Defense Responses In Tobacco Research Articles

Lactic acid induced defense responses in tobacco against Phytophthora nicotianae

Induced resistance is considered an eco-friendly disease control strategy, which can enhance plant disease resistance by inducing the plant’s immune system to activate the defense response. In recent years, studies have shown that lactic acid can play a role in plant defense against biological stress; however, whether lactic acid can improve tobacco resistance to Phytophthora nicotianae, and its molecular mechanism remains unclear. In our study, the mycelial growth and sporangium production of P. nicotianae were inhibited by lactic acid in vitro in a dose-dependent manner. Application of lactic acid could reduce the disease index, and the contents of total phenol, salicylic acid (SA), jasmonic acid (JA), lignin and H2O2, catalase (CAT) and phenylalanine ammonia–lyase (PAL) activities were significantly increased. To explore this lactic acid-induced protective mechanism for tobacco disease resistance, RNA-Seq analysis was used. Lactic acid enhances tobacco disease resistance by activating Ca2+, reactive oxygen species (ROS) signal transduction, regulating antioxidant enzymes, SA, JA, abscisic acid (ABA) and indole-3-acetic acid (IAA) signaling pathways, and up-regulating flavonoid biosynthesis-related genes. This study demonstrated that lactic acid might play a role in inducing resistance to tobacco black shank disease; the mechanism by which lactic acid induces disease resistance includes direct antifungal activity and inducing the host to produce direct and primed defenses. In conclusion, this study provided a theoretical basis for lactic acid-induced resistance and a new perspective for preventing and treating tobacco black shank disease.

Harpin-loaded chitosan nanoparticles induced defense responses in tobacco

Phytopathogens entry into plant cell is restricted by several chemical substances which hamper the natural ecosystem and are expensive. Induction of defense responses in plants by elicitors is a better alternative approach to combat phytopathogens. Controlled and steady release of elicitors from nanoparticles core will improve the bioavailability and alleviate the repeated application of molecules that triggers the hypersensitive response (HR) in plants. Here we reported the efficacy of chitosan-harpin as nanoparticles in eliciting defense responses in tobacco wherein harpin (30 ng/mL) loaded chitosan nanoparticles (H-CSNPs) were prepared by ionotropic gelation method using chitosan and sodium tripolyphosphate (TPP). The nanoparticles were spherical and possess a size of about ∼160 nm with a zeta potential of 30.2 ± 0.7 mV. The FT-IR and XRD studies confirmed the formation of chitosan nanoparticles. The encapsulation efficiency of harpin was 85% and steady release was observed up to 4 h in releasing kinetics. Topical application of H-CSNPs resulted histopathological evidences which coincided with the induction HR specific marker genes viz., pathogenises related proteins and induction of defense enzymes. The harpin at nanogram concentrations with chitosan improved the bioavailability than the harpin or chitosan alone in the tobacco plant.

Retrieving the in vivo Scopoletin Fluorescence Excitation Band Allows the Non-invasive Investigation of the Plant-Pathogen Early Events in Tobacco Leaves.

In this study, we developed and applied a new spectroscopic fluorescence method for the in vivo detection of the early events in the interaction between tobacco (Nicotiana tabacum L.) plants and pathogenic bacteria. The leaf disks were infiltrated with a bacterial suspension in sterile physiological solution (SPS), or with SPS alone as control. The virulent Pseudomonas syringae pv. tabaci strain ATCC 11528, its non-pathogenic ΔhrpA mutant, and the avirulent P. syringae pv. tomato strain DC3000 were used. At different post-infiltration time–points, the in vivo fluorescence spectra on leaf disks were acquired by a fiber bundle-spectrofluorimeter. The excitation spectra of the leaf blue emission at 460 nm, which is mainly due to the accumulation of coumarins following a bacterial infiltration, were processed by using a two-bands Gaussian fitting that enabled us to isolate the scopoletin (SCT) contribution. The pH-dependent fluorescence of SCT and scopolin (SCL), as determined by in vitro data and their intracellular localization, as determined by confocal microscopy, suggested the use of the longer wavelength excitation band at 385 nm of 460 nm emission (F385_460) to follow the metabolic evolution of SCT during the plant–bacteria interaction. It was found to be directly correlated (R2 = 0.84) to the leaf SCT content, but not to that of SCL, determined by HPLC analysis. The technique applied to the time-course monitoring of the bacteria–plant interaction clearly showed that the amount and the timing of SCT accumulation, estimated by F385_460, was correlated with the resistance to the pathogen. As expected, this host defense response was delayed after P. syringae pv. tabaci ATCC 11528 infiltration, in comparison to P. syringae pv. tomato DC3000. Furthermore, no significant increase of F385_460 (SCT) was observed when using the non-pathogenic ΔhrpA mutant of P. syringae pv. tabaci ATCC 11528, which lacks a functional Type Three Secretion System (TTSS). Our study showed the reliability of the developed fluorimetric method for a rapid and non-invasive monitoring of bacteria-induced first events related to the metabolite-based defense response in tobacco leaves. This technique could allow a fast selection of pathogen-resistant cultivars, as well as the on-site early diagnosis of tobacco plant diseases by using suitable fluorescence sensors.

