Increase In Polyphenol Oxidase Activity Research Articles

Role of ascorbic acid in the inhibition of polyphenol oxidase and the prevention of browning in different browning-sensitive Lactuca sativa var. capitata (L.) and Eruca sativa (Mill.) stored as fresh-cut produce.

Polyphenol oxidase (PPO) and, to a minor extent, peroxidase (POD) represent the key enzymes involved in enzymatic browning, a negative process induced by cutting fresh-cut produce such as lettuce (Lactuca sativa) and rocket salad (Eruca sativa). Although ascorbic acid is frequently utilised as an anti-browning agent, its mechanism in the prevention of the browning phenomenon is not clearly understood. The activity of PPO and POD and their isoforms in lettuce (a high-browning and low-ascorbic acid species) and rocket salad (a low-browning and high-ascorbic species) was characterised. The kinetic parameters of PPO and in vitro ascorbic acid-PPO inhibition were also investigated. In rocket salad, PPO activity was much lower than that in lettuce and cutting induced an increase in PPO activity only in lettuce. Exogenous ascorbic acid (5 mmol L(-1)) reduced PPO activity by about 90% in lettuce. POD did not appear to be closely related to browning in lettuce. PPO is the main enzyme involved in the browning phenomenon; POD appears to play a minor role. The concentration of endogenous ascorbic acid in rocket salad was related to its low-browning sensitivity after cutting. In lettuce, the addition of ascorbic acid directly inhibited PPO activity. The results suggest that the high ascorbic acid content found in rocket salad plays an effective role in reducing PPO activity.

Expression of a nematode symbiotic bacterium-derived protease inhibitor protein in tobacco enhanced tolerance against Myzus persicae

Fusion proteins of a protease inhibitor from an entomopathogenic nematode symbiotic bacterium (PIN1) and green fluorescent protein (GFP) were expressed in tobacco (Nicotiana tobacum cv. Samsun NN). The PIN1-GFP protein expressed under the control of the CaMV-35S promoter was detected in leaves of transgenic tobacco plants. The effect of PIN1 on anti-pest activity for Myzus persicae was tested by feeding neonate aphids on three independent homozygous lines. For nymphs fed on PIN1-GFP-expressing plants, no effects on insect survival were observed but average insect weight and fecundity were significantly reduced. The aphid biomass was decreased by 30-35% compared to those reared on control plants. The effects of PIN1 on M. persicae were correlated with the decrease of the leucine aminopeptidase and total protease activities of whole insect extracts. Furthermore, an increase in polyphenoloxidase activity was observed in PIN1-GFP-expressing plants. These results revealed that the transgenic expression of PIN1 in tobacco enhanced tolerance against aphids. Key message This study suggests that entomopathogenic nematode symbiotic bacterium is another valuable resource of protease inhibitors which can be engineered into plants for insect pest management.

Influence of nitrogen fertilization on polyphenol oxidase activity in peach fruits

Generally, when nitrogen effects on fruit properties are assessed, different samples harvested at different maturity stages are assayed. This method makes it difficult to investigate the direct effects of fertilization on the different quality parameters without the influence of the ripening stage. In this piece of work, the effects of three levels of nitrogen fertilization (0, 60 and 120kg/ha) on polyphenol oxidase (PPO) activity and consistency of peach fruits at the same maturity stage were studied. The results showed that nitrogen application (regardless the dose) had a direct effect on the studied parameters, causing an increase of PPO activity and also an increase of fruit consistency. However, no significant differences were found between the application of 60 and 120kgN/ha. This enhanced activity of the enzyme may represent a problem in peach postharvest handling and processing.

