Nitrogen metabolism including nitrate reductase (EC 1.6.6.1), glutamate dehydroge‐nase (EC 1.4.1.... more Nitrogen metabolism including nitrate reductase (EC 1.6.6.1), glutamate dehydroge‐nase (EC 1.4.1.2) and glutamate‐oxalacetate aminotransferase (EC 2.6.1.1) activities were studied during growth of petals taken from carnation flowers (Dianthus caryophyllus L. cv. Sir Arthur) together with senescence parameters (lipid hydroper‐oxides, soluble amino acids and permeability). A slight decline in nitrogen percentage on a dry weight basis was found together with a sharp decrease in nitrate reduct‐ase, glutamate‐oxalacetate aminotransferase and glutamate dehydrogenase activities during the maximum growth phase, which was characterized by increase in respiration, dry weight, length, organic nitrogen and DNA per petal. Changes generally associated with senescence, like lipid hydroperoxide and soluble ammo nitrogen accumulation and increases in permeability began to appear already during early growth. The results indicate that permeability and proteolysis may be closely related. The possible significance of the decrease in nitrogen percentage and enzyme activities during growth of petals is discussed.
Sucrose synthase (SS), the key sucrose hydrolytic enzyme (EC 2.4.1.13), plays an important role i... more Sucrose synthase (SS), the key sucrose hydrolytic enzyme (EC 2.4.1.13), plays an important role in N 2-®xing nodule metabolism. It has also been proposed that N 2 ®xation in soybean nodules could be mediated by the potential to metabolize sucrose. The isolation and characterization of a nodule-enhanced SS full-length cDNA clone from the bean Phaseolus vulgaris is reported here. Southern blot analysis indicated that there are at least two SS genes in beans. Using a 3¢ speci®c probe from this SS cDNA clone, it was possible to identify a nodule-enhanced SS gene (PvSSn), which is expressed almost exclusively in nodules. A second gene (PvSS), which is expressed in all tissues tested, was detected using a coding region probe. Nodule-enhanced PvSSn transcript levels, but not the enzyme activity or protein amount, is reduced during nodule development. These data indicated that this reduction could be due to a limitation on the carbon availability in the nodule. PvSSn expression is reduced in the asparagine-treated nodules. By contrast, PvSSn transcript levels in nodules increased in the presence of glutamine, allantoin and allopurinol. This result suggests a relationship between ureide transport and SS regulation and could help in understanding why the ureide transport mechanism is activated during nitrogen ®xation in bean.
The effect of water stress on nitrate reductase (NR) activity was studied in detached oat leaves ... more The effect of water stress on nitrate reductase (NR) activity was studied in detached oat leaves exposed to various levels of oxygen. Drought inhibited NADH: NR only in the presence of oxygen, the higher the concentration of oxygen in the atmosphere, the stronger the effect of water stress. The decrease in NADH: NR was always accompanied by an increase in FMNH: NR and NADH: cytochrome c reductase (CR) partial activities. Infiltration of segments with free-radical scavengers prior to drought and oxygen treatment completely impeded the decrease in NADH: NR and the increase in FMNH: NR and CR activities. A trypsin-like inhibitor only partially reversed the loss of NADH: NR. Phenylmethylsulfonyl fluoride (PMSF), a protease inhibitor, and EDTA-Na had no effect on NADH: NR. The evidence presented here strongly suggests that drought and oxygen stress inhibit NR mainly via oxygen free radicals, which may stimulate the hydrolysis of the protein.