Effect of Feeding Stage and Density of Whiteflies on Subsequent Aphid Performance on Tobacco Plants

Bemisia tabaci Middle East-Asia Minor 1 (MEAM1) is a cosmopolitan, highly polyphagous agricultural pest, which has the capacity to displace other native insect herbivores. Here, equipped with an integrated approach, the effect of developmental stages and feeding density of whiteflies on Myzus persicae performance in tobacco plants are investigated. Bioassay results showed that B. tabaci nymphs, but not adult, pre-infestation significantly reduced survival and fecundity of M. persicae, and the strongest resistance to M. persicae was detected at the medium density (9–10 nymphs/cm2). Neither low nor high feeding density of B. tabaci nymphs triggered visible resistance to aphids. However, no significant results were detected in salicylate-deficient NahG plants after B. tabaci nymph infestation. In addition to performance distinctions, hormone quantification and qPCR results revealed very different effects for nymph and adult whitefly stages on the defense responses in tobacco. B. tabaci nymph infestation significantly increased SA accumulation and SA-responsive genes (PR-1a, PR-2a) expression but suppressed JA-regulated responses. In contrast, tobacco plants responded to adult infestation by slightly increasing in both SA- and JA-regulated defenses. Furthermore, higher transcription level of Bt56, coding gene of a secretory salivary effector, was recorded in nymphs vs. adults, while silencing of Bt56 by virus-induced gene silencing (VIGS) partly impaired the aphid resistance induced by B. tabaci nymphs. These results proved that the induction of tobacco defense responses varied with the feeding stages of whiteflies: nymphs of B. tabaci, but not adults, induced a defense response against aphids, with a density threshold for this induced resistance.

Internalization and induction of defense responses in tobacco by harpinPss conjugated gold nanoparticles as a foliar spray

Controlled and targeted delivery of elicitor molecules with improved bioavailability is required to treat plant diseases. Here we reported stable, reproducible, and nano-sized (20–22 nm) negatively (H-AuNPs) and positively (H-CAuNPs) charged harpin adsorbed nanoparticles to induce defense responses in tobacco. The foliar application of NPs facilitated their entry possibly through epidermal stomata accumulated in the apoplast and symplast eliciting the hypersensitive response (HR) with the up-regulation of pathogenesis-related gene transcripts (PR-1, PR-2, PR-3, chitinase, HSR203, and HR specific marker gene ‘HIN1’) in 6 h. Immunogold labeling of harpinPss revealed its internalization (symplast & apoplast) and correlated with the induction of defense responses. Studies with fluorescently labeled NPs demonstrated the uptake and distribution within the leaf. Transmission electron microscopy observations revealed the internalization of H-CAuNPs and long-distance transport, eliciting systemic acquired resistance. Our observation holds promise in effectively minimizing the concentration of harpin for field application.

Method for the Production and Purification of Plant Immuno-Active Xylanase from Trichoderma.

