water loss rate and antioxidant enzyme activities.

RNA-seq derived identification of coronatine-regulated genes putatively involved in terpenoid biosynthetic pathway in the rubber tree Hevea brasiliensis

Rubber tree (Hevea brasiliensis) is a woody plant of the Spurge family (Euphorbiaceae) and the most economically important member of the genus Hevea. Rubber trees are often damaged and exposed to the attack of pathogens through wounds during and after tapping periods. Jasmonic acid (JA) and methyl jasmonate (MeJA), collectively termed jasmonates (JAs), are signaling molecules that are produced by plants, especially when subjected to environmental stresses. JA/MeJA treatment triggers the biosynthesis of many specialized metabolites including terpenoids through transcriptional reprogramming. To gain molecular insights into terpenoid metabolism and mechanism underlying rubber tree’s responses to stress signals such as wounding and pathogenicity, we performed an RNA-seq analysis of H. brasiliensis treated with either water or coronatine (COR) - a structural and functional mimic of JAs. We reconstructed the transcriptomes from the published RNA-sequencing data under the guidance of the newly released high-quality chromosome-level genome assembly of H. brasiliensis cultivar GT1 and screened for genes that are differently expressed between the two treatments. The putative functions and pathways of the differently expressed genes (DEGs) were obtained by using BLASTX search against NCBI non-redundant database, followed by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) mapping. As a result, six out of the 446 identified DEGs were mapped to terpenoid metabolism pathway, five of which were up-regulated by coronatine treatment. This result provides a clue to gain insight into the genetic basis underlying terpenoid-related yield and resistance traits in the rubber tree.

Transcriptome dynamic landscape underlying the improvement of maize lodging resistance under coronatine treatment

BackgroundLodging is one of the important factors causing maize yield. Plant height is an important factor in determining plant architecture in maize (Zea mays L.), which is closely related to lodging resistance under high planting density. Coronatine (COR), which is a phytotoxin and produced by the pathogen Pseudomonas syringae, is a functional and structural analogue of jasmonic acid (JA).ResultsIn this study, we found COR, as a new plant growth regulator, could effectively reduce plant height and ear height of both hybrids (ZD958 and XY335) and inbred (B73) maize by inhibiting internode growth during elongation, thus improve maize lodging resistance. To study gene expression changes in internode after COR treatment, we collected spatio-temporal transcriptome of inbred B73 internode under normal condition and COR treatment, including the three different regions of internode (fixed, meristem and elongation regions) at three different developmental stages. The gene expression levels of the three regions at normal condition were described and then compared with that upon COR treatment. In total, 8605 COR-responsive genes (COR-RGs) were found, consist of 802 genes specifically expressed in internode. For these COR-RGs, 614, 870, 2123 of which showed expression changes in only fixed, meristem and elongation region, respectively. Both the number and function were significantly changed for COR-RGs identified in different regions, indicating genes with different functions were regulated at the three regions. Besides, we found more than 80% genes of gibberellin and jasmonic acid were changed under COR treatment.ConclusionsThese data provide a gene expression profiling in different regions of internode development and molecular mechanism of COR affecting internode elongation. A putative schematic of the internode response to COR treatment is proposed which shows the basic process of COR affecting internode elongation. This research provides a useful resource for studying maize internode development and improves our understanding of the COR regulation mechanism based on plant height.

Pseudmonas cannabina pv. alisalensis TrpA Is Required for Virulence in Multiple Host Plants.

Pseudomonas cannabina pv. alisalensis (Pcal) causes bacterial leaf spot and blight of Brassicaceae and Poaceae. We previously identified several potential Pcal virulence factors with transposon mutagenesis. Among these a trpA mutant disrupted the tryptophan synthase alpha chain, and had an effect on disease symptom development and bacterial multiplication. To assess the importance of TrpA in Pcal virulence, we characterized the trpA mutant based on inoculation test and Pcal gene expression profiles. The trpA mutant showed reduced virulence when dip- and syringe-inoculated on cabbage and oat. Moreover, epiphytic bacterial populations of the trpA mutant were also reduced compared to the wild-type (WT). These results suggest that TrpA contributes to bacterial multiplication on the leaf surface and in the apoplast, and disease development. Additionally, several Brassicaceae (including Japanese radish, broccoli, and Chinese cabbage) also exhibited reduced symptom development when inoculated with the trpA mutant. Moreover, trpA disruption led to downregulation of bacterial virulence genes, including type three effectors (T3Es) and the phytotoxin coronatine (COR), and to upregulation of tryptophan biosynthesis genes. These results indicate that a trade-off between virulence factor production and Pcal multiplication with tryptophan might be regulated in the infection processes.