NADH-dependent glutamate synthase (NADH-GOGAT) is a key enzyme in primary ammonia assimilation in... more NADH-dependent glutamate synthase (NADH-GOGAT) is a key enzyme in primary ammonia assimilation in Phaseolus vulgaris nodules. Two different types of cDNA clones of PvNADH-GOGAT were isolated from the nodule cDNA libraries. The full-length cDNA clones of PvNADH-GOGAT-I (7.4 kb) and PvNADH-GOGAT-II (7.0 kb), which displayed an 83% homology between them, were isolated using cDNA library screening, 'cDNA library walking' and RT-PCR amplification. Southern analysis employing specific 5Ј cDNA probes derived from PvNADH-GOGAT-I and PvNADH-GOGAT-II indicated the existence of a single copy of each gene in the bean genome. Both these proteins contain~100 amino acid sequences theoretically addressing each isoenzyme to different subcellular compartments. RT-PCR analysis indicated that PvNADH-GOGAT-II expression is higher than PvNADH-GOGAT-I during nodule development. Expression analysis by RT-PCR also revealed that both of these genes are differentially regulated by sucrose. On the other hand, the expression of PvNADH-GOGAT-I, but not PvNADH-GOGAT-II, was inhibited with nitrogen compounds. In situ hybridization and promoter expression analyses demonstrated that the NADH-GOGAT-I and-II genes are differentially expressed in bean root and nodule tissues. In silico analyses of the NADH-GOGAT promoters revealed the presence of potential cis elements in them that could mediate differential tissue-specific, and sugar and amino acid responsive expression of these genes.
Soybean (Glycine max L.) is an important source of protein for human and animal nutrition, as wel... more Soybean (Glycine max L.) is an important source of protein for human and animal nutrition, as well as a major source of vegetable oil. The soybean crop requires adequate water all through its growth period to attain its yield potential, and the lack of soil moisture at critical stages of growth profoundly impacts the productivity. In this study, utilizing 1H NMR-based metabolite analysis combined with the physiological studies we assessed the effects of short-term water stress on overall growth, nitrogen fixation, ureide and proline dynamics, as well as metabolic changes in drought tolerant (NA5009RG) and sensitive (DM50048) genotypes of soybean in order to elucidate metabolite adjustments in relation to the physiological responses in the nitrogen-fixing plants towards water limitation. The results of our analysis demonstrated critical differences in physiological responses between these two genotypes, and identified the metabolic pathways that are affected by short-term water limit...
ABSTRACT Leaf segments of 8-day-old oat plants were incubated with 100μM glycolate or xanthine in... more ABSTRACT Leaf segments of 8-day-old oat plants were incubated with 100μM glycolate or xanthine inthe dark or with 100μM paraquat in the light. These treatments were designed to enhance the internalproduction of activated oxygen species and to allow us to study the possible role of such species in the regulation of the activity of NO 3 -induced nitrate reductase (NR) in illuminated green tissues. Each of the treatments inhibited NO 3 -induced NR activity. HPMS (an inhibitor of glycolate oxidase) and allopurinol (an inhibitor of xanthine oxidase) protected NR from inactivation by glycolate and by xanthine. Free radical scavengers also protected NR from inactivation by xanthine and paraquat but they had no effect on the inhibition by glycolate. The activities of catalase and super oxide dismutase did not increase in response to the various treatments. Thus, while the production of active oxygen species appears to be stimulated under such conditions, the rates of the enzymatic destruction and “scavenging” of these species seem to remain unchanged. Hence, the endogenous levels of these active oxygen species can be expected to increase in the treated leaves. We interpret our results as evidence that inhibition by glycolate of NR is caused by the H 2 O 2 generated during the oxidation of glycolate while the effects of xanthine and paraquat depend on the enhanced production of free radicals. It is concluded that activated oxygen species that are formed endogenously may play a role in the regulation of NR in green tissues.
... Sucrose-Modulated Morphogenesis in Anagallis arvensis L. Sonia T. Silvente and Victorio S. Tr... more ... Sucrose-Modulated Morphogenesis in Anagallis arvensis L. Sonia T. Silvente and Victorio S. Trippi ... pp. 1-116. Reveuni, O. and I. Adata (1974) Endogenous carbohydrates, root promoters and root inhibitors in easy and difficult to root date palm {Phoenix dactylifera L.) of shoots. ...