In situ Inactivation of Polyphenol Oxidase in Mamey Fruit (Pouteria sapota) by Microwave Treatment

Polyphenol oxidase (PPO) is the enzyme responsible for quality loss in most fruits and vegetables. Quality loss is mainly because of oxidative chemical reactions which generate the darkening of tissues. Mamey fruit (Pouteria sapota) after harvesting suffers a rapid quality decay trough activation of PPO. However, PPO may be inactivated in situ by chemical or thermal treatment. In food processing, microwave treatment (MT) has been used recently as an alternative for PPO inactivation. In this study, it was observed that mamey fruit pulp subjected to a gently MT resulted in a higher PPO activity as the generated heat induced in situ the increase in PPO activity. In contrast, PPO was completely inactivated after long MT by using a high microwave power. Temperature in mamey pulp after MT reached a maximum of 79 °C; although PPO was active up to 60 °C. PPO was completely inactivated when conventional blanching treatment was performed but required a higher temperature (92 °C/300 s). The optimum energy intensity (E(opt)) for PPO inactivation by MT was 0.51 kJ/g or 937 W/165 s. Under this condition, the remaining PPO activity was inversely proportional to energy intensity (E). Interestingly, MT resulted in a negligible damage in microstructure of mamey pulp, although blanching treatment resulted in large damaging effects on tissue organization and shape. Therefore, MT is proposed as an effective way to completely inactivate PPO without causing any significant damage to fruit tissues and shape; as preservation of color, flavor, and taste would be favored.

A novel postharvest UV-C treatment to reduce chilling injury (membrane damage, browning and chlorophyll degradation) in banana peel

To investigate the mode of action of UV-C to alleviate CI in banana. Banana [Musa (AAA group), Cavendish subgroup cv. Cavendish] fruits were treated with UV-C at dosages of 0.03 kJ m −2 prior to storage at 8 or 25 °C. UV-C treatment reduced both the incidence and severity of banana fruits stored under low temperature. UV-C treatment reduced membrane damage indicated by lower activity of lipoxygenase (LOX) and malondialdehyde (MDA) content. In additions, the in vitro polyphenol oxidase (PPO) activity was activated when fruits were stored at CI temperature and UV-C treatment could inhibit the PPO activity. UV-C treatment also delayed yellowing and chlorophyll (Chl) degradation due to the inhibition of chlorophyllase and chl-degrading peroxidase activities. Furthermore, the reduction of ethylene production and respiration rate by UV-C treatment results in extended postharvest shelf life of banana. These finding suggest that the loss of cellular compartments in consequence of membrane degradation, combined with the increase of PPO activity, might contribute to the development of CI in banana peel. UV-C treatment may play an important role in maintained membrane integrity and inhibited PPO activity, reducing the severity of CI symptom and delayed ripening in banana. This novel technique may offer an advance in postharvest handling of bananas and other chilling-sensitive commodities in order to reduce postharvest losses resulting from CI.

Induction of Polyphenol Oxidase Activity in Dormant Wild Oat (Avena fatua) Seeds and Caryopses: A Defense Response to Seed Decay Fungi

Persistence of the soil seed bank requires both dormancy and resistance to seed decay organisms. However, there is little or no information evaluating biochemical responses of dormant weed seeds to pathogens. Wild oat caryopses were incubated with four pathogenic fungal isolates to evaluate the response of the pathogen defense enzyme, polyphenol oxidase (PPO). Caryopsis PPO activity was induced by threeFusariumspp. isolates previously obtained from whole seeds incubated in the field whereas caryopsis PPO activity was decreased by aPythiumisolate.Fusarium avenaceumisolateF.a.1 caused the greatest PPO induction and was studied in more detail. When whole wild oat seeds were incubated onF.a.1, PPO activity was induced in seeds, hulls (lemma and palea), and caryopses. Incubation of whole seeds onF.a.1 gradually induced caryopsis PPO activity over an 8-d period, whereas incubation of caryopses onF.a.1 over a 4-d period caused a greater and more rapid induction of PPO activity. Very little PPO activity could be leached from untreated caryopses, but nearly all of the induced PPO activity inF.a.1-treated caryopses was readily leached when incubated in buffer. In Western blots, both untreated andF.a.1-treated leachates contained a ∼57-kilodalton (kD) protein, putatively the mature and relatively inactive form of PPO. However, lower molecular weight antigenic proteins between ∼52 and ∼25 kD were strongly induced inF.a.1-treated caryopses, with this induction being correlated with the increase in PPO activity. We hypothesize that dormant weed seeds possess biochemical defenses against pathogens and, more specifically, that proteolysis in the presence of fungal pathogens may release an activated form of PPO from the surface of wild oat caryopses and hulls.

