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PubMed2006

Comparing contributions of soil versus root colonization to variations in stomatal behavior and soil drying in mycorrhizal Sorghum bicolor and Cucurbita pepo.

In prior studies we learned that colonization of soil can be as important as colonization of roots in determining mycorrhizal influence on the water relations of host plants. Here we use a path analysis modeling approach to test (a) whether quantity of hyphae in soil contributes to variations in stomatal behavior and soil drying, and (b) whether soil colonization or root colonization has a stronger influence on these stomatal and soil drying responses. Experiments were performed on Sorghum bicolor and Cucurbita pepo, with soils and roots colonized by a mixture of Glomus intraradices and Gigaspora margarita. Soil colonization generally made more significant contributions to stomatal conductance than did root colonization. Soil colonization did not make significant direct contributions to soil water potential measures (soil water potential at stomatal closure or soil drying rate), whereas root colonization did contribute a potentially important path to each. The findings further support a role for mycorrhization of the soil itself in contributing to the regulation of stomatal behavior of host plants.

Cucurbita↗
PubMed2005

Influence of UV-B exclusion and selenium treatment on photochemical efficiency of photosystem II, yield and respiratory potential in pumpkins (Cucurbita pepo L.).

The influence of ambient and filtered solar ultraviolet-B (UV)-B radiation and of selenium treatment was determined on photochemical efficiency, respiratory potential measured by electron transport system (ETS) activity, and yield in pumpkins, Cucurbita pepo L. Yield and ETS activity were higher when solar UV-B was filtered out. The results suggested that the decreased yield was related to the UV-B impaired flow of electrons in the respiratory chain. Selenium increased yield under ambient radiation conditions. However, no significant effect of excluding UV-B radiation or of treatment with selenium was observed on the photochemical efficiency of photosystem II (PSII).

Cucurbita↗
PubMed2002

Kinetic mechanism and order of substrate binding for sn-glycerol-3-phosphate acyltransferase from squash (Cucurbita moschata).

sn-Glycerol-3-phosphate acyltransferase (G3PAT, EC 2.3.1.15), a component of glycerolipid biosynthesis, is an important enzyme in chilling sensitivity in plants. The three-dimensional structure of the enzyme from squash (Cucurbita moschata), without bound substrate, has been determined [Turnbull et al. (2001) Acta Crystallogr. D 57, 451-453; Turnbull et al. (2001) Structure 9, 347-353]. Here we report the kinetic mechanism of plastidial G3PAT from squash and the order of substrate binding using acyl-acyl carrier protein (acyl-ACP) substrates. The reaction proceeds via a compulsory-ordered ternary complex with acyl-ACP binding before glycerol-3-phosphate. We have also determined that the reaction will proceed with C(4:0)-CoA, C(6:0)-CoA and C(12:0)-ACP substrates, allowing a wider choice of acyl groups for future co-crystallisation studies.

Acyl Carrier Protein↗
PubMed2003

A defect in the yeast plasma membrane urea transporter Dur3p is complemented by CpNIP1, a Nod26-like protein from zucchini (Cucurbita pepo L.), and by Arabidopsis thaliana delta-TIP or gamma-TIP.

Dur3 encodes the yeast plasma membrane urea transporter and Deltadur3 mutants are unable to grow on media containing low concentrations of urea as sole nitrogen source. Complementation of the Deltadur3 mutant line with expression libraries generated from whole Arabidopsis thaliana seedlings or from zucchini (Cucurbita pepo L.) vascular tissue yielded numerous lines that had regained the capacity to grow on low urea as sole nitrogen source. Analysis of several of these yeast lines revealed that the Deltadur3 mutation was complemented either by delta-TIP (TIP=tonoplast intrinsic protein) or gamma-TIP from Arabidopsis or by CpNIP1, a new NOD26-like protein from zucchini. delta-TIP (At3g16240) and gamma-TIP (At2g36830) had previously been characterized as proteins facilitating the transport of water across the tonoplast membrane, and Nod26-like proteins were characterized as glycerol transporters. So far, transport of urea has not been described for any of the proteins described in this paper. Further analyses support this function of TIPs and nodulin 26-like intrinsic proteins in urea transport.

Amino Acid Sequence↗
PubMed2003

Metabolic networks of Cucurbita maxima phloem.

