In light of ongoing climate changes in wine-growing regions, the selection of drought-tolerant ro... more In light of ongoing climate changes in wine-growing regions, the selection of drought-tolerant rootstocks is becoming a crucial factor for developing a sustainable viticulture. In this study, M4, a new rootstock genotype that shows tolerance to drought, was compared from a genomic and transcriptomic point of view with the less drought-tolerant genotype 101.14. The root and leaf transcriptome of both 101.14 and the M4 rootstock genotype was analysed, following exposure to progressive drought conditions. Multifactorial analyses indicated that stress treatment represents the main factor driving differential gene expression in roots, whereas in leaves the genotype is the prominent factor. Upon stress, M4 roots and leaves showed a higher induction of resveratrol and flavonoid biosynthetic genes, respectively. The higher expression of VvSTS genes in M4, confirmed by the accumulation of higher levels of resveratrol in M4 roots compared with 101.14, was coupled to an up-regulation of severa...
A proteomic analysis was conducted on peach fruit mesocarp in order to better elucidate the bioch... more A proteomic analysis was conducted on peach fruit mesocarp in order to better elucidate the biochemical and physiological events which characterize the transition of fruit from the "unripe" to the "ripe" phase. The first goal of the present work was to set-up a protocol suitable for improving protein extraction from peach mesocarp. The use of freeze-dried powdered tissue, together with the addition of phenol prior to the extraction with an aqueous buffer, significantly increased the protein yield and the quality of 2-DE gels. The proteomic profiles of the mesocarp from peach fruit of a non-melting flesh (NMF; 'Oro A') and a melting flesh (MF; 'Bolero') cultivar, at "unripe" and "ripe" stages as defined by some parameters typical of ripening, were then analyzed. The comparative analysis of the 2-DE gels showed that in NMF and MF peaches the relative volumes of 53 protein spots significantly changed in relation to both the ripeni...
In this study a new selected genotype, named M4, was compared with the commercial rootstock 101.1... more In this study a new selected genotype, named M4, was compared with the commercial rootstock 101.14, analyzing the responses evoked by water stress (WS) in both the shoot and the root organs.The effects of progressive WS were studied in controlled environmental conditions on plants grown in pots.Sixty-six plants for each genotype were randomized to obtain two pools that were used as i) control, plants that were maintained at the 80% of the soil field capacity; ii) water stress, plants in which water supply was progressively reduced until the 30% of the field capacity. The effects of WS was studied for an experiment period of 10 days.Shoot growth, plant water status and leaf photosynthetic parameters were measured. Moreover, the levels of sugars, amino acids and total proteins as well as the contents of the more abundant ions were determined. M4 genotype resulted to have a greater capacity to tolerate WS, maintaining photosynthetic activity also under severe stress conditions. In this genotype, the root system appeared to play a central role to sustain biochemical and physiological responses evoked by WS, as suggested by the greater capacity to adequate inorganic and organic osmolytes in this organ. The experiments conducted in grafted plants, in which Cabernet was the used as scion, confirmed the good performances of M4.This study produced a first characterization of the new putative rootstock M4, in which the greater tolerance to WS, resulted strictly related to root integrity/functionality, so confirming that these aspects have to be considered in the further selection programs.
In the light of possible climate change, a crucial aspect for future Mediterranean Europe viticul... more In the light of possible climate change, a crucial aspect for future Mediterranean Europe viticulture is the selection of new rootstocks exhibiting adaptability to drought and salt stress. In this context and considering recent literature, it is fundamental to increase our knowledge of the biochemical and physiological events that characterise stress responses in grapevine roots. In this study, leaf and root responses induced by water stress (WS) and sodium chloride (NaCl) exposure in a new selected genotype, named M4 (Vitis vinifera × V. berlandieri) × V. berlandieri cv. Resseguier n. 1), were compared with those of the commercial rootstock 101.14. Methods and Results: The effect of progressive WS and NaCl exposure was studied under controlled environmental conditions on ungrafted plants grown in pots. Shoot growth, plant water status and leaf photosynthetic parameters were measured. The concentration of sugars, amino acids and total proteins, as well as the concentration of the more abundant ions, was determined in both leaf and root organs. The M4 genotype showed greater capacity to maintain photosynthetic activity, to accumulate osmotic compounds as well as to counteract Na and Cl accumulation. Conclusion: The M4 genotype exhibited a greater capacity to tolerate both WS and exposure to an increasing concentration of Na and Cl, maintaining photosynthetic activity also under severe stress conditions. The root system appeared to play a central role in sustaining biochemical and physiological responses experienced under these adverse conditions. Significance of the Study: This study showed that the tolerance to abiotic stress conditions, such as WS and NaCl exposure, depends on root integrity/functionality, confirming that these aspects must be considered in further selection programs.