Nitrogen metabolism including nitrate reductase (EC 1.6.6.1), glutamate dehydroge‐nase (EC 1.4.1.... more Nitrogen metabolism including nitrate reductase (EC 1.6.6.1), glutamate dehydroge‐nase (EC 1.4.1.2) and glutamate‐oxalacetate aminotransferase (EC 2.6.1.1) activities were studied during growth of petals taken from carnation flowers (Dianthus caryophyllus L. cv. Sir Arthur) together with senescence parameters (lipid hydroper‐oxides, soluble amino acids and permeability). A slight decline in nitrogen percentage on a dry weight basis was found together with a sharp decrease in nitrate reduct‐ase, glutamate‐oxalacetate aminotransferase and glutamate dehydrogenase activities during the maximum growth phase, which was characterized by increase in respiration, dry weight, length, organic nitrogen and DNA per petal. Changes generally associated with senescence, like lipid hydroperoxide and soluble ammo nitrogen accumulation and increases in permeability began to appear already during early growth. The results indicate that permeability and proteolysis may be closely related. The possible significance of the decrease in nitrogen percentage and enzyme activities during growth of petals is discussed.
Sucrose synthase (SS), the key sucrose hydrolytic enzyme (EC 2.4.1.13), plays an important role i... more Sucrose synthase (SS), the key sucrose hydrolytic enzyme (EC 2.4.1.13), plays an important role in N 2-®xing nodule metabolism. It has also been proposed that N 2 ®xation in soybean nodules could be mediated by the potential to metabolize sucrose. The isolation and characterization of a nodule-enhanced SS full-length cDNA clone from the bean Phaseolus vulgaris is reported here. Southern blot analysis indicated that there are at least two SS genes in beans. Using a 3¢ speci®c probe from this SS cDNA clone, it was possible to identify a nodule-enhanced SS gene (PvSSn), which is expressed almost exclusively in nodules. A second gene (PvSS), which is expressed in all tissues tested, was detected using a coding region probe. Nodule-enhanced PvSSn transcript levels, but not the enzyme activity or protein amount, is reduced during nodule development. These data indicated that this reduction could be due to a limitation on the carbon availability in the nodule. PvSSn expression is reduced in the asparagine-treated nodules. By contrast, PvSSn transcript levels in nodules increased in the presence of glutamine, allantoin and allopurinol. This result suggests a relationship between ureide transport and SS regulation and could help in understanding why the ureide transport mechanism is activated during nitrogen ®xation in bean.
The effect of water stress on nitrate reductase (NR) activity was studied in detached oat leaves ... more The effect of water stress on nitrate reductase (NR) activity was studied in detached oat leaves exposed to various levels of oxygen. Drought inhibited NADH: NR only in the presence of oxygen, the higher the concentration of oxygen in the atmosphere, the stronger the effect of water stress. The decrease in NADH: NR was always accompanied by an increase in FMNH: NR and NADH: cytochrome c reductase (CR) partial activities. Infiltration of segments with free-radical scavengers prior to drought and oxygen treatment completely impeded the decrease in NADH: NR and the increase in FMNH: NR and CR activities. A trypsin-like inhibitor only partially reversed the loss of NADH: NR. Phenylmethylsulfonyl fluoride (PMSF), a protease inhibitor, and EDTA-Na had no effect on NADH: NR. The evidence presented here strongly suggests that drought and oxygen stress inhibit NR mainly via oxygen free radicals, which may stimulate the hydrolysis of the protein.
NADH-dependent glutamate synthase (NADH-GOGAT) is a key enzyme in primary ammonia assimilation in... more NADH-dependent glutamate synthase (NADH-GOGAT) is a key enzyme in primary ammonia assimilation in Phaseolus vulgaris nodules. Two different types of cDNA clones of PvNADH-GOGAT were isolated from the nodule cDNA libraries. The full-length cDNA clones of PvNADH-GOGAT-I (7.4 kb) and PvNADH-GOGAT-II (7.0 kb), which displayed an 83% homology between them, were isolated using cDNA library screening, 'cDNA library walking' and RT-PCR amplification. Southern analysis employing specific 5Ј cDNA probes derived from PvNADH-GOGAT-I and PvNADH-GOGAT-II indicated the existence of a single copy of each gene in the bean genome. Both