Metabolomic analysis aims at a comprehensive characterization of biological samples. Yet, biologically meaningful interpretations are often limited by the poor spatial and temporal resolution of the acquired data sets. One way to remedy this is to limit the complexity of the cell types being studied. Cucurbita maxima Duch. vascular exudates provide an excellent material for metabolomics in this regard. Using automated mass spectral deconvolution, over 400 components have been detected in these exudates, but only 90 of them were tentatively identified. Many amino compounds were found in vascular exudates from leaf petioles at concentrations several orders of magnitude higher than in tissue disks from the same leaves, whereas hexoses and sucrose were found in far lower amounts. In order to find the expected impact of assimilation rates on sugar levels, total phloem composition of eight leaves from four plants was followed over 4.5 days. Surprisingly, no diurnal rhythm was found for any of the phloem metabolites that was statistically valid for all eight leaves. Instead, each leaf had its own distinct vascular exudate profile similar to leaves from the same plant, but clearly different from leaves harvested from plants at the same developmental stage. Thirty to forty per cent of all metabolite levels of individual leaves were different from the average of all metabolite profiles. Using metabolic co-regulation analysis, similarities and differences between the exudate profiles were more accurately characterized through network computation, specifically with respect to nitrogen metabolism.

Circadian Rhythm↗
PubMed2003

Expression of plastid-encoded photosynthetic genes during chloroplast or chromoplast differentiation in Cucurbitae pepo L. fruits.

The objective of the study was to determine the patterns of expression of two photosynthetic genes rbcL and psbA, during chloroplast and chromoplast differentiation in fruit tissues of three Cucurbitae pepo L. cultivars: Early Prolific, Foodhook Zucchini and Bicolor Gourds. In two Early Prolific isogenic lines, YYBB and YYB+B+, the steady-state amounts of rbcL and psbA transcripts increased with fruit development upto 14 days post-pollination. The YYB+B+ line in which chloroplast differentiates into chromoplast at about pollination, did not show significantly higher amounts of both transcripts compared to YYBB, in which chromoplast develops early prior to pollination. In the Bicolor Gourds, in which the chromoplast and chloroplast containing tissues lie in juxtaposition on the same fruit, showed little differences in rbcL and psbA transcripts between the two tissues, if any the chromoplast containing tissue contained more of both transcripts than the chloroplast containing tissue. In Fordhook Zucchini fruits, where the chloroplast containing tissue developed early prior to pollination and was maintained, the steady-state amounts of rbcL transcripts increased to a maximum at 3 days post-pollination and levelled at 14 and 21 days post-pollination. In contrast, in Fordhook Zucchini fruits, the psbA transcript increased gradually up to 21 days post-pollination. In Fordhook Zucchini, the apparent ratios of psbA transcripts versus rbcL transcripts ranged from 2.5 to 3.9, at day 3 to 21 post-pollination, while in Bicolor Gourds were 2.9 and 4.5 at days 14 and 21 post-pollination. The two photosynthetic genes, psbA and rbcL were developmentally regulated and differentially expressed. However, their expression in chloroplast containing fruit tissues was not higher than in the chromoplast containing fruit tissues.

Blotting, Northern↗
PubMed2002

Evaluation of the hypoglycemic effect of Cucurbita ficifolia Bouché (Cucurbitaceae) in different experimental models.

Acute hypoglycemic effects of freeze-dried juice of Cucurbita ficifolia Bouché (Cucurbitaceae) fruits were studied in healthy and alloxan-diabetic mice. C. ficifolia fruit administered by intraperitoneal route produced, in a dose-dependent manner, a significant decrease of the glycemia in healthy mice. Although oral route of C. ficifolia fruit juice also caused significant reductions of blood glucose levels in healthy mice, the effect was minor. The juice administered by intraperitoneal route showed an acute hypoglycemic effect in alloxan-diabetic mice. In addition, daily oral administration of this preparation showed a highly significant reduction of the glycemia after 14 days of treatment. Freeze- dried juice caused acute toxicity when administered intraperitoneally, and also when it was administered daily by the oral route.

Administration, Oral↗
PubMed2002

Correlation of binding-loop internal dynamics with stability and function in potato I inhibitor family: relative contributions of Arg(50) and Arg(52) in Cucurbita maxima trypsin inhibitor-V as studied by site-directed mutagenesis and NMR spectroscopy.