Plants lack a circulating adaptive immune system to protect themselves against pathogens. Therefore, they have evolved an innate immune system based upon complicated and efficient defense mechanisms, either constitutive or inducible. Plant defense responses are triggered by elicitors such as microbe-associated molecular patterns (MAMPs). These components are recognized by pattern recognition receptors (PRRs) which include plant cell surface receptors. Upon recognition, PRRs trigger pattern-triggered immunity (PTI). Ethylene Inducing Xylanase (EIX) is a fungal MAMP protein from the plant-growth-promoting fungi (PGPF)–Trichoderma. It elicits plant defense responses in tobacco (Nicotiana tabacum) and tomato (Solanum lycopersicum), making it an excellent tool in the studies of plant immunity. Xylanases such as EIX are hydrolytic enzymes that act on xylan in hemicellulose. There are two types of xylanases: the endo-1, 4-β-xylanases that hydrolyze within the xylan structure, and the β-d-xylosidases that hydrolyze the ends of the xylan chain. Xylanases are mainly synthesized by fungi and bacteria. Filamentous fungi produce xylanases in high amounts and secrete them in liquid cultures, making them an ideal system for xylanase purification. Here, we describe a method for cost- and yield-effective xylanase production from Trichoderma using wheat bran as a growth substrate. Xylanase produced by this method possessed xylanase activity and immunogenic activity, effectively inducing a hypersensitive response, ethylene biosynthesis, and ROS burst.

The oligomeric states of elicitins affect the hypersensitive response and resistance in tobacco.

Successful plant defence against microbial pathogens is based on early recognition and fast activation of inducible responses. Key mechanisms include detection of microbe-associated molecular patterns by membrane-localized pattern recognition receptors that induce a basal resistance response. A well-described model of such responses to pathogens involves the interactions between Solanaceae plants and proteinaceous elicitors secreted by oomycetes, called elicitins. It has been hypothesized that the formation of oligomeric structures by elicitins could be involved in their recognition and activation of defensive transduction cascades. In this study, we tested this hypothesis using several approaches, and we observed differences in tobacco plant responses induced by the elicitin β-cryptogein (β-CRY) and its homodimer, β-CRYDIM. We also found that the C-terminal domain of elicitins of other ELI (true-elicitin) clades plays a significant role in stabilization of their oligomeric structure and restraint in the cell wall. In addition, covalently cross-linking β-CRYDIM impaired the formation of signalling complexes, thereby reducing its capacity to elicit the hypersensitive response and resistance in the host plant, with no significant changes in pathogenesis-related protein expression. By revealing the details of the effects of β-CRY dimerization on recognition and defence responses in tobacco, our results shed light on the poorly understood role of elicitins' oligomeric structures in the interactions between oomycetes and plants.

Root Exudates Metabolic Profiling Suggests Distinct Defense Mechanisms Between Resistant and Susceptible Tobacco Cultivars Against Black Shank Disease.

There is increasing evidence that root exudates play important roles in plant disease resistance. Black shank, caused by Phytophthora nicotianae, is a destructive soil-borne disease in tobacco (Nicotiana tabacum L.). The aim of the present study was to investigate the activity and composition of the root exudates from resistant and susceptible tobacco cultivars. The root exudates of the resistant cultivar Gexin 3 showed inhibitory activity against P. nicotianae, while the exudates of susceptible cultivar Xiaohuangjin 1025 stimulated the colony growth but had no effect on spore germination. Metabolic profiling using liquid chromatography/electrospray ionization-quadrupole-time-of-flight mass spectrometry depicted differing metabolic patterns of root exudates between Gexin 3 and Xiaohuangjin 1025. The activity and composition of root exudates was altered by P. nicotianae inoculation. Multivariate analysis showed that root exudates (including organic acids, alkaloids, fatty acids, and esters) were different between the two varieties. The defense substances in root exudates, such as tartaric acid, ferulic acid, and lauric acid, may represent a pre-infection prevention strategy for tobacco. Phenylpropanoids as well as inducers of salicylic acid, fatty acids, 6-hydroxyhexanoic acid, and hydrojasmonate may be involved in tobacco defense responses. Compared to the susceptible cultivar, the roots of the resistant cultivar exhibited high enzyme activities of phenylalanine ammonia-lyase, cinnamate-4-hydroxylase and 4-coumarate-CoA ligase, which may prompt the synthesis and secretion of phenylpropanoids. Our results indicated that the root exudates not only provide a pre-infection prevention strategy by exuding antimicrobial substances, but also increase tobacco disease resistance by eliciting plant defense responses. In addition, some defense compounds as well as compounds that play a role in inducing plant defense responses, showed potential for disease control application. This study provides an insight into possible disease resistance mechanisms of root exudates, and attempts the beneficial utilization of these secondary metabolites of plants.