The acidification of endomembrane compartments is essential for enzyme activities, sorting, traff... more The acidification of endomembrane compartments is essential for enzyme activities, sorting, trafficking, and trans-membrane transport of various compounds. Vacuoles are mildly acidic in most plant cells because of the action of V-ATPase and/or pyrophosphatase proton pumps but are hyperacidified in specific cells by mechanisms that remained unclear. Here, we show that the blue petal color of petunia ph mutants is due to a failure to hyperacidify vacuoles. We report that PH1 encodes a P 3B -ATPase, hitherto known as Mg2 + transporters in bacteria only, that resides in the vacuolar membrane (tonoplast). In vivo nuclear magnetic resonance and genetic data show that PH1 is required and, together with the tonoplast H + P 3A -ATPase PH5, sufficient to hyperacidify vacuoles. PH1 has no H + transport activity on its own but can physically interact with PH5 and boost PH5 H + transport activity. Hence, the hyperacidification of vacuoles in petals, and possibly other tissues, relies on a heteromeric P-ATPase pump.
Proteomic analysis of fruits of three peach (Prunus persica) cultivars with different flesh firmn... more Proteomic analysis of fruits of three peach (Prunus persica) cultivars with different flesh firmness characteristics during ripening / C. Fedeli, AS Negri, B. Prinsi, S. Morgutti, N. Negrini, M. Cocucci, L. Espen. ((Intervento presentato al 2. convegno Annual National Conference of ...
Peach fruit (Prunus persica [L.] Batsch) ripening : a proteomic comparative analysis of two culti... more Peach fruit (Prunus persica [L.] Batsch) ripening : a proteomic comparative analysis of two cultivars with different flesh firmness characteristics at the transition from the pre-climacteric to the climacteric stage / C. Fedeli, AS Negri, B. Prinsi, S. Morgutti, N. Negrini, M. Cocucci, L. Espen. (( ...
Background: Iron deficiency induces in Strategy I plants physiological, biochemical and molecular... more Background: Iron deficiency induces in Strategy I plants physiological, biochemical and molecular modifications capable to increase iron uptake from the rhizosphere. This effort needs a reorganization of metabolic pathways to efficiently sustain activities linked to the acquisition of iron; in fact, carbohydrates and the energetic metabolism has been shown to be involved in these responses. The aim of this work was to find both a confirmation of the already expected change in the enzyme concentrations induced in cucumber root tissue in response to iron deficiency as well as to find new insights on the involvement of other pathways. Results: The proteome pattern of soluble cytosolic proteins extracted from roots was obtained by 2-DE. Of about two thousand spots found, only those showing at least a two-fold increase or decrease in the concentration were considered for subsequent identification by mass spectrometry. Fifty-seven proteins showed significant changes, and 44 of them were identified. Twenty-one of them were increased in quantity, whereas 23 were decreased in quantity. Most of the increased proteins belong to glycolysis and nitrogen metabolism in agreement with the biochemical evidence. On the other hand, the proteins being decreased belong to the metabolism of sucrose and complex structural carbohydrates and to structural proteins. Conclusions: The new available techniques allow to cast new light on the mechanisms involved in the changes occurring in plants under iron deficiency. The data obtained from this proteomic study confirm the metabolic changes occurring in cucumber as a response to Fe deficiency. Two main conclusions may be drawn. The first one is the confirmation of the increase in the glycolytic flux and in the anaerobic metabolism to sustain the energetic effort the Fe-deficient plants must undertake. The second conclusion is, on one hand, the decrease in the amount of enzymes linked to the biosynthesis of complex carbohydrates of the cell wall, and, on the other hand, the increase in enzymes linked to the turnover of proteins.