Transcriptome Analysis Identified Gene Regulation Networks in Soybean Leaves Perturbed by the Coronatine Toxin

The non-host specific Pseudomonas syringae phytotoxin Coronatine (COR) causes chlorosis and promotes toxicity by inducing physiological changes in plants. We performed transcriptome analysis to better understand plants' transcriptional and metabolic response to COR. Toward this end, mock-treated and COR-treated soybean plants were analyzed by RNA-Seq. A total of 4,545 genes were differentially expressed between the two treatments, of which 2,170 were up-regulated whereas 2,375 were down-regulated in COR treated samples. Gene annotation and pathway analysis conducted using the Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) databases revealed that the differential genes were involved in photosynthesis, jasmonic acid (JA) synthesis, signal transduction, and phenylpropane metabolism. This study will provide new insights into COR mediated responses and extend our understanding of COR function in plants.

Improved biotechnological production of paclitaxel in Taxus media cell cultures by the combined action of coronatine and calix[8]arenes

Paclitaxel (PTX), a widely used anticancer agent, is found in the inner bark of several Taxus species, although at such low levels that its extraction is ecologically unsustainable. Biotechnological platforms based on Taxus sp. cell cultures offer an eco-friendlier approach to PTX production, with yields that can be improved by elicitation. However, the also limited excretion of target compounds from the producer cells to the medium hampers their extraction and purification. In this context, we studied the effect of treating T. media cell cultures with the elicitor coronatine (COR) and calix[8]arenes (CAL), nanoparticles that can host lipophilic compounds within their macrocyclic scaffold. The highest taxane production (103.5 mg.L−1), achieved after treatment with COR (1 μM) and CAL (10 mg.L−1), was 15-fold greater than in the control, and PTX represented 82% of the total taxanes analyzed. Expression levels of the flux-limiting PTX biosynthetic genes, BAPT and DBTNBT, increased after the addition of COR, confirming its elicitor action, but not CAL. The CAL treatment significantly enhanced taxane excretion, especially when production levels were increased by COR; 98% of the total taxanes were found in the culture medium after COR + CAL treatment. By forming complexes with PTX, the nanoparticles facilitated its excretion to the medium, and by protecting cells from PTX toxicity, its intra-and extra-cellular degradation may have been avoided. The addition of COR and CAL to T. media cell cultures is therefore a bio-sustainable and economically viable system to improve the yield of this important anticancer compound.

New Insight into Justicidin B Pathway and Production in Linum austriacum.

Lignans are the main secondary metabolites synthetized by Linum species as plant defense compounds but they are also valuable for human health, in particular, for novel therapeutics. In this work, Linum austriacum in vitro cultures, cells (Cc), adventitious roots (ARc) and hairy roots (HRc) were developed for the production of justicidin B through elicitation with methyl jasmonate (MeJA) and coronatine (COR). The performances of the cultures were evaluated for their stability, total phenols content and antioxidant ability. NMR was used to identify justicidin B and isojusticidin B and HPLC to quantify the production, highlighting ARc and HRc as the highest productive tissues. MeJA and COR treatments induced the synthesis of justicidin B more than three times and the synthesis of other compounds. RNA-sequencing and a de novo assembly of L. austriacum ARc transcriptome was generated to identify the genes activated by MeJA. Furthermore, for the first time, the intracellular localization of justicidin B in ARc was investigated through microscopic analysis. Then, HRc was chosen for small-scale production in a bioreactor. Altogether, our results improve knowledge on justicidin B pathway and cellular localization in L. austriacum for future scale-up processes.