The side chains of Arg(50) and Arg(52) at positions P(6)' and P(8)', respectively, anchor the binding loop to the protein scaffold by means of hydrogen bonds in Cucurbita maxima trypsin inhibitor-V (CMTI-V), a potato I family member. Here, we have investigated the relative contributions of Arg(50) and Arg(52) to the binding-loop flexibility and stability by determining changes in structure, dynamics, and proteolytic stability as a consequence of individually mutating them into an alanine. We have compared chemical shift assignments of main-chain hydrogens and nitrogens, and (1)H-(1)H interresidue nuclear Overhauser effects (NOEs) for the two mutants with those of the wild-type protein. We have also measured NMR longitudinal and transverse relaxation rates and (15)N-(1)H NOE enhancements for all backbone and side-chain NH groups and calculated the model-free parameters for R50A-rCMTI-V and R52A-rCMTI-V. The three-dimensional structures and backbone dynamics of the protein scaffold region remain very similar for both mutants, relative to the wild-type protein. The flexibility of the binding loop is increased in both R50A- and R52A-rCMTI-V. In R52A-rCMTI-V, the mean generalized order parameter ( ) of the P(6)-P(1) residues of the binding loop (39-44) decreases to 0.68 +/- 0.02 from 0.76 +/- 0.04 observed for the wild-type protein. However, in R50A-rCMTI-V, the flexibility of the whole binding loop increases, especially that of the P(1)'-P(3)' residues (45-47), whose value drops dramatically to 0.35 +/- 0.03 from 0.68 +/- 0.03 determined for rCMTI-V. More strikingly, S(2) values of side-chain N epsilon Hs reveal that, in the R50A mutant, removal of the R50 hydrogen bond results in the loss of the R52 hydrogen bond too, whereas in R52A, the R50 hydrogen bond remains unaffected. Kinetic data on trypsin-catalyzed hydrolysis of the reactive-site peptide bond (P(1)-P(1)') suggest that the activation free energy barrier of the reaction at 25 degrees C is reduced by 2.1 kcal/mol for R50A-rCMTI-V and by 1.5 kcal/mol for R52A-rCMTI-V, relative to rCMTI-V. Collectively, the results suggest that although both the P(6') and P(8)' anchors are required for optimal inhibitor function and stability in the potato I family, the former is essential for the existence of the latter and has greater influence on the binding-loop structure, dynamics, and stability.

Arginine↗
PubMedDate not supplied

Kinetic and thermodynamic investigation on ascorbate oxidase activity and stability of a Cucurbita maxima extract.

The kinetic and thermodynamic properties of ascorbate oxidase (AO) activity and stability of a Cucurbita maxima extract were investigated. Activity tests performed at 25 degrees C using initial ascorbic acid concentration in the range 50-750 M allowed estimating the Michaelis constant for this substrate (Km = 126 microM) and the maximum initial rate of ascorbic acid oxidation (A0,max = 1.57 mM min-1). The main thermodynamic parameters of the enzyme reaction (DeltaH* = 10.3 kJ mol-1; DeltaG* = 87.2 kJ mol-1; DeltaS* = -258 J mol-1 K-1) were estimated through activity tests performed at 25-48 C. Within such a temperature range, no decrease in the initial reaction rate was detected. The long-term thermostability of the raw extract was then investigated by means of residual activity tests carried out at 10-70 degrees C, which allowed estimating the thermodynamic parameters of the irreversible enzyme inactivation as well (DeltaH*D = 51.7 kJ mol-1; DeltaG*D = 103 kJ mol-1; S*D = -160 J mol-1 K-1). Taking into account the specific rate of AO inactivation determined at different temperatures, we also estimated the enzyme half-life (1047 min at 10 degrees C and 21.2 min at 70 degrees C) and predicted the integral activity of a continuous system using this enzyme preparation. This work should be considered as a preliminary attempt to characterize the AO activity of a C. maxima extract before its concentration by liquid-liquid extraction techniques.

Ascorbate Oxidase↗
PubMed2003

Subspecies-level variation in the phytoextraction of weathered p,p'-DDE by Cucurbita pepo.