MiR159 Represses a Constitutive Pathogen Defense Response in Tobacco.

MicroR159 (miR159) regulation of GAMYB expression is highly conserved in terrestrial plants; however, its functional role remains poorly understood. In Arabidopsis (Arabidopsis thaliana), although GAMYB-like genes are constitutively transcribed during vegetative growth, their effects are suppressed by strong and constitutive silencing by miR159. GAMYB expression occurs only if miR159 function is inhibited, which results in detrimental pleiotropic defects, questioning the purpose of the miR159-GAMYB pathway. Here, miR159 function was inhibited in tobacco (Nicotiana tabacum) and rice (Oryza sativa) using miRNA MIM159 technology. Similar to observations in Arabidopsis, inhibition of miR159 in tobacco and rice resulted in pleiotropic defects including stunted growth, implying functional conservation of the miR159-GAMYB pathway among angiosperms. In MIM159 tobacco, transcriptome profiling revealed that genes associated with defense and programmed cell death were strongly activated, including a suite of 22 PATHOGENESIS-RELATED PROTEIN (PR) genes that were 100- to 1,000-fold upregulated. Constitutive expression of a miR159-resistant GAMYB transgene in tobacco resulted in phenotypes similar to that of MIM159 tobacco and activated PR gene expression, verifying the dependence of the above-mentioned changes on GAMYB expression. Consistent with the broad defense response, MIM159 tobacco appeared immune to Phytophthora infection. These findings suggest that the tobacco miR159-GAMYB pathway functions in the biotic defense response, which becomes activated upon miR159 inhibition. However, PR gene expression was not upregulated in Arabidopsis or rice when miR159 was inhibited, suggesting that miR159-GAMYB pathway functional differences exist between species, or factors in addition to miR159 inhibition are required in Arabidopsis and rice to activate this broad defense response.

Parasitic Wasp Mediates Plant Perception of Insect Herbivores.

Microplitis croceipes is a solitary parasitoid that specializes on noctuid larvae of Helicoverpa zea and Heliothis virescens. Both the parasitoid and its hosts are naturally distributed across a large part of North America. When parasitoids deposit their eggs into hosts, venom and polydnaviruses (PDVs) are also injected into the caterpillars, which can suppress host immune responses, thus allowing parasitoid larvae to develop. In addition, PDVs can regulate host oral cues, such as glucose oxidase (GOX). The purpose of this study was to determine if parasitized caterpillars differentially induce plant defenses compared to non-parasitized caterpillars using two different caterpillar host/plant systems. Heliothis virescens caterpillars parasitized by M. croceipes had significantly lower salivary GOX activity than non-parasitized caterpillars, resulting in lower levels of tomato defense responses, which benefited parasitoid performance by increasing the growth rate of parasitized caterpillars. In tobacco plants, parasitized Helicoverpa zea caterpillars had lower GOX activity but induced higher plant defense responses. The higher tobacco defense responses negatively affected parasitoid performance by reducing the growth rate of parasitized caterpillars, causing longer developmental periods, and reduced cocoon mass and survival of parasitoids. These studies demonstrate a species-specific effect in different plant-insect systems. Based on these results, plant perception of insect herbivores can be affected by parasitoids and lead to positive or negative consequences to higher trophic levels depending upon the particular host-plant system.

A novel protein elicitor PeFOC1 from Fusarium oxysporum triggers defense response and systemic resistance in tobacco