Coronatine Contributes to Pseudomonas cannabina pv. alisalensis Virulence by Overcoming Both Stomatal and Apoplastic Defenses in Dicot and Monocot Plants.

Pseudomonas cannabina pv. alisalensis is a causative agent of bacterial blight of crucifers including cabbage, radish, and broccoli. Importantly, P. cannabina pv. alisalensis can infect not only a wide range of Brassicaceae spp. but, also, green manure crops such as oat. However, P. cannabina pv. alisalensis virulence mechanisms have not been investigated and are not fully understood. We focused on coronatine (COR) function, which is one of the well-known P. syringae pv. tomato DC3000 virulence factors, in P. cannabina pv. alisalensis infection processes on both dicot and monocot plants. Cabbage and oat plants dip-inoculated with a P. cannabina pv. alisalensis KB211 COR mutant (ΔcmaA) exhibited reduced virulence compared with P. cannabina pv. alisalensis wild type (WT). Moreover, ΔcmaA failed to reopen stomata on both cabbage and oat, suggesting that COR facilitates P. cannabina pv. alisalensis entry through stomata into both plants. Furthermore, cabbage and oat plants syringe-infiltrated with ΔcmaA also showed reduced virulence, suggesting that COR is involved in overcoming not only stomatal-based defense but also apoplastic defense. Indeed, defense-related genes, including PR1 and PR2, were highly expressed in plants inoculated with ΔcmaA compared with WT, indicating that COR suppresses defense-related genes of both cabbage and oat. Additionally, salicylic acid accumulation increases after ΔcmaA inoculation compared with WT. Taken together, COR contributes to causing disease by suppressing stomatal-based defense and apoplastic defense in both dicot and monocot plants. Here, we investigated COR functions in the interaction of P. cannabina pv. alisalensis and different host plants (dicot and monocot plants), using genetically and biochemically defined COR deletion mutants.[Formula: see text] The author(s) have dedicated the work to the public domain under the Creative Commons CC0 "No Rights Reserved" license by waiving all of his or her rights to the work worldwide under copyright law, including all related and neighboring rights, to the extent allowed by law, 2021.

Genome-Wide Characterization of Jasmonates Signaling Components Reveals the Essential Role of ZmCOI1a-ZmJAZ15 Action Module in Regulating Maize Immunity to Gibberella Stalk Rot.

Gibberella stalk rot (GSR) by Fusarium graminearum causes significant losses of maize production worldwide. Jasmonates (JAs) have been broadly known in regulating defense against pathogens through the homeostasis of active JAs and COI-JAZ-MYC function module. However, the functions of different molecular species of JAs and COI-JAZ-MYC module in maize interactions with Fusarium graminearum and regulation of diverse metabolites remain unknown. In this study, we found that exogenous application of MeJA strongly enhanced resistance to GSR. RNA-seq analysis showed that MeJA activated multiple genes in JA pathways, which prompted us to perform a genome-wide screening of key JA signaling components in maize. Yeast Two-Hybrid, Split-Luciferase, and Pull-down assays revealed that the JA functional and structural mimic coronatine (COR) functions as an essential ligand to trigger the interaction between ZmCOIa and ZmJAZ15. By deploying CRISPR-cas9 knockout and Mutator insertional mutants, we demonstrated that coi1a mutant is more resistant, whereas jaz15 mutant is more susceptible to GSR. Moreover, JA-deficient opr7-5opr8-2 mutant displayed enhanced resistance to GSR compared to wild type. Together, these results provide strong evidence that ZmJAZ15 plays a pivotal role, whereas ZmCOIa and endogenous JA itself might function as susceptibility factors, in maize immunity to GSR.

Data-Independent Acquisition Proteomics Unravels the Effects of Iron Ions on Coronatine Synthesis in Pseudomonas syringae pv. tomato DC3000.