Previous studies indicate that Cucurbita pepo can phytoextract highly weathered persistent organic pollutants (POPs) from soil and translocate large quantities to aerial tissues. To investigate intraspecific variability in uptake potential, a field study was conducted to quantify the phytoextraction of weathered p,p'-DDE by 21 cultivar varieties of summer squash from two distinct subspecies, C. pepo ssp texana and C. pepo ssp pepo. Significant differences exist between the two subspecies, with average root and stem to soil bioconcentration factors (BCF, dry weight ratio of contaminant concentration in the vegetation to that in the soil) of 7.22 and 5.40 for ssp pepo and of 2.37 and 0.454 for spp texana, respectively. The amounts of weathered p,p-DDE extracted from the soil by ssp pepo and ssp texana were 0.301 and 0.065%, respectively, with maximum values within each subspecies of 0.780 and 0.182%, respectively. The quantities of 14 inorganic elements were determined in both the soil and tissues (roots, stems, leaves, and fruit) of all 21 cultivar varieties. Phosphorus concentrations in the tissues of ssp pepo were 14 (fruit)-73% (stems) greater than those of ssp texana. These data support our hypothesis that the unique ability of certain cultivars of C. pepo to phytoextract highly weathered POPs from soil is the result of low molecular weight organic acid exudation as a unique phosphorus acquisition mechanism.

Biodegradation, Environmental↗
PubMed2005

Dual role of the pigmentation gene B in affecting carotenoid and vitamin E content in squash (Cucurbita pepo) mesocarp.

The yellow and orange colorations of the mesocarp of pumpkins and squash of Cucurbita pepo are due to the presence of carotenoid pigments that also greatly contribute to their nutritional value. Carotenoids, as well as tocopherols (vitamin E), are isoprenoid compounds formed by branches of a common biosynthetic pathway. Photodiode array HPLC analysis was used to simultaneously determine the content and composition of fruit-flesh carotenoids and tocopherols in five pairs of near-isogenic lines differing in the allelic state of genes previously identified as having profound effects on fruit color. The dominant B allele promoted carotenoid accumulation up to 5-fold and prevented tocopherol accumulation in all genetic backgrounds. The dominant L-2 allele doubled carotenoid content and, in combination with the dominant B allele, increased carotenoid content by 10-15-fold as compared to the recessive l-2 allele. The genes D and L-1 had no significant effects on mesocarp tocopherols or carotenoids content. These results indicate that the B gene, which affects both carotenoids and tocopherols, may play a regulatory role in the flux of the isoprenoid pathway products.

Carotenoids↗
PubMed2005

Cucurbitosides F-M, acylated phenolic glycosides from the seeds of Cucurbita pepo.

Eight new phenolic glycosides, cucurbitosides F-M (1-8), were isolated from the seeds of Cucurbita pepo. Their structures were elucidated as 4-(2-hydroxyethyl)phenyl 5-O-(2-S-2-methylbutyryl)-beta-d-apiofuranosyl(l-->2)-beta-d-glucopyranoside (1), 4-(2-hydroxyethyl)phenyl 5-O-(3-methylbutyryl)-beta-d-apiofuranosyl(l-->2)-beta-d-glucopyranoside (2), 4-(2-hydroxyethyl)phenyl 5-O-nicotinyl-beta-d-apiofuranosyl(l-->2)-beta-d-glucopyranoside (3), 4-(2-hydroxyethyl)phenyl 5-O-(4-aminobenzoyl)-beta-d-apiofuranosyl(l-->2)-beta-d-glucopyranoside (4), 4-(2-hydroxyethyl)-2-methoxyphenyl 5-O-(2-S-2-methylbutyryl)-beta-d-apiofuranosyl(l-->2)-beta-d-glucopyranoside (5), 4-(hydroxymethyl)phenyl 5-O-(2-S-2-methylbutyryl)-beta-d-apiofuranosyl(l-->2)-beta-d-glucopyranoside (6), 4-(hydroxymethyl)phenyl 5-O-nicotinyl-beta-d-apiofuranosyl(l-->2)-beta-d-glucopyranoside (7), and 4-(hydroxymethyl)phenyl 5-O-(4-aminobenzoyl)-beta-d-apiofuranosyl(l-->2)-beta-d-glucopyranoside (8) on the basis of various spectroscopic analyses and analyses of hydrolysis products.

Acetylation↗
PubMed2003

Purification and characterization of Moschatin, a novel type I ribosome-inactivating protein from the mature seeds of pumpkin (Cucurbita moschata), and preparation of its immunotoxin against human melanoma cells.

A novel ribosome-inactivating protein designated Moschatin from the mature seeds of pumpkin (Cucurbita moschata) has been successively purified to homogeneity, using ammonium sulfate precipitation, CM-cellulose 52 column chromatography, Blue Sepharose CL-6B Affinity column chromatography and FPLC size-exclusion column chromatography. Moschatin is a type 1 RIP with a pI of 9.4 and molecular weight of approximately 29 kD. It is a rRNA N-glycosidase and potently blocked the protein synthesis in the rabbit reticulocyte lysate with a IC50 of 0.26 nM. Using the anti-human melanoma McAb Ng76, a novel immunotoxin Moschatin-Ng76 was prepared successfully and it efficiently inhibited the growth of targeted melanoma cells M21 with a IC50 of 0.04 nM, 1500 times lower than that of free Moschatin. The results implied that Moschatin could be used as a new potential anticancer agent.