Enhanced stilbene production in cell cultures of Cayratia trifolia through co-treatment with abiotic and biotic elicitors and sucrose

This report demonstrates the elicitation effect on growth and stilbene accumulation in cell cultures of Cayratia trifolia (Vitaceae) by an extract of the angiosperm parasite Cuscuta reflexa and salicylic acid in combination with sucrose feeding. Cell cultures of C. trifolia, a tropical liana, were maintained in liquid Murashige and Skoog's basal medium containing 0.25 mg l−1 naphthalene acetic acid, 0.2 mg l−1 kinetin with 3% sucrose and 250 mg l−1 casein hydrolysate. The cells treated with Cuscuta elicitor showed increased polyphenol oxidase activity with increasing concentration of the elicitor, while total phenol content remained almost unchanged. Enhanced yield of stilbenes (∼8-fold) was recorded in the cells treated with 200 mg l−1 Cuscuta elicitor for 7 d. Optimum accumulation of stilbenes with a non-significant decrease in cell growth as compared with control was recorded with the addition of 3% sucrose on the seventh day of cell culture. Addition of 3% sucrose with salicylic acid at 500 μM and Cuscuta extract at 200 mg l−1 on the seventh day enhanced total stilbene yield up to 50.1 mg l−1, which was ∼14-fold higher than in control cultures. Piceid content increased ∼200-fold in such cultures.

Polyphenol oxidase activity, phenolic acid composition and browning in cashew apple ( Anacardium occidentale, L.) after processing

This study describes the extraction and characterisation of cashew apple polyphenol oxidase (PPO) and the effect of wounding on cashew apple phenolic acid composition, PPO activity and fruit browning. Purification factor was 59 at 95% (NH 4) 2SO 4 saturation. For PPO activity, the optimal substrate was catechol and the optimum pH was 6.5. PPO K m and V max values were 18.8 mM and 13.6 U min −1 ml −1, respectively. Ascorbic acid, citric acid, sodium sulphite and sodium metabisulphite decreased PPO activity, while sodium chloride increased PPO activity. Wounding at 2 °C and 27 °C for 24 h increased PPO activity but storage at 40 °C reduced PPO activity. Gallic acid, protocatechuic acid and cinnamic acid (free and conjugate) were identified in cashew apple juice. Cutting and subsequent storage at 40 °C hydrolysed cinnamic acid. 5-Hydroxymethylfurfural content in cashew apple juice increased after injury and storage at higher temperatures, indicating non-enzymatic browning.

Effects of pulsed light treatments on quality and antioxidant properties of fresh-cut mushrooms ( Agaricus bisporus)

This study investigated the impact of pulsed light treatments on microbial quality, enzymatic browning, texture and antioxidant properties of fresh-cut mushrooms. The reduction of the native microflora of sliced mushrooms ranged from 0.6 to 2.2 log after 15 days of refrigerated storage by flashing at 4.8, 12 and 28 J cm −2. Pulsed light treatments allowed extension of the microbiological shelf life of fresh-cut mushrooms by 2–3 days in comparison to untreated samples, while providing a high quality product. The use of high pulsed light fluencies (12 and 28 J cm −2) dramatically affected the texture of sliced mushrooms due to thermal damage induced by the treatments. Enzymatic browning was also promoted by an increase in polyphenol oxidase activity when the highest dose of pulsed light was applied. At 28 J cm −2, phenolic compounds, vitamin C and antioxidant capacity were significantly reduced. Our results suggest that the application of pulsed light at doses of 4.8 J cm −2 could extend the shelf life of fresh-cut mushrooms without dramatically affecting texture and antioxidant properties.