Coronatine (COR) is a new type of plant growth regulator that is produced by Pseudomonas syringae pathovars and plays an important role in modulating plant growth, development, and tolerance to multiple stresses. However, the factors affecting COR production are not very clear. In this study, the effects of FeCl3 on COR production were researched. The data-independent acquisition (DIA) approach, which is a proteomic quantitative analysis method, was applied to quantitatively trace COR production and proteomic changes in P. syringae pv. tomato DC3000 under different FeCl3 culture conditions. The results showed that COR production increased with the addition of FeCl3 and that there was significant upregulation in the expression of proteins related to COR synthesis and regulation. In addition, FeCl3 also affected the expression of related proteins involved in various metabolic pathways such as glycolysis and the tricarboxylic acid cycle. Moreover, various precursors such as isoleucine and succinate semialdehyde, as well as other related proteins involved in the COR synthesis pathway, were significantly differentially expressed. Our findings revealed the dynamic regulation of COR production in response to FeCl3 at the protein level and showed the potential of using the DIA method to track the dynamic changes of the P. syringae pv. tomato DC3000 proteome during COR production, providing an important reference for future research on the regulatory mechanism of COR biosynthesis and theoretical support for COR fermentation production.

Coronatine Enhances Stalk Bending Resistance of Maize, Thickens the Cell Wall and decreases the Area of the Vascular Bundles

Coronatine (COR) is a phytotoxin produced by the pathogen Pseudomonas syringae, it has a structure similar to that of jasmonates (JAs), but it is much more active as a plant growth regulator. The goal of this study was to gain more insight into the effect and the mechanism of COR effects on stalk characteristics are related lodging resistance of maize. The agronomic traits, stalk ultrastructure, and endogenous hormones in maize stalks were studied in field trails and greenhouses, using hybrid cultivar “Xianyu 335” (XY335), “Zhengdan 958” (ZD958) and inbred line B73 as materials in 2018 and 2019. Different concentration of COR were sprayed onto maize foliar surfaces at the seven-expanded-leaves (V7) stage. Foliar application with 10 µmol L−1 of COR at the V7 stage decreased plant and ear height, increased weight and diameter of the basal internodes, and increased penetration strength and stalk bending resistance. Compared to the control treatment, in COR-treated plants, salicylic acid (SA) and jasmonic acid (JA) were decreased significantly in stalks. The treatment of 10 µmol L−1 of COR enhanced lignin accumulation, the integrity, and the thickness of cell walls in maize stalks in the early stages of stem growth in the inbred line B73, as revealed by autofluorescence microscopy and scanning electron micrographs. Our results indicated that COR improved stalk bending resistance of maize not only by optimizing stalk morphological characteristics, but also by altering hormone levels, which may led to greater lignin accumulation, thickens cell wall, and decreased the area of vascular bundles.

The changes of wheat seedlings in drought condition by exogenous coronatine (COR)

ABSTRACT Under drought and normal conditions, exogenous coronatine (COR), by the way of spraying, could induce the resistance of wheats (JiMai22 and LiangXing99). As a kind analogue of JA-Ile and compared to water-spraying, COR-spraying could improve water metabolism, photosynthesis, antioxidant enzymes activity and reduce the MDA and super anion contents. Moreover, though the way of JA acted with other hormones, especially the ABA, to regulate the plant growth, and then improve the drought resistance.

Differential Response of Tomato Plants to the Application of Three Trichoderma Species When Evaluating the Control of Pseudomonas syringae Populations.