Animals↗
PubMed1965

THE REDUCTION OF NITRATE, NITRITE AND HYDROXYLAMINE TO AMMONIA BY ENZYMES FROM CUCURBITA PEPO L. IN THE PRESENCE OF REDUCED BENZYL VIOLOGEN AS ELECTRON DONOR.

1. Enzyme systems from Cucurbita pepo have been shown to catalyse the reduction of nitrite and hydroxylamine to ammonia in yields about 90-100%. 2. Reduced benzyl viologen serves as an efficient electron donor for both systems. Activity of the nitrite-reductase system is directly related to degree of dye reduction when expressed in terms of the function for oxidation-reduction potentials, but appears to decrease to negligible activity below about 9% dye reduction. 3. NADH and NADPH alone produce negligible nitrite loss, but NADPH can be linked to an endogenous diaphorase system to reduce nitrite to ammonia in the presence of catalytic amounts of benzyl viologen. 4. The NADH- or NADPH-nitrate-reductase system that is also present can accept electrons from reduced benzyl viologen, but shows relationships opposite to that for the nitrite-reductase system with regard to effect of degree of dye reduction on activity. The product of nitrate reduction may be nitrite alone, or nitrite and ammonia, or ammonia alone, according only to the degree of dye reduction. 5. The relative activities of nitrite-reductase and hydroxylamine-reductase systems show different relationships with degree of dye reduction and may become reversed in magnitude when effects of degree of dye reduction are tested over a suitable range. 6. Nitrite severely inhibits the rate of reduction of hydroxylamine without affecting the yield of ammonia as a percentage of total substrate loss, but hydroxylamine has a negligible effect on the activity of the nitrite-reductase system. 7. The apparent K(m) for nitrite (1 mum) is substantially less than that for hydroxylamine, for which variable values between 0.05 and 0.9mm (mean 0.51 mm) have been observed. 8. The apparent K(m) values for reduced benzyl viologen differ for the nitrite-reductase and hydroxylamine-reductase systems: 60 and 7.5 mum respectively. 9. It is concluded that free hydroxylamine may not be an intermediate in the reduction of nitrite to ammonia by plants, and a possible mechanism for reduction of both compounds by the same enzyme system is discussed in the light of current ideas relating to other organisms.

Ammonia↗
PubMed2001

Fecundity of transgenic wild-crop hybrids of Cucurbita pepo (Cucurbitaceae): implications for crop-to-wild gene flow.

Hybridization between crops and their weedy or wild relatives is an area of concern because the widespread use of genetically engineered crops may allow novel, beneficial transgenes to enter nearby populations. We compared fitness components of wild Cucurbita pepo from Arkansas, USA, with wild-crop hybrids derived from yellow squash (a cultivar of C. pepo with transgenic resistance to two viruses). Wild and hybrid progeny were grown in agricultural fields in Arkansas (1996-98) and Ohio (1996) in six similar experiments. Cross types (wild and hybrid) did not differ significantly in seedling survival, which exceeded 85% in all cases. In Ohio, where more detailed observations were made, hybrid plants produced 41% as many male flowers, 21% as many female flowers, and 28% as many seeds as wild plants. At all sites, flowering periods of the two cross types overlapped extensively. Putative virus symptoms were more common in wild plants than in hybrids. Lifetime fecundity varied considerably among sites and years. The average fecundity of hybrids ranged from 453 to 4497 seeds per plant and represented 15% - 53% of the numbers of seeds produced by wild plants in the same experiments. These results suggest that the F1 generation does not represent a strong barrier to the introgression of neutral or beneficial crop genes into free-living populations of C. pepo.

Crops, Agricultural↗
PubMed2005

Changes in the subcellular distribution of glutathione during virus infection in Cucurbita pepo (L.).