Background: Nitrogen nutrition is one of the major factors that limit growth and production of cr... more Background: Nitrogen nutrition is one of the major factors that limit growth and production of crop plants. It affects many processes, such as development, architecture, flowering, senescence and photosynthesis. Although the improvement in technologies for protein study and the widening of gene sequences have made possible the study of the plant proteomes, only limited information on proteome changes occurring in response to nitrogen amount are available up to now. In this work, two-dimensional gel electrophoresis (2-DE) has been used to investigate the protein changes induced by NO 3concentration in both roots and leaves of maize (Zea mays L.) plants. Moreover, in order to better evaluate the proteomic results, some biochemical and physiological parameters were measured.
We propose an integrated approach, obtained by the combination of multivariate statistics and pro... more We propose an integrated approach, obtained by the combination of multivariate statistics and proteomics, useful to isolate candidate biomarkers for the evaluation of grape ripening. We carried out a comparative 2-DE analysis of grape skins collected in three moments of ripening and analyzed the spot volume dataset through the application of principal component analysis followed by forward stepwiselinear discriminant analysis. This technique allowed to discriminate véraison, quite mature and mature samples, and to sort the matched spots according to their significance. We identified 36 spots showing high discriminating coefficients through liquid chromatography -electrospray ionization -tandem mass spectrometry (LC-ESI-MS/MS). Most of them were involved in biotic and abiotic stress responses indicating these enzymes as good candidate markers of berry ripening. These evidences hint at a likely developmental role of these proteins, in addition to their reported activity in stress events. Restricting the same statistical analysis to the samples belonging to the two last stages, it was indicated that this approach can clearly distinguish these close and similar phases of berry development. Taken all together, these results bear out that the employment of the combination of 2-DE and multivariate statistics is a reliable tool in the identification of new protein markers for describing the ripening phases and to assess the overall quality of the fruit.
In light of ongoing climate changes in wine-growing regions, the selection of drought-tolerant ro... more In light of ongoing climate changes in wine-growing regions, the selection of drought-tolerant rootstocks is becoming a crucial factor for developing a sustainable viticulture. In this study, M4, a new rootstock genotype that shows tolerance to drought, was compared from a genomic and transcriptomic point of view with the less drought-tolerant genotype 101.14. The root and leaf transcriptome of both 101.14 and the M4 rootstock genotype was analysed, following exposure to progressive drought conditions. Multifactorial analyses indicated that stress treatment represents the main factor driving differential gene expression in roots, whereas in leaves the genotype is the prominent factor. Upon stress, M4 roots and leaves showed a higher induction of resveratrol and flavonoid biosynthetic genes, respectively. The higher expression of VvSTS genes in M4, confirmed by the accumulation of higher levels of resveratrol in M4 roots compared with 101.14, was coupled to an up-regulation of severa...
A proteomic analysis was conducted on peach fruit mesocarp in order to better elucidate the bioch... more A proteomic analysis was conducted on peach fruit mesocarp in order to better elucidate the biochemical and physiological events which characterize the transition of fruit from the "unripe" to the "ripe" phase. The first goal of the present work was to set-up a protocol suitable for improving protein extraction from peach mesocarp. The use of freeze-dried powdered tissue, together with the addition of phenol prior to the extraction with an aqueous buffer, significantly increased the protein yield and the quality of 2-DE gels. The proteomic profiles of the mesocarp from peach fruit of a non-melting flesh (NMF; 'Oro A') and a melting flesh (MF; 'Bolero') cultivar, at "unripe" and "ripe" stages as defined by some parameters typical of ripening, were then analyzed. The comparative analysis of the 2-DE gels showed that in NMF and MF peaches the relative volumes of 53 protein spots significantly changed in relation to both the ripeni...