Cloning, microbial expression and structure–activity relationship of polyphenol oxidases from Camellia sinensis

Polyphenol oxidase (PPO) can be used for organic synthesis and degradation of wastes and dyes in industries. Lack of enzyme sources is a major barrier for its application. A PPO gene, with a full length of 1.8 kb without introns, was cloned by PCR from genomic DNA of five common cultivars of Camellia sinensis. They had a 98.2–99.9% degree of identity in nucleotides and 94.7–96.1% in amino acids and encoded a polypeptide of 599 amino acids with a signal peptide targeting the chloroplast and three Cu-binding domains. The mature PPO showed high expression and enzyme activity after refolding the inclusion bodies in Escherichia coli BL21 (DE3) using pET30c expression vector, but low expression in Pichia pastoris GS115 using both the secretory and non-secretory vectors pPICZαA and pPICZA. The expression of PPO mutants demonstrated that the signal sequences prevented recombinant gene expression in E. coli. PPO activity was not affected by the C-terminus and was slightly inhibited by the CuC domain. Other domains were important for its activity. A 3.1-fold increase in PPO activity over non-recombinant controls was obtained by expressing the PPO fragment without signal sequences and the CuC domain in E. coli BL21 (DE3) using the pET30c vector.

Defense mechanisms of Solanum tuberosum L. in response to attack by plant-pathogenic bacteria.

The natural resistance of plants to disease is based not only on preformed mechanisms, but also on induced mechanisms. The defense mechanisms present in resistant plants may also be found in susceptible ones. This study attempted to analyze the metabolic alterations in plants of the potato Solanum tuberosum L. cv. Agata that were inoculated with the incompatible plant-pathogenic bacteria X. axonopodis and R. solanacearum, and the compatible bacterium E. carotovora. Levels of total phenolic compounds, including the flavonoid group, and the activities of polyphenol oxidase (PPO) and peroxidase (POX) were evaluated. Bacteria compatibility was evaluated by means of infiltration of tubers. The defense response was evaluated in the leaves of the potato plants. Leaves were inoculated depending on their number and location on the stem. Multiple-leaf inoculation was carried out on basal, intermediate, and apical leaves, and single inoculations on intermediate leaves. Leaves inoculated with X. axonopodis and with R. solanacearum showed hypersensitive responses within 24 hours post-inoculation, whereas leaves inoculated with E. carotovora showed disease symptoms. Therefore, the R. solanacearum isolate used in the experiments did not exhibit virulence to this potato cultivar. Regardless of the bacterial treatments, the basal leaves showed higher PPO and POX activities and lower levels of total phenolic compounds and flavonoids, compared to the apical leaves. However, basal and intermediate leaves inoculated with R. solanacearum and X. axonopodis showed increases in total phenolic compounds and flavonoid levels. In general, multiple-leaf inoculation showed the highest levels of total phenolics and flavonoids, whereas the single inoculations resulted in the highest increase in PPO activity. The POX activity showed no significant difference between single- and multiple-leaf inoculations. Plants inoculated with E. carotovora showed no significant increase in defense mechanisms such as enzyme activity and phenolic compounds. Therefore, resistance or susceptibility in S. tuberosum cv. Agata might depend on leaf age, type of inoculation performed (single or multiple), and the interaction between plant and pathogen.