In recent years, it is a hotspot research field on interaction mechanism between elicitor and plant. In this study, a novel hypersensitive response (HR)-inducing protein elicitor was isolated from the culture filtrate of Fusarium oxysporum f. sp. cubense and named PeFOC1, which consisted of 321 amino acids with a molecular weight of approximately 35 kDa. After the inducible expression in Escherichia coli and purification by ÄKTA explore system, the recombinant PeFOC1 also triggered a typical HR in tobacco. In addition, PeFOC1 induced a cascade of defense response in tobacco including production of hydrogen peroxide, deposition of callose, and accumulation of phenolic compounds. Moreover, PeFOC1 significantly improved systemic resistance of tobacco seedlings to tobacco mosaic virus and Pseudomonas syringae pv. tabaci. Real-time quantitative-PCR analysis indicated that several defense-related genes in tobacco, such as NtPR1a, NtNPR1, NtPAL, NtEDS1, NtPDF, and NtLOX, were all up-regulated by the treatment of PeFOC1. All these results collectively demonstrated that PeFOC1 triggered defense response and systemic acquired resistance (SAR) in tobacco. This research not only provides further research on immune mechanism between plant and elicitor, but also sheds new light on strategy for biocontrol in the future.

Comparative transcriptome analysis reveals differential gene expression in resistant and susceptible tobacco cultivars in response to infection by cucumber mosaic virus

Cucumber mosaic virus (CMV) is one of the most severe viral diseases transmitted by aphids infecting Solanum crops in China, causing great losses of crop yields and income in rural communities. The tobacco cultivars NC82 and Taiyan 8 are closely related but differ in resistance to CMV. NC82 is susceptible to infection and Taiyan 8 is resistant, but the mechanisms underlying this difference in resistance are not clear. In this study, we conducted RNA sequencing to analyze changes in gene expression induced in the leaves of Taiyan 8 and NC82 upon systemic infection with CMV, compared with gene expression in the leaves of mock-inoculated plants. Leaves were sampled at one, three, eight, and 15 days after infection. In total, 3443 and 747 differentially expressed genes were identified in Taiyan 8 and NC82, respectively. Gene ontology and pathway enrichment analyses revealed that the different responses to CMV infection between cultivars were based on microtubule-based processes, pentose and glucuronate interconversions, plant–pathogen interaction, and hormone signal transduction pathways. Genes encoding pathogenesis-related proteins, disease-resistance proteins, lipoxygenase, cellulose synthase, an auxin response factor, and an ethylene receptor showed different expression patterns. The differences in gene expression following CMV infection likely contributed to the different resistance levels of these two tobacco cultivars. The comprehensive transcriptome dataset described here, which includes candidate response genes, will serve as a resource for further studies of the molecular mechanisms associated with tobacco defense responses against CMV.

Ethyl Gallate Displays Elicitor Activities in Tobacco Plants.

Alkyl gallates showed elicitor activities on tobacco in both whole plants and cell suspensions. Methyl gallate (MG), ethyl gallate (EG), and propyl gallate (PG) infiltration into tobacco leaves induced hypersensitive reaction-like lesions and topical production of autofluorescent compounds revealed under UV light. When sprayed on tobacco plants at 5 mM, EG promoted upregulation of defense-related genes such as the antimicrobial PR1, β-1,3-glucanase PR2, Chitinase PR3, and osmotin PR5 target genes. Tobacco BY-2 cells challenged with EG underwent cell death in 48 h, which was significantly reduced in the presence of the protease inhibitor aprotinin. The three alkyl gallates all caused alkalinization of the BY-2 extracellular medium, whereas gallic acid did not trigger any pH variation. Using EGTA or LaCl3, we showed that Ca2+ mobilization occurred in BY-2 cells elicited with EG. Overall, our findings are the first evidence of alkyl gallate elicitor properties with early perception events on the plasma membrane, potential hypersensitive reactions, and PR-related downstream defense responses in tobacco.

Transcriptome analysis of resistant and susceptible tobacco (Nicotiana tabacum) in response to root-knot nematode Meloidogyne incognita infection

The root-knot nematode (RKN) Meloidogyne incognita reproduces on the roots of tobacco (Nicotiana tabacum), damaging crops, reducing crop yield, and causing economic losses annually. The development of resistant genotypes is an alternative strategy to effectively control these losses. However, the molecular mechanism responsible for host pathogenesis and defense responses in tobacco specifically against RKNs remain poorly understood. Here, root transcriptome analysis of resistant (Yuyan12) and susceptible (Changbohuang) tobacco varieties infected with RKNs was performed. Moreover, 2623 and 545 differentially expressed genes (DEGs) in RKN-infected roots were observed in Yuyan12 and Changbohuang, respectively, compared to those in non-infected roots, including 289 DEGs commonly expressed in the two genotypes. Among these DEGs, genes encoding cell wall modifying proteins, auxin-related proteins, the ROS scavenging system, and transcription factors involved in various biological and physiochemical processes were significantly expressed in both the resistant and susceptible genotypes. This work is thus the first report on the relationships in the RKN-tobacco interaction using transcriptome analysis, and the results provide important information on the mechanism of RKN resistance in tobacco.