In this study a new selected genotype, named M4, was compared with the commercial rootstock 101.1... more In this study a new selected genotype, named M4, was compared with the commercial rootstock 101.14, analyzing the responses evoked by water stress (WS) in both the shoot and the root organs.The effects of progressive WS were studied in controlled environmental conditions on plants grown in pots.Sixty-six plants for each genotype were randomized to obtain two pools that were used as i) control, plants that were maintained at the 80% of the soil field capacity; ii) water stress, plants in which water supply was progressively reduced until the 30% of the field capacity. The effects of WS was studied for an experiment period of 10 days.Shoot growth, plant water status and leaf photosynthetic parameters were measured. Moreover, the levels of sugars, amino acids and total proteins as well as the contents of the more abundant ions were determined. M4 genotype resulted to have a greater capacity to tolerate WS, maintaining photosynthetic activity also under severe stress conditions. In this genotype, the root system appeared to play a central role to sustain biochemical and physiological responses evoked by WS, as suggested by the greater capacity to adequate inorganic and organic osmolytes in this organ. The experiments conducted in grafted plants, in which Cabernet was the used as scion, confirmed the good performances of M4.This study produced a first characterization of the new putative rootstock M4, in which the greater tolerance to WS, resulted strictly related to root integrity/functionality, so confirming that these aspects have to be considered in the further selection programs.
In the light of possible climate change, a crucial aspect for future Mediterranean Europe viticul... more In the light of possible climate change, a crucial aspect for future Mediterranean Europe viticulture is the selection of new rootstocks exhibiting adaptability to drought and salt stress. In this context and considering recent literature, it is fundamental to increase our knowledge of the biochemical and physiological events that characterise stress responses in grapevine roots. In this study, leaf and root responses induced by water stress (WS) and sodium chloride (NaCl) exposure in a new selected genotype, named M4 (Vitis vinifera × V. berlandieri) × V. berlandieri cv. Resseguier n. 1), were compared with those of the commercial rootstock 101.14. Methods and Results: The effect of progressive WS and NaCl exposure was studied under controlled environmental conditions on ungrafted plants grown in pots. Shoot growth, plant water status and leaf photosynthetic parameters were measured. The concentration of sugars, amino acids and total proteins, as well as the concentration of the more abundant ions, was determined in both leaf and root organs. The M4 genotype showed greater capacity to maintain photosynthetic activity, to accumulate osmotic compounds as well as to counteract Na and Cl accumulation. Conclusion: The M4 genotype exhibited a greater capacity to tolerate both WS and exposure to an increasing concentration of Na and Cl, maintaining photosynthetic activity also under severe stress conditions. The root system appeared to play a central role in sustaining biochemical and physiological responses experienced under these adverse conditions. Significance of the Study: This study showed that the tolerance to abiotic stress conditions, such as WS and NaCl exposure, depends on root integrity/functionality, confirming that these aspects must be considered in further selection programs.
The acidification of endomembrane compartments is essential for enzyme activities, sorting, traff... more The acidification of endomembrane compartments is essential for enzyme activities, sorting, trafficking, and trans-membrane transport of various compounds. Vacuoles are mildly acidic in most plant cells because of the action of V-ATPase and/or pyrophosphatase proton pumps but are hyperacidified in specific cells by mechanisms that remained unclear. Here, we show that the blue petal color of petunia ph mutants is due to a failure to hyperacidify vacuoles. We report that PH1 encodes a P 3B -ATPase, hitherto known as Mg2 + transporters in bacteria only, that resides in the vacuolar membrane (tonoplast). In vivo nuclear magnetic resonance and genetic data show that PH1 is required and, together with the tonoplast H + P 3A -ATPase PH5, sufficient to hyperacidify vacuoles. PH1 has no H + transport activity on its own but can physically interact with PH5 and boost PH5 H + transport activity. Hence, the hyperacidification of vacuoles in petals, and possibly other tissues, relies on a heteromeric P-ATPase pump.
Proteomic analysis of fruits of three peach (Prunus persica) cultivars with different flesh firmn... more Proteomic analysis of fruits of three peach (Prunus persica) cultivars with different flesh firmness characteristics during ripening / C. Fedeli, AS Negri, B. Prinsi, S. Morgutti, N. Negrini, M. Cocucci, L. Espen. ((Intervento presentato al 2. convegno Annual National Conference of ...