Changes in the subcellular distribution and quantification of glutathione were studied with electron microscopic immunogold cytochemistry in Zucchini yellow mosaic virus (ZYMV)-infected Styrian pumpkin plants (Cucurbita pepo L. ssp. pepo var. styriaca Greb.) two weeks after inoculation. The amount of gold particles bound to glutathione was statistically evaluated for different cell structures, including mitochondria, plastids, nuclei, peroxisomes, and cytosol. In general, ZYMV-infected plants showed higher gold labelling density in intact mesophyll cells of the 5th (older leaves) and the youngest fully developed leaves (younger leaves), and decreased levels of glutathione within root tip cells when compared to the control. In general, within older and younger leaves the highest amount of gold particles was found in mitochondria and the lowest amount in plastids. In ZYMV-infected older leaves, an increase in glutathione was found in peroxisomes (1.7-fold), the cytosol (1.6-fold), mitochondria (1.4-fold), and nuclei (1.2-fold), whereas glutathione levels in plastids did not differ significantly when compared to control cells. In ZYMV-infected younger leaves elevated glutathione contents were found in the cytosol (3-fold), nuclei (2.1-fold), peroxisomes (1.8-fold), and plastids (1.5-fold), whereas mitochondria showed an insignificant decrease in glutathione levels in comparison to the control. In root tip cells of ZYMV-infected plants the amount of gold particles bound to glutathione was decreased in all investigated cell structures by between 0.7- to 0.8-fold. Additionally, total glutathione contents were determined in older and younger leaves using high-performance liquid chromatography (HPLC), which revealed no significant differences between control and ZYMV-infected leaves. The relevance of the results of both methods were compared and are discussed.

Cell Nucleus↗
PubMed2006

Nectar resorption and translocation in Cucurbita pepo L. and Platanthera chlorantha Custer (Rchb.).

Nectar resorption and sugar translocation were studied in Cucurbita pepo (Cucurbitaceae) and Platanthera chlorantha (Orchidaceae) by micro-autoradiography. In both species, nectar was resorbed in pollinated and unpollinated flowers and ovules developing into seeds were found to be the main sugar sink. In C. pepo, the mobility of resorbed sugars in pollinated female flowers was higher than in unpollinated ones; male flowers showed lower mobility of resorbed sugar. In P. chlorantha, radioactivity was detected in pollinated flowers below and above labelled unpollinated ones: the nearer the flower, the stronger the accumulation of label in developing fruits.

Autoradiography↗
PubMed2006

Virus-induced changes in the subcellular distribution of glutathione precursors in Cucurbita pepo (L.).

Changes in glutathione contents occur in plants during environmental stress situations, such as pathogen attack, as the formation of reactive oxygen species leads to the activation of the antioxidative defence system. As glutathione is synthesized out of its constituents cysteine, glycine, and glutamate the availability of these components will limit glutathione synthesis in plants especially during stress situations and therefore the ability of the plant to fight oxidative stress. To gain a deeper insight into possible limitations of glutathione synthesis during pathogen attack the present investigations were aimed to study how the subcellular distribution of glutathione precursors correlates with the subcellular distribution of glutathione during virus attack in plants. Selective antibodies against cysteine, glutamate, and glycine were used to study the impact of Zucchini yellow mosaic virus (ZYMV) infection on glutathione precursor contents within different cell compartments of cells from Cucurbita pepo (L.) plants with the transmission electron microscope (TEM). Generally, levels of cysteine and glutamate were found to be strongly decreased in most cell compartments of younger and older leaves including glutathione-producing cell compartments such as plastids and the cytosol. The strongest decrease of cysteine was found in plastids (- 54 %) and mitochondria (- 51 %) of younger leaves and in vacuoles (- 37 %) and plastids (- 29 %) of older leaves. The strongest decrease of glutamate in younger leaves occurred in peroxisomes (- 67 %) and nuclei (- 58 %) and in peroxisomes (- 64 %) and plastids (- 52 %) of the older ones. Glycine levels were found to be strongly decreased (- 63 % in mitochondria and - 53 % in plastids) in most cell compartments of older leaves and strongly increased (about 50 % in plastids and peroxisomes) in all cell compartments of the younger ones. These results indicate that low glycine contents in the older leaves were responsible for low levels of glutathione in these organs during ZYMV infection rather than limited amounts of cysteine or glutamate. Glutathione precursors were virtually absent in cell walls and intercellular spaces and play therefore no important role during ZYMV attack in the apoplast. While glutamate was absent in vacuoles, elevated levels of glycine (up to 30 %) and decreased cysteine contents (up to - 37 %) were observed in vacuoles during ZYMV infection. The impact of the present results on the current knowledge about glutathione synthesis and degradation on the cellular level during ZYMV infection are discussed.

Cucurbita↗