Accumulation Of Pathogenesis-related Proteins Research Articles

Antimicrobial peptide CB-M exhibits direct antifungal activity against Botrytis cinerea and induces disease resistance to gray mold in cherry tomato fruit

Gray mold is a common postharvest disease caused by Botrytis cinerea that causes fruit decay and reduces fruit storage quality. In this study, the antifungal activity of antimicrobial peptide CB-M against B. cinerea and its mechanism of action, as well as the ability of CB-M to induce disease resistance in cherry tomato fruit, were investigated. CB-M had a strong inhibitory effect on spore survival and mycelial growth of B. cinerea, and these effects were enhanced with increased CB-M concentration. Moreover, significant leakages of cellular constituents and fluorescence microscopy assay confirmed that CB-M treatment could destroy the cell membrane integrity of B. cinerea. Scanning electron microscopy observations further demonstrated obvious distortion and shriveling of B. cinerea hyphae. Gel retardation and SDS-PAGE assay showed that CB-M could likely bind to cellular DNA and degrade/inhibit protein synthesis in B. cinerea. Furthermore, preincubation with CB-M before pathogen inoculation significantly decreased lesion diameter and incidence of gray mold caused by B. cinerea in tomato, cherry tomato, and grape fruit throughout whole storage time. Cherry tomato was used to explore the ability of CB-M to induce gray mold resistance, and the results showed that CB-M preincubation enhanced the activities of superoxide dismutase, peroxidase, and catalase, accelerated the accumulation of pathogenesis-related proteins, flavonoids, and total phenolics, and increased the activities of β-1,3-glucanase, chitinase, and phenylalanine ammonia-lyase during storage. These results indicated that CB-M is a good substitute for fungicides and has excellent prospects for controlling postharvest gray mold in horticultural crops.

Design, Synthesis, and Fungicidal Activity of Novel Plant Elicitors Based on a Diversity-Oriented Synthesis Strategy

The novel plant elicitors, 3-benzyl-5-[1-(2-oxo-4-phenyl-1-oxaspiro[4.5]dec-3-en-3-yl)ethylidene]-2-aminoimidazolin-4-one derivatives, were designed based on the diversity-oriented synthesis strategy and synthesized in four steps via the Knoevenagel condensation reaction as the key step. They were characterized by 1H NMR, 13C NMR, HR-ESI-MS, and X-ray diffraction. The position of the C═N bond of Z- and E-configuration compounds was determined by X-ray diffraction. The in vivo fungicidal activity evaluation revealed that most of these compounds exhibited remarkable activities (100%) against Pseudoperonospora cubensis at 400 μg/mL, among which compound 8e still exhibited excellent protective activity with a 50% inhibition rate at 0.1 μg/mL. Because the in vitro effect on tested phytopathogens was poor, the mechanism to induce the immune responses and reinforce the resistance of cucumber against Botrytis cinerea was studied. The results indicated that the compound 8e-mediated defense response against B. cinerea was based on the accumulation of pathogenesis-related proteins and cell wall reinforcement by callose deposition. Quantitative analysis of salicylic acid (SA) and jasmonic acid (JA) and the increased expression of induced resistance-related genes and the defense-associated phenylalanine ammonia lyase revealed that the immune response triggered by compound 8e was highly associated with the SA signaling pathway. Significant upregulation of JA-related genes Cs-AOS2 indicated that the JA signaling pathway was also influenced. It was also shown that the plants treated with compound 8e promoted primary root elongation, which resulted in enhanced plant growth. Most importantly, these compounds have completely new structures compared with the traditional plant elicitors. Further research of 8e-mediated plant disease resistance might have a great influence on the development of plant elicitors.

Transient proteolysis reduction of Nicotiana benthamiana-produced CAP256 broadly neutralizing antibodies using CRISPR/Cas9.