Peach fruit (Prunus persica [L.] Batsch) ripening : a proteomic comparative analysis of two culti... more Peach fruit (Prunus persica [L.] Batsch) ripening : a proteomic comparative analysis of two cultivars with different flesh firmness characteristics at the transition from the pre-climacteric to the climacteric stage / C. Fedeli, AS Negri, B. Prinsi, S. Morgutti, N. Negrini, M. Cocucci, L. Espen. (( ...
Background: Iron deficiency induces in Strategy I plants physiological, biochemical and molecular... more Background: Iron deficiency induces in Strategy I plants physiological, biochemical and molecular modifications capable to increase iron uptake from the rhizosphere. This effort needs a reorganization of metabolic pathways to efficiently sustain activities linked to the acquisition of iron; in fact, carbohydrates and the energetic metabolism has been shown to be involved in these responses. The aim of this work was to find both a confirmation of the already expected change in the enzyme concentrations induced in cucumber root tissue in response to iron deficiency as well as to find new insights on the involvement of other pathways. Results: The proteome pattern of soluble cytosolic proteins extracted from roots was obtained by 2-DE. Of about two thousand spots found, only those showing at least a two-fold increase or decrease in the concentration were considered for subsequent identification by mass spectrometry. Fifty-seven proteins showed significant changes, and 44 of them were identified. Twenty-one of them were increased in quantity, whereas 23 were decreased in quantity. Most of the increased proteins belong to glycolysis and nitrogen metabolism in agreement with the biochemical evidence. On the other hand, the proteins being decreased belong to the metabolism of sucrose and complex structural carbohydrates and to structural proteins. Conclusions: The new available techniques allow to cast new light on the mechanisms involved in the changes occurring in plants under iron deficiency. The data obtained from this proteomic study confirm the metabolic changes occurring in cucumber as a response to Fe deficiency. Two main conclusions may be drawn. The first one is the confirmation of the increase in the glycolytic flux and in the anaerobic metabolism to sustain the energetic effort the Fe-deficient plants must undertake. The second conclusion is, on one hand, the decrease in the amount of enzymes linked to the biosynthesis of complex carbohydrates of the cell wall, and, on the other hand, the increase in enzymes linked to the turnover of proteins.
Background: Nitrogen nutrition is one of the major factors that limit growth and production of cr... more Background: Nitrogen nutrition is one of the major factors that limit growth and production of crop plants. It affects many processes, such as development, architecture, flowering, senescence and photosynthesis. Although the improvement in technologies for protein study and the widening of gene sequences have made possible the study of the plant proteomes, only limited information on proteome changes occurring in response to nitrogen amount are available up to now. In this work, two-dimensional gel electrophoresis (2-DE) has been used to investigate the protein changes induced by NO 3concentration in both roots and leaves of maize (Zea mays L.) plants. Moreover, in order to better evaluate the proteomic results, some biochemical and physiological parameters were measured.
We propose an integrated approach, obtained by the combination of multivariate statistics and pro... more We propose an integrated approach, obtained by the combination of multivariate statistics and proteomics, useful to isolate candidate biomarkers for the evaluation of grape ripening. We carried out a comparative 2-DE analysis of grape skins collected in three moments of ripening and analyzed the spot volume dataset through the application of principal component analysis followed by forward stepwiselinear discriminant analysis. This technique allowed to discriminate véraison, quite mature and mature samples, and to sort the matched spots according to their significance. We identified 36 spots showing high discriminating coefficients through liquid chromatography -electrospray ionization -tandem mass spectrometry (LC-ESI-MS/MS). Most of them were involved in biotic and abiotic stress responses indicating these enzymes as good candidate markers of berry ripening. These evidences hint at a likely developmental role of these proteins, in addition to their reported activity in stress events. Restricting the same statistical analysis to the samples belonging to the two last stages, it was indicated that this approach can clearly distinguish these close and similar phases of berry development. Taken all together, these results bear out that the employment of the combination of 2-DE and multivariate statistics is a reliable tool in the identification of new protein markers for describing the ripening phases and to assess the overall quality of the fruit.
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Papers by B. Prinsi