Overexpression of SpWRKY1 promotes resistance to Phytophthora nicotianae and tolerance to salt and drought stress in transgenic tobacco.

WRKY transcription factors are key regulatory components of plant responses to biotic and abiotic stresses. SpWRKY1, a pathogen-induced WRKY gene, was isolated from tomato (Solanum pimpinellifolium L3708) using in silico cloning and reverse transcriptase-polymerase chain reaction (RT-PCR) methods. SpWRKY1 expression was significantly induced following oomycete pathogen infection and treatment with salt, drought, salicylic acid (SA), methyl jasmonate (MeJA) and abscisic acid (ABA). Overexpression of SpWRKY1 in tobacco conferred greater resistance to Phytophthora nicotianae infection, as evidenced by lower malondialdehyde (MDA) content; relative electrolyte leakage (REL); higher chlorophyll content; and higher peroxidase (POD, EC 1.11.1.7), superoxide dismutase (SOD, EC 1.15.1.1) and phenylalanine ammonia-lyase (PAL, EC 4.3.1.24) activities. This resistance was also coupled with enhanced expression of SA- and JA-associated genes (NtPR1, NtPR2, NtPR4, NtPR5 and NtPDF1.2), as well as of various defense-related genes (NtPOD, NtSOD and NtPAL). In addition, transgenic tobacco plants also displayed an enhanced tolerance to salt and drought stresses, mainly demonstrated by the transgenic lines exhibiting lower accumulation of MDA content and higher POD (EC 1.11.1.7), SOD (EC 1.15.1.1) activities, chlorophyll content, photosynthetic rate and stomatal conductance, accompanied by enhanced expression of defense-related genes (NtPOD, NtSOD, NtLEA5, NtP5CS and NtNCED1) under salt and drought stresses. Overall, these findings suggest that SpWRKY1 acts as a positive regulator involved in tobacco defense responses to biotic and abiotic stresses.

A novel elicitor identified from Magnaporthe oryzae triggers defense responses in tobacco and rice.

Our studies indicate a potential important elicitor candidate which can aid in the fight against a worldwide disease, rice blast. In this study, we report the purification, identification, characterization, and gene cloning of a novel hypersensitive response-inducing protein elicitor (MoHrip2) secreted from an important pathogenic fungus, Magnaporthe oryzae. The protein fraction was isolated from the culture filtrate of M. oryzae and identified by de novo sequencing. The elicitor-encoding gene mohrip2 was cloned following sequence comparison and PCR amplification. This 459-bp gene encodes a 152-residue polypeptide that contains an 18-residue signal peptide and exhibits a pI of 4.72 and an apparent molecular mass of 16kDa. The hypothetical protein, MoHrip2, was expressed in Escherichia coli, and both the recombinant and the endogenous protein caused necrotic lesions in tobacco leaves. In addition to phenolic compound deposition and alkalization of the extracellular medium, MoHrip2 also induced hydrogen peroxide production and nitric oxide accumulation in tobacco cells. Moreover, rice seedlings treated with MoHrip2 exhibited pronounced resistance to M. oryzae compared with control seedlings.

Leucine zipper-like motifs of HrpZPss are not essential to induce hypersensitive response in tobacco.

Harpins are heat-stable, glycine-rich proteins secreted by several Gram-negative phytopathogenic bacteria, involved in the induction of hypersensitive response in non-host plants. HrpZPss is highly heat stable, exists as a polydispersed, multimeric protein in nature and the functional significance for its existence in many oligomeric forms, remains still unclear. It was suggested that the leucine zipper-like motifs (LZMs) may take part in the formation of oligomeric aggregates, which may be responsible for HR elicitation by harpins and for their high thermal stability. To test the involvement of LZMs in HR elicitation and induction of defense responses, we have generated deletion mutants with or without LZMs and analysed their ability to induce HR, and/or activate the defense responses. The deletion mutant HrpZMM1, with no LZM also elicited HR as well as defense responses in tobacco suggesting the LZMs are not essential for elicitor activity of HrpZPss. However, the ability to elicit defense responses may be linked with the ability to induce HR.

Metabolic Survey of Defense Responses to a Compatible Hemibiotroph, Phytophthora parasitica var. nicotianae, in Ethylene Signaling-Impaired Tobacco

Reactive oxygen species (ROS) and ethylene play an important role in determining the resistance or susceptibility of plants to pathogen attack. A previous study of the response of tobacco cultivar ( Nicotiana tabacum L. cv. Wisconsin 38) to a compatible hemibiotroph, Phytophthora parasitica var. nicotianae (Ppn) showed that biphasic bursts of ROS and ethylene are positively associated with disease severity. The levels of ethylene and ROS might influence the susceptibility of plants to pathogens, with changing levels of metabolite related to disease resistance or susceptibility. In this study, to obtain more detailed information on the interaction of ROS and ethylene signaling related to resistance and/or susceptibility of plants to pathogen, Ppn-induced metabolic profiles from wild type (WT) and ethylene signaling-impaired transgenic plants that expressed Ein3 antisense (Ein3-AS) were compared using ultraperformance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS). Nonredundant mass ions (576 in ESI+ mode and 336 in ESI- mode) were selected, and 56 mass ions were identified on the basis of their accurate mass ions and MS/MS spectra. Two-way hierarchical clustering analysis of the selected mass ions revealed that nicotine and phenylpropanoid-polyamine conjugates, such as caffeoyl-dihydrocaffeoyl-spermidine, dicaffeoyl-spermidine, caffeoyl-feruloyl-spermidine, and two bis(dihydrocaffeoyl)-spermine isomers, and their intermediates, such as arginine and putrecine, were present at lower levels in Ein3-AS transgenic plants during Ppn interaction than in WT, whereas galactolipid and oxidized free fatty acid levels were higher in Ein3-AS transgenic plants. Taken together, these results reveal a function for ethylene signaling in tobacco defense responses during Ppn interaction.

The overexpression of a maize mitogen‐activated protein kinase gene (ZmMPK5) confers salt stress tolerance and induces defence responses in tobacco

As sessile organisms, plants are exposed to potential dangers, including multiple biotic and abiotic stresses. The mitogen-activated protein kinase (MAPK) is a universal signalling pathways involved in these processes. A previous study showed that maize ZmMPK5 is induced by various stimuli at transcriptional and post-translational levels. In this study, ZmMPK5 was overexpressed in tobacco to further analyse its biological functions. Under salt and oxidative stresses, ZmMPK5-overexpressing lines displayed less severe damage and stronger growth phenotypes corresponding to a series of physiological changes. In addition, the transgenic lines accumulated less reactive oxygen species (ROS) and had higher levels of antioxidant enzyme activity and metabolites than wild-type (WT) plants following NaCl treatment. Quantitative RT-PCR revealed that the expression of ROS-related and stress-responsive genes was higher in transgenic plants than in WT plants. Furthermore, transgenic lines exhibited enhanced resistance to viral pathogens, and expressed constitutively higher transcript levels of pathogenesis-related genes, such as PR1a, PR4, PR5 and EREBP. Taken together, these results demonstrated that ZmMPK5 is involved in salt stress, oxidative stress and pathogen defence signalling pathways, and its function may be at least partly devoted to efficiently eliminating ROS accumulation under salt stress.

The splice variant Ntr encoded by the tobacco resistance gene N has a role for negative regulation of antiviral defense responses

Tobamovirus infection or transient elicitor expression induces a hypersensitive response (HR) in tobacco plants carrying the N resistance gene, which produces two gene products (N and Ntr) through alternative splicing. In this study, the biological significance of Ntr was investigated. The transient overexpression of Ntr markedly inhibited the induction of HR and N-dependent antiviral defense responses in tobacco and Nicotiana benthamiana, respectively. In addition, there were no drastic changes observed in the ratio of the two alternative transcripts during early tobamovirus infection. Our data suggest that Ntr may play a role as a negative regulator of N-dependent antiviral defenses.