Inactivation kinetics of apple polyphenol oxidase in different pressure–temperature domains

The impact of high hydrostatic pressure and temperature on the stability of polyphenol oxidase (PPO) was studied in cloudy apple juice. Application of 200–500 MPa near room temperature or heat treatment at 45–55 °C at ambient pressure caused an increase of PPO activity of up to 65% in freshly squeezed apple juice. Combined pressure–temperature inactivation experiments with fully activated PPO (5 min treatment at 400 MPa and 20 °C) were carried out in the range of 0.1–700 MPa and 20–80 °C. Enzyme inactivation kinetics followed a 2.2 order reaction scheme at all pressure–temperature conditions tested. A polynomial model was successfully applied to describe the rate of PPO inactivation as a function of pressure and temperature and was used to construct a pressure–temperature isokinetic diagram. This diagram clearly showed synergistic effects of pressure and temperature on the inactivation of apple PPO at pressures above 300 MPa and antagonistic effects at lower pressures. Compared to ambient pressure conditions, temperatures required to inactivate PPO in apple juice were increased 10–15 °C at 100–300 MPa. High pressure processing of fresh fruits is gaining popularity in the food industry because of its ability to inactivate microorganisms and some enzymes near room temperature with little impact on flavour or nutritional attributes of the food. However, quantitative data regarding the impact of process parameters on the target reaction are required to economically utilise this technology. This paper provides a mathematical model describing the combined effect of pressure, temperature and treatment time on the inactivation of PPO in cloudy apple juice.

Constitutive and Induced Activities of Defense-Related Enzymes in Aphid-Resistant and Aphid-Susceptible Cultivars of Wheat

Phenylalanine ammonia-lyase (PAL), polyphenol oxidase (PPO), and peroxidase (POD) are considered important biochemical markers in host plant resistance against pest insects. Constitutive activity of these enzymes was analyzed in resistant and susceptible wheat cultivars against cereal aphid Sitobion avenae (F.) at various developmental stages, i.e., tillering, stem elongation, flag leaf, and ear. Following aphid infestation, the activity of these enzymes was determined at the flag leaf and ear stages. Resistant cultivars exhibited greater constitutive PAL activity than susceptible ones at the tillering, stem elongation, and flag leaf stages. Aphid infestation enhanced levels of PAL activity in the flag leaf and ear stages in both resistant and susceptible cultivars. Constitutive PPO activity was higher in the resistant cultivars at all developmental stages. Aphid infestation induced increases in PPO activity in the flag leaf and ear stages of one susceptible cultivar, whereas induction in resistant cultivars was weaker. Resistant cultivars showed greater constitutive POD activity in the tillering, stem elongation, and flag leaf stages, while aphid infestation induced POD activity in all cultivars, especially in susceptible ones. The potential role of PAL, PPO, and POD in wheat defense against aphid infestation is discussed.

Defense mechanisms of Solanum tuberosum L. in response to attack by plant-pathogenic bacteria

The natural resistance of plants to disease is based not only on preformed mechanisms, but also on induced mechanisms. The defense mechanisms present in resistant plants may also be found in susceptible ones. This study attempted to analyze the metabolic alterations in plants of the potato Solanum tuberosum L. cv. Agata that were inoculated with the incompatible plant-pathogenic bacteria X. axonopodis and R. solanacearum, and the compatible bacterium E. carotovora. Levels of total phenolic compounds, including the flavonoid group, and the activities of polyphenol oxidase (PPO) and peroxidase (POX) were evaluated. Bacteria compatibility was evaluated by means of infiltration of tubers. The defense response was evaluated in the leaves of the potato plants. Leaves were inoculated depending on their number and location on the stem. Multiple-leaf inoculation was carried out on basal, intermediate, and apical leaves, and single inoculations on intermediate leaves. Leaves inoculated with X. axonopodis and with R. solanacearum showed hypersensitive responses within 24 hours post-inoculation, whereas leaves inoculated with E. carotovora showed disease symptoms. Therefore, the R. solanacearum isolate used in the experiments did not exhibit virulence to this potato cultivar. Regardless of the bacterial treatments, the basal leaves showed higher PPO and POX activities and lower levels of total phenolic compounds and flavonoids, compared to the apical leaves. However, basal and intermediate leaves inoculated with R. solanacearum and X. axonopodis showed increases in total phenolic compounds and flavonoid levels. In general, multiple-leaf inoculation showed the highest levels of total phenolics and flavonoids, whereas the single inoculations resulted in the highest increase in PPO activity. The POX activity showed no significant difference between single- and multiple-leaf inoculations. Plants inoculated with E. carotovora showed no significant increase in defense mechanisms such as enzyme activity and phenolic compounds. Therefore, resistance or susceptibility in S. tuberosum cv. Agata might depend on leaf age, type of inoculation performed (single or multiple), and the interaction between plant and pathogen.