Trichoderma species are well known biocontrol agents that are able to induce responses in the host plants against an array of abiotic and biotic stresses. Here, we investigate, when applied to tomato seeds, the potential of Trichoderma strains belonging to three different species, T. parareesei T6, T. asperellum T25, and T. harzianum T34, to control the fully pathogenic strain Pseudomonas syringae pv. tomato (Pst) DC3000, able to produce the coronatine (COR) toxin, and the COR-deficient strain Pst DC3118 in tomato plants, and the molecular mechanisms by which the plant can modulate its systemic defense. Four-week old tomato plants, seed-inoculated, or not, with a Trichoderma strain, were infected, or not, with a Pst strain, and the changes in the expression of nine marker genes representative of salicylic acid (SA) (ICS1 and PAL5) and jasmonic acid (JA) (TomLoxC) biosynthesis, SA- (PR1b1), JA- (PINII and MYC2) and JA/Ethylene (ET)-dependent (ERF-A2) defense pathways, as well as the abscisic acid (ABA)-responsive gene AREB2 and the respiratory burst oxidase gene LERBOH1, were analyzed at 72 hours post-inoculation (hpi) with the bacteria. The significant increase obtained for bacterial population sizes in the leaves, disease index, and the upregulation of tomato genes related to SA, JA, ET and ABA in plants inoculated with Pst DC3000 compared with those obtained with Pst DC3118, confirmed the COR role as a virulence factor, and showed that both Pst and COR synergistically activate the JA- and SA-signaling defense responses, at least at 72 hpi. The three Trichoderma strains tested reduced the DC3118 levels to different extents and were able to control disease symptoms at the same rate. However, a minor protection (9.4%) against DC3000 was only achieved with T. asperellum T25. The gene deregulation detected in Trichoderma-treated plus Pst-inoculated tomato plants illustrates the complex system of a phytohormone-mediated signaling network that is affected by the pathogen and Trichoderma applications but also by their interaction. The expression changes for all nine genes analyzed, excepting LERBOH1, as well as the bacterial populations in the leaves were significantly affected by the interaction. Our results show that Trichoderma spp. are not adequate to control the disease caused by fully pathogenic Pst strains in tomato plants.

Physiological and Transcriptome Profiling Analyses Reveal Important Roles of Coronatine in Improving Drought Tolerance of Tobacco

Coronatine (COR) is a phytotoxin produced by Pseudomonas syringae and is a functional analogue of the bioactive hormone JA-Ile, which is widely involved in plant defence responses. In this study, we explored the effects of exogenous applications of COR on tobacco plants under polyethylene glycol-induced drought stress. Compared with control (CK), COR-treated tobacco plants exhibited higher leaf relative water content and better photosynthetic performance under drought exposure. Ultrastructural examination revealed that drought led to stomatal closure and disorganization of granum stacking in the chloroplasts (with obvious accumulation of plastoglobuli), and mitochondria in the CK samples presented injured cristae. In the leaf tissue of the COR-treated plants, regularly stacked granum thylakoids, few plastoglobuli and intact mitochondrial membranes and cristae were observed. Totals of 1803 and 6207 differentially expressed genes (DEGs) were identified between the samples from the COR-treated and CK plants under well-watered and drought conditions. Functional annotation analysis revealed that these DEGs were involved mainly in plant hormone signal transduction, cellular carbohydrate metabolic processes and photosynthesis processes. Six hundred forty transcription factor genes were also identified among the DEGs. This study provides a global view of COR-induced drought stress tolerance in tobacco from both physiological and transcriptional aspects.

Combining Physiological and Metabolomic Analysis to Unravel the Regulations of Coronatine Alleviating Water Stress in Tobacco (Nicotiana tabacum L.).

Drought is a major abiotic stress that restricts plants growth, development, and yield. Coronatine (COR), a mimic of JA-Ile, functions in plant tolerance to multiple stresses. In our study, we examined the effects of COR in tobacco under polyethylene glycol (PEG) stress. COR treatment improved plant growth under stress as measured by fresh weight (FW) and dry weight (DW). The enzyme activity assay indicated that, under osmotic stress conditions, the activities of superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), and glutathione reductase (GR) were enhanced by COR treatment. Histochemical analyses via nitrotetrazolium blue chloride (NBT) and 3,3′-diaminobenzidine (DAB) staining showed that COR reduced reactive oxygen species (ROS) accumulation during osmotic stress. Metabolite profiles revealed that COR triggered significant metabolic changes in tobacco leaves under osmotic stress, and many essential metabolites, such as sugar and sugar derivatives, organic acids, and nitrogen-containing compounds, which might play active roles in osmotic-stressed tobacco plants, were markedly accumulated in the COR-treated tobacco. The work presented here provides a comprehensive understanding of the COR-mediated physiological, biochemical, and metabolic adjustments that minimize the adverse impact of osmotic stress on tobacco.