The hypersensitive response is elicited by Agrobacterium infiltration of Nicotiana benthamiana, including the induction and accumulation of pathogenesis-related proteins, such as proteases. This includes the induction of the expression of several cysteine proteases from the C1 (papain-like cysteine protease) and C13 (legumain-like cysteine protease) families. This study demonstrates the role of cysteine proteases: NbVPE-1a, NbVPE-1b, and NbCysP6 in the proteolytic degradation of Nicotiana benthamiana (glycosylation mutant ΔXTFT)-produced anti-human immunodeficiency virus broadly neutralizing antibody, CAP256-VRC26.25. Three putative cysteine protease cleavage sites were identified in the fragment crystallizable region. We further demonstrate the transient coexpression of CAP256-VRC26.25 with CRISPR/Cas9-mediated genome editing vectors targeting the NbVPE-1a, NbVPE-1b, and NbCysP6 genes which resulted in a decrease in CAP256-VRC26.25 degradation. No differences in structural features were observed between the human embryonic kidney 293 (HEK293)-produced and ΔXTFT broadly neutralizing antibodies produced with and without the coexpression of genome-editing vectors. Furthermore, despite the presence of proteolytically degraded fragments of plant-produced CAP256-VRC26.25 without the coexpression of genome editing vectors, no influence on the in vitro functional activity was detected. Collectively, we demonstrate an innovative in planta strategy for improving the quality of the CAP256 antibodies through the transient expression of the CRISPR/Cas9 vectors.

Systemic resistance: Plant responses to interaction with fungal bio-control agents

Systemic resistance in plant against phytopathogens is mainly associated with systemic acquired resistance (SAR) and induced systemic resistance (ISR). SAR and ISR are mechanisms of induced defense that confers long-lasting protection against a broad spectrum of phytopathogens. SAR requires the signal molecule salicylic acid (SA) while ISR signal molecules are jasmonic acid (JA) and ethylene. These are associated with accumulation of pathogenesis-related proteins (PRP) which contribute resistance inside plant. Bio-control fungi (BCF) such as Trichoderma spp. and Pseudomonas spp. are agents that control plant diseases. They have the ability to control numerous soil-borne as well as many foliar and fruit pathogens. BCF also augments nutrient uptake and increase nitrogen use efficiency in crop plants. Some strains also have abilities to improve photosynthetic efficiency of plants. Furthermore, PcPCL1606 has displayed additional traits regarding its fitness in soil and plant root environments such as soil survival, efficient plant root colonization, promotion of plant growth. The present review highlights systemic resistance in plant concerns with SAR and ISR-mediated functional traits, such as enhancement of plant growth and bio-control of phytopathogens.

A novel glycoprotein from Streptomyces sp. triggers early responses of plant defense

GP-1, a novel glycoprotein from Streptomyces sp. ZX01 has a plant immunity-inducing effect. GP-1-treated plants exhibited enhanced systemic resistance with a significant reduction in TMV lesions on tobacco leaves, but its antiviral mechanism remains unclear. In this study, early plant defense-related responses, such as Ca2+ influx, callose apposition, oxidative burst, hypersensitive response, programmed cell death, increase in nitric oxide (NO), and stomatal closure, were investigated under GP-1 treatment, and the mechanism of how GP-1 induces viral resistance in Nicotiana benthamiana was studied. Results showed that GP-1 induced [Ca2+]cyt rapidly in tobacco leaves and suspended cells, followed by reactive oxygen species (ROS) and NO elevation. Transcriptome analysis showed significant differences in expression levels between the GP-1-treated N. benthamiana and the control and showed significantly upregulated and enriched pathways including defense and immune reaction. Similar to typical pathogen-associated molecular patterns, GP-1 induced callose deposition and stomatal closure to form defense barriers against pathogen invasion. The expression of defense-related genes further confirmed the above conclusions. By analyzing transcriptome in N. benthamiana and the contents of salicylic acid (SA) and jasmonic acid (JA), GP-1 enhanced viral resistance of tobacco by improving the SA and JA contents, strengthening plant secondary metabolites activities, promoting systemic accumulation of pathogenesis-related proteins in TMV- inoculated tobacco there by producing antiviral activity.

Mechanizmy obronne roślin drzewiastych uruchamiane w odpowiedzi na atak patogenów

This paper presents a comprehensive study on the mechanisms of tree responses to pathogens. We describe natural resistance concerning the presence of different types of barriers protecting plants from invasion and also give them strength after a pathogen attack. We put emphasis on induced resistance functioning both locally and systemically and involvement of phytohormones signaling networks. Systemic acquired resistance (SAR) involved the action of salicylic acid and H202 and accumulation of pathogenesis-related proteins. In turn, jasmonates and ethylene are signaling molecules in the induced resistance (SIR). All these substances play a crucial role in the forest management and can be applied in the tree protection strategies based on the natural and synthetic active compounds.

ITRAQ-based proteomics of sunflower cultivars differing in resistance to parasitic weed Orobanche cumana.

Orobanche cumana is an obligate root parasite causing severe damage to many economically important crops, including sunflowers worldwide. For efficient control measures, it is necessary to understand the resistant mechanism during interaction at molecular level. The present study emphasizes on comparative proteomics to investigate the mechanistic basis of compatible and incompatible interaction of O. cumana with resistant (JY207) and susceptible (TK0409) sunflowers. More than 3500 proteins were identified from two cultivars by iTRAQ analysis. Identified proteins associated with general functions, posttranslational modification, energy production and conversion, carbohydrate transport and metabolism, and signal transduction mechanisms were the most represented category of induced proteins in both cultivars. The resistant interaction was characterized by alteration of defense-related proteins involved in recognition of parasites, accumulation of pathogenesis-related proteins, biosynthesis of lignin, and detoxification of toxic metabolites in JY207 after inoculation. The susceptible interaction was characterized by decreased abundance of proteins involved in biosynthesis and signaling of plant growth regulators including auxin, gibberellin, brassinosteroid, and ethylene in TK0409 after inoculation. The present study provides comprehensive details of proteins and differential modulation of pathways regulated under compatible and incompatible interaction, allowing the identification of important molecular components for development of sustainable resistance against this parasite.

Plant immunity induced by COS-OGA elicitor is a cumulative process that involves salicylic acid

Plant innate immunity offers considerable opportunities for plant protection but beside flagellin and chitin, not many molecules and their receptors have been extensively characterized and very few have successfully reached the field. COS-OGA, an elicitor that combines cationic chitosan oligomers (COS) with anionic pectin oligomers (OGA), efficiently protected tomato (Solanum lycopersicum) grown in greenhouse against powdery mildew (Leveillula taurica). Leaf proteomic analysis of plants sprayed with COS-OGA showed accumulation of Pathogenesis-Related proteins (PR), especially subtilisin-like proteases. qRT-PCR confirmed upregulation of PR-proteins and salicylic acid (SA)-related genes while expression of jasmonic acid/ethylene-associated genes was not modified. SA concentration and class III peroxidase activity were increased in leaves and appeared to be a cumulative process dependent on the number of sprayings with the elicitor. These results suggest a systemic acquired resistance (SAR) mechanism of action of the COS-OGA elicitor and highlight the importance of repeated applications to ensure efficient protection against disease.

The effects of grapevine trunk diseases (GTDs) on vine physiology

Esca disease as well as Botryosphaeria and Eutypa dieback cause considerable economic problems for vineyards worldwide, and currently, no efficient treatment is available to control these diseases. For these three grapevine trunk diseases (GTDs), the main physiological effects reported concern carbohydrate metabolism and defence responses in the different organs of vine. In the trunk, a depletion of starch reserves in woody tissues is associated with fungal colonization; in the leaves, where pathogens are not present, the carbohydrate metabolism is also affected as revealed by a decline of the photosynthetic rate. A consequence of these disturbances is a lower pool of carbon reserves that might contribute to a decrease of plant development and vigour during the subsequent year. Other metabolic activities such as lipid and amino acid metabolism are down regulated. The perturbation of these primary metabolisms is often associated with the induction of defence responses. The development of biochemical barriers resulting from the accumulation of both tyloses and gummosis is observed during the infection of the wood causing blockage of the xylem vessels and thus limiting the fungal invasion. Their progression in the wood is also inhibited by the formation of polyphenol-rich reaction zones and by the accumulation of pathogenesis-related proteins, and the oxidative burst and the production of reactive oxygen species. Additionally, detoxification processes of the vine are involved; this reaction could be linked to the production of extracellular compounds by GTD agents some of which are phytotoxic. As a consequence, the sensory quality of berries and probably the wine made from these berries decrease. This review presents an overview of the physiological modifications described in vines affected by GTDs.

Leaf proteome rebalancing in Nicotiana benthamiana for upstream enrichment of a transiently expressed recombinant protein.

A key factor influencing the yield of biopharmaceuticals in plants is the ratio of recombinant to host proteins in crude extracts. Postextraction procedures have been devised to enrich recombinant proteins before purification. Here, we assessed the potential of methyl jasmonate (MeJA) as a generic trigger of recombinant protein enrichment in Nicotiana benthamiana leaves before harvesting. Previous studies have reported a significant rebalancing of the leaf proteome via the jasmonate signalling pathway, associated with ribulose 1,5-bisphosphate carboxylase oxygenase (RuBisCO) depletion and the up-regulation of stress-related proteins. As expected, leaf proteome alterations were observed 7 days post-MeJA treatment, associated with lowered RuBisCO pools and the induction of stress-inducible proteins such as protease inhibitors, thionins and chitinases. Leaf infiltration with the Agrobacterium tumefaciens bacterial vector 24 h post-MeJA treatment induced a strong accumulation of pathogenesis-related proteins after 6 days, along with a near-complete reversal of MeJA-mediated stress protein up-regulation. RuBisCO pools were partly restored upon infiltration, but most of the depletion effect observed in noninfiltrated plants was maintained over six more days, to give crude protein samples with 50% less RuBisCO than untreated tissue. These changes were associated with net levels reaching 425 μg/g leaf tissue for the blood-typing monoclonal antibody C5-1 expressed in MeJA-treated leaves, compared to less than 200 μg/g in untreated leaves. Our data confirm overall the ability of MeJA to trigger RuBisCO depletion and recombinant protein enrichment in N. benthamiana leaves, estimated here for C5-1 at more than 2-fold relative to host proteins.

Trichoderma viride induces pathogenesis related defense response against rot pathogen infection in groundnut (Arachis hypogaea L.)

The study examine induction of defense enzymes involved in phenylpropanoid pathway and accumulation of pathogenesis related proteins in rot pathogen (Aspergillus niger Van Tieghem) challenged groundnut seedlings in response to Trichoderma viride JAU60. Seeds of five groundnut varieties differing in collar rot susceptibility were sown under non-infested, pathogen infested and pathogen+T. viride JAU60 seed treatment. Collar rot disease evident between 31.0% (J-11, GG-2) and 67.4% (GG-20) in different groundnut varieties under pathogen infested which was significantly reduced from 58.1% (J-11, GG-2) to 51.6% (GG-20) by Trichoderma treatment. The specific activities of polyphenol oxidase (EC 1.14.18.1) and β-1,3 glucanase (EC 3.2.1.6) elevated 3.5 and 2.3-fold, respectively, at 3days; phenylalanine ammonia lyase (EC 4.3.1.5) evident 1.6-fold higher at 6days; and chitinase (EC 3.2.1.14) sustained 2.3–2.8 folds up to 9days in Trichoderma treated+pathogen infested seedlings of tolerant varieties (J-11, GG-2) compared with moderate and susceptible (GAUG-10, GG-13, GG-20). T. viride JAU60 induces defense enzymes in a different way for tolerant and susceptible varieties to combat the disease. This study indicates the synergism activation of defense enzymes under the pathogenic conditions or induced resistance by T. viride JAU60 in a different groundnut varieties susceptible to collar rot disease.

Comparative Analysis of Defense Responses in Chocolate Spot-Resistant and -Susceptible Faba Bean (Vicia faba) Cultivars Following Infection by the Necrotrophic Fungus Botrytis fabae.

In this study, resistance responses were investigated during the interaction of Botrytis fabae with two faba bean cultivars expressing different levels of resistance against this pathogen, Nubaria (resistant) and Giza 40 (susceptible). Disease severity was assessed on leaves using a rating scale from 1 to 9. Accumulation levels of reactive oxygen species (ROS), lipid peroxidation and antioxidant enzymes (superoxide dismutase, catalase and ascorbate peroxidase) were measured in leaf tissues at different times of infection. The expression profiles of two pathogenesis-related proteins (PRPs) encoded by the genes PR-1 and β-1,3-glucanase were also investigated using reverse transcription RT-PCR analysis. The accumulation of these defense responses was induced significantly in both cultivars upon infection with B. fabae compared with un-inoculated controls. The resistant cultivar showed weaker necrotic symptom expression, less ROS accumulation, a lower rate of lipid peroxidation and higher activity of the enzymatic ROS scavenging system compared with susceptible cultivar. Interestingly, ROS accumulated rapidly in the resistant leaf tissues and peaked during the early stages of infection, whereas accumulation was stronger and more intense in the susceptible tissues in later stages. Moreover, the response of the resistant cultivar to infection was earlier and stronger, exhibiting high transcript accumulation of the PR genes. These results indicated that the induction of oxidant/antioxidant responses and the accumulation of PRPs are part of the faba bean defense mechanism against the necrotrophic fungus B. fabae with a different intensity and timing of induction, depending on the resistance levels.

The pathogenesis-related protein PR-4b from Theobroma cacao presents RNase activity, Ca(2+) and Mg(2+) dependent-DNase activity and antifungal action on Moniliophthora perniciosa.

BackgroundThe production and accumulation of pathogenesis-related proteins (PR proteins) in plants in response to biotic or abiotic stresses is well known and is considered as a crucial mechanism for plant defense. A pathogenesis-related protein 4 cDNA was identified from a cacao-Moniliophthora perniciosa interaction cDNA library and named TcPR-4b.ResultsTcPR-4b presents a Barwin domain with six conserved cysteine residues, but lacks the chitin-binding site. Molecular modeling of TcPR-4b confirmed the importance of the cysteine residues to maintain the protein structure, and of several conserved amino acids for the catalytic activity. In the cacao genome, TcPR-4b belonged to a small multigene family organized mainly on chromosome 5. TcPR-4b RT-qPCR analysis in resistant and susceptible cacao plants infected by M. perniciosa showed an increase of expression at 48 hours after infection (hai) in both cacao genotypes. After the initial stage (24-72 hai), the TcPR-4b expression was observed at all times in the resistant genotypes, while in the susceptible one the expression was concentrated at the final stages of infection (45-90 days after infection). The recombinant TcPR-4b protein showed RNase, and bivalent ions dependent-DNase activity, but no chitinase activity. Moreover, TcPR-4b presented antifungal action against M. perniciosa, and the reduction of M. perniciosa survival was related to ROS production in fungal hyphae.ConclusionTo our knowledge, this is the first report of a PR-4 showing simultaneously RNase, DNase and antifungal properties, but no chitinase activity. Moreover, we showed that the antifungal activity of TcPR-4b is directly related to RNase function. In cacao, TcPR-4b nuclease activities may be related to the establishment and maintenance of resistance, and to the PCD mechanism, in resistant and susceptible cacao genotypes, respectively.

Induction of systemic resistance against tobacco mosaic virus by Ningnanmycin in tobacco

Ningnanmycin (NNM) is an antiviral agent firstly isolated from Strepcomces noursei var·xichangensisn. Studies have shown that NNM promotes PAL, POD and SOD activity and possesses antiviral activity against tobacco mosaic virus (TMV). In this study, our results demonstrated that NNM inhibited the polymerization process of TMV coat protein (TMV-CP) in vitro and promoted the systemic accumulation of pathogenesis-related proteins (PRs), which are the markers of systemic acquired resistance (SAR). An non-expressor, pathogenesis-related genes 1 (NPR1) that regulates SAR and induces systemic resistance (ISR), increased. In addition, the Jaz3 expression increase showed that NNM also induced ISR. Based on the results of this work and earlier reports, it is suggesting that NNM induces tobacco systemic resistance against TMV via activating multiple plant defense signaling pathways.

Induce the plant resistance to pathogen infection

Systemic acquired resistance (SAR) is a mechanism of induced defense that confers long-lasting protection against a broad spectrum of microorganisms. Salicylic acid (SA) is the signal molecule which is required for induce SAR and is associated with accumulation of pathogenesis-related proteins, which are thought to contribute to resistance. SA paly vital role in some related resistance gene expression in plant cell which have direct or indirect effect on pathogen growth as SA has direct toxicity for pathogen and in the same time has stimulation effect for some enzyme related to reduce the oxidative burst.

Induction of extracellular defense-related proteins in suspension cultured-cells of Daucus carota elicited with cyclodextrins and methyl jasmonate

Suspension cultured-cells (SCC) of Daucus carota were used to evaluate the effect of methyl jasmonate and cyclodextrins, separately or in combination, on the induction of defense responses, particularly the accumulation of pathogenesis-related proteins. A comparative study of the extracellular proteome (secretome) between control and elicited carrot SCC pointed to the presence of amino acid sequences homologous to glycoproteins which have inhibitory activity against the cell-wall-degrading enzymes secreted by pathogens and/or are induced when carrot cells are exposed to a pathogen elicitor. Other amino acid sequences were homologous to Leucine-Rich Repeat domain-containing proteins, which play an essential role in defense against pathogens, as well as in the recognition of microorganisms, making them important players in the innate immunity of this plant. Also, some tryptic peptides were shown to be homologous to a thaumatin-like protein, showing high specificity to abiotic stress and to different reticuline oxidase-like proteins that displayed high levels of antifungal activity, suggesting that methyl jasmonate and cyclodextrins could play a role in mediating defense-related gene product expression in SCC of D. carota. Apart from these elicitor-inducible proteins, we observed the presence of PR-proteins in both control and elicited carrot SCC, suggesting that their expression is mainly constitutive. These PR-proteins are putative class IV chitinases, which also have inhibitory activity against pathogen growth and the class III peroxidases that participate in response to environmental stress (e.g. pathogen attack and oxidative), meaning that they are involved in defense responses triggered by both biotic and abiotic factors.

Identification of upregulated genes in laminarin-treated poplar (Populus alba × P. tremula var. glandulosa) suspension cells by suppression subtractive hybridization and cDNA microarray

Abstract Elicitors trigger defence responses in plant cells through signal transduction pathways, leading to accumulation of pathogenesis-related (PR) proteins and, eventually, pathogen resistance. To understand defence responses of hybrid poplar (Populus alba × P. tremula var. glandulosa), we isolated and characterized upregulated genes in poplar cells by laminarin-induced elicitation using suppression subtractive hybridization (SSH) and cDNA microarray approaches. A total of 1,269 clones in the SSH library were sequenced and a cDNA microarray, containing 265 unique subtracted clones, was fabricated. From the microarray results, 37 clones were found to be upregulated by laminarin treatment and their putative functions are discussed. Genes involved in signal transduction, transcriptional regulation, and phytohormone biosynthesis were upregulated. Other genes encoding PR proteins, peptidases, and an ABC transporter, as well as genes involved in lignification and protein synthesis and turnover, were also identified. Our results suggest that well-organized defence responses, from signal transduction to accumulation of PR proteins, are activated in poplar cells by laminarininduced elicitation and could contribute to resistance against pathogens.

Methyl jasmonate induced defense responses increase resistance to Fusarium oxysporum f. sp. cubense race 4 in banana

Fusarium wilt, caused by the fungal pathogen Fusarium oxysporum, is a severe banana (Musa spp.) root disease, currently threatening banana production worldwide. Methyl jasmonate (MeJA) has been recognized for its ability to induce systemic defense against various pathogens. In this study, we treated banana plantlets with 1.5mM exogenous MeJA. The effects of treatment on the level of chitinase, β-1,3-glucanase, total phenols (TP), hydrogen peroxide (H2O2), superoxide radicals (O2−), malondialdehyde (MDA), as well as the activities of superoxide dismutase (SOD), peroxidase (POD), polyphenol oxidase (PPO), catalase (CAT) and phenylalanine ammonia-lyase (PAL), plus disease incidence (DI) and disease severity (DS) were investigated. The results demonstrated that MeJA regulated the contents of H2O2 and O2−, and decreased MDA in banana roots. Meanwhile, plantlets treated with MeJA exhibited considerably higher chitinase, β-1,3-glucanase and TP contents, and higher activities of SOD, POD, PPO, CAT and PAL. In addition, MeJA treatment dramatically reduced disease incidence (DI) and disease severity (DS) of banana plants compared to control. The higher activities of scavenger antioxidant enzymes account for the reduction of cellular levels of reactive oxygen species (ROS), which is of vital importance for maintaining cell membrane integrity. The accumulation of pathogenesis-related proteins, chitinases and β-1,3-glucanase, as well as total phenols and PAL activity in the treated roots also indicated that MeJA triggered key enzymes of secondary metabolite biosynthetic pathways, suggesting that MeJA was involved in the activation of a disease-related defense system.

Differences in the Induction of Defence Responses in Resistant and Susceptible Muskmelon Plants Infected with Colletotrichum lagenarium

Defence reactions occurring in resistant (cv. Gankezaomi) and susceptible (cv. Ganmibao) muskmelon leaves were investigated after inoculating with Colletotrichum lagenarium. Lesion restriction in resistant cultivars was associated with the accumulation of hydrogen peroxide (H2O2). The activity of antioxidants catalase (CAT) and peroxidase (POD) significantly increased in both cultivars after inoculation, while levels of both CAT and POD activity were significantly higher in the resistant cultivar. Ascorbate peroxidase (APX) increased in both cultivars after inoculation, and level of APX activity was significantly higher in the resistant cultivar. Glutathione reductase (GR) activity significantly increased in both cultivars following inoculation, but was higher in the resistant cultivar, resulting in higher levels of ascorbic acid (AsA) and reduced glutathione (GSH). Phenylalanine ammonia lyase (PAL) significantly increased in inoculated leaves of both cultivars, resulting in higher levels of total phenolic compounds and flavonoids. The pathogenesis-related proteins chitinase (CHT) and β-1, 3-glucanase (GLU) significantly increased following inoculation with higher activity in the resistant cultivar. These findings show that resistance of muskmelon plants against C. lagenarium is associated with the rapid accumulation of H2O2, resulting in altered cellular redox status, accumulation of pathogenesis-related proteins, activation of phenylpropanoid pathway to accumulation of phenolic compounds and flavonoids.

Induction of trans-resveratrol and extracellular pathogenesis-related proteins in elicited suspension cultured cells of Vitis vinifera cv Monastrell

Suspension-cultured cells of Vitis vinifera cv Monastrell were used to investigate the effects of methyljasmonate, ethylene and salicylic acid separately or in combination with cyclodextrins on both trans-resveratrol production and the induction of defense responses.The results showed that the addition of methyljasmonate or ethylene to suspension-cultured cells jointly treated with cyclodextrins and salicylic acid provoked a decrease of trans-resveratrol levels suggesting that salicylic acid has a negative and antagonistic effect with methyljasmonate or ethylene on trans-resveratrol production. Likewise, the exogenous application of these compounds induced the accumulation of pathogenesis-related proteins. Analysis of the extracellular proteome showed the presence of amino acid sequences homologous to an specific β-1,3-glucanase, class III peroxidases and a β-1,4-mannanase, which suggests that these signal molecules could play a role in mediating defense-related gene product expression in V. vinifera cv Monastrell. Apart from these inducible proteins, other proteins were found in both the control and elicited cell cultures of V. vinifera. These included class IV chitinase, polygalacturonase inhibitor protein and reticuline oxidase-like protein, suggesting that their expression is constitutive being involved in the modification of the cell wall architecture during cell culture growth and in the prevention of pathogen attack.