Combined effects of chemical dip and/or carrageenan coating and/or controlled atmosphere on quality of fresh-cut banana

The combined effect of chemical dip and/or edible coating and/or controlled atmosphere (CA) on quality of fresh-cut banana was investigated. Banana slices were subject to a 3-min dip into a solution containing 1% (w/v) calcium chloride, 0.75% (w/v) ascorbic acid and 0.75% (w/v) cysteine and/or combined with a carrageenan coating and/or combined with controlled atmosphere (3% O 2 + 10% CO 2). Physico-chemical and microbiological qualities were evaluated during 5 days of storage at 5 °C. Dip combined with CA treatment prevented product weight loss and increase of polyphenol oxidase activity during the 5 days of storage. Colour, firmness, pH, tritatable acidity and total soluble solids values and total phenolic content presented the smallest changes. Microbial analysis showed that minimally processed bananas were within the acceptable limits during 5 days of storage at 5 °C.

Ativação de defesa em cacaueiro contra a murcha-de-verticílio por extratos naturais e acibenzolar-S-metil

O objetivo deste trabalho foi avaliar o potencial de extratos fúngicos e vegetais na redução da murcha-de-verticílio do cacaueiro, as atividades da peroxidase e polifenoloxidase e o conteúdo de lignina. Mudas de cacaueiro foram pulverizadas com filtrado de micélio de Rhizopus sp. (FMR), quitosana de Rhizopus sp. (QMR) e Trichoderma sp. (QMT), extratos de casca in natura e seca de maracujá, extrato metanólico de casca seca de frutos de maracujá (MMS) e acibenzolar-S-metil (ASM - 0.2 mg mL-1) e sete dias depois, submetidas à inoculação de Verticillium dahliae. O ASM reduziu a murcha-de-verticílio em 38,0%, seguido dos extratos FMR, QMT, MMS e QMR, que apresentaram reduções em 22,8, 20,1, 19,2 e 15,7%, respectivamente, em relação à testemunha. Plantas pulverizadas com ASM ou FMR seguidas de inoculação apresentaram aumento da atividade de peroxidase aos oito dias após a pulverização, comparadas às respectivas testemunhas, com pico aos 18 dias após a pulverização. ASM e FMR aumentaram a atividade de polifenoloxidase aos quatro dias após a pulverização. Maiores concentrações de lignina foram obtidas em plantas tratadas com FMR e FMR seguido de inoculação. FMR é um potencial indutor de resistência para manejo de murcha-de-verticílio em cacaueiro.

Does Secondary Plant Metabolism Provide a Mechanism for Plant Defenses in the Tropical Soda Apple Solanum Viarum (Solanales: Solanaceae) against Spodoptera exigua and S. eridania (Lepidoptera: Noctuidae)?

Abstract Survival assays were conducted with beet armyworm Spodoptera exigua (Hubner) and southern armyworm S. eridania (Stoll) with tropical soda apple Solanum viarum Dunal, a relative of tomato. In addition, polyphenol oxidase (PPO) enzyme assays were conducted to determine if plant defense compounds are being produced by tropical soda apple in response to herbivory. Both S. exigua and S. eridania induced plant defenses in tropical soda apple. Significantly more S. exigua and S. eridania neonate larvae survived to 2nd instar on non-induced plants and artificial diet when compared with plants with induced defenses. Tropical soda apple plants fed on by S. exigua and S. eridania had significantly increased PPO activity.

Application of magnetic field and iron in order to change medicinal products of Ocimum basilicum

In the present research, the effects of static magnetic field, with or without iron on the growth, the activity of certain enzymes (like polyphenol oxidase and phenyle alanine ammonialyase), the content of phenolic compounds and the essential oil composition of sweet basil (Ocimum basilicum) were studied. Treatment of plants with static magnetic field with or without iron led to decrease of the growth of plants as well as decrease in the activity of phenyle alanine ammonialyase and of phenolic compounds content. In addition in those plants exposed to magnetic field the amount of essential oils, notably of methyl chavicol, was increased. Exposure of plants to the magnetic field decreased the activity of polyphenol oxidase, whereas treatment of plants with iron when exposed to magnetic field increased polyphenol oxidase activity. In this group the content of essential oil was reduced.

(65) Jasmonate-induced Changes in Polyphenol Oxidase, Peroxidase, and Proteinase Inhibitors in Horticultural Species

Jasmonates are a class of plant hormones involved in plant defense and stress responses. For example, jasmonate-induced defense responses in Lycopersicon esculentum include increases in activity of proteinase inhibitors, polyphenol oxidases, and peroxidases. As part of our efforts to reduce or control greenhouse pest infestations, we hypothesized that methyl jasmonate (MeJA) could induce these biochemical changes in common greenhouse crops. We studied Impatiens wallerana `Super Elfin Pink', L. esculentum `Big Boy', Petunia ×hybrida `Bravo Lavendar', Viola ×wittrockiana `Imperial Beaconsfield', Coleus ×hybridus `Wizard Jade', Nicotiana alata `Saratoga Lime', Pelargonium ×hortorum `Pinto Pink', and Tagetes erecta `Antigua Primrose'. Polyphenol oxidase and peroxidase activity was studied in the first four species, and proteinase inhibitors were studied in all eight. We sprayed plants with 0, 5 × 10-6, or 10-4 molar MeJA and made measurements after 24 hours. We detected a small increase in polyphenol oxidase activity of plants treated with 10-4 molar MeJA; 5 × 10-6 molar had no effect, and L. esculentum had the highest polyphenol oxidase activity. Peroxidase activity was not affected by MeJA. I. wallerana had the highest peroxidase activity, L. esculentum and V. ×wittrockiana had the lowest. 5 × 10-6 molar MeJA increased proteinase inhibitor activity in most species, and 10-4 molar increased activity in every species except P. ×hortorum.

LePLDβ1 activation and relocalization in suspension‐cultured tomato cells treated with xylanase

Phospholipase D (PLD) has been implicated in various cellular processes including membrane degradation, vesicular trafficking and signal transduction. Previously, we described a PLD gene family in tomato (Lycopersicon esculentum) and showed that expression of one of these genes, LePLDbeta1, was induced by treatment with the fungal elicitor xylanase. To further investigate the function of this PLD, a gene-specific RNAi construct was used to knock down levels of LePLDbeta1 transcript in suspension-cultured tomato cells. Silenced cells exhibited a strong decrease in xylanase-induced PLD activity and responded to xylanase treatment with a disproportionate oxidative burst. Furthermore, LePLDbeta1-silenced cell-suspension cultures were found to have increased polyphenol oxidase activity, to secrete less of the beta-d-xylosidase LeXYL2 and to secrete and express more of the xyloglucan-specific endoglucanase inhibitor protein XEGIP. Using an LePLDbeta1-green fluorescent protein (GFP) fusion protein for confocal laser scanning microscopy-mediated localization studies, untreated cells displayed a cytosolic localization, whereas treatment with xylanase induced relocalization to punctuate structures within the cytosol. Possible functions for PLDbeta in plant-pathogen interactions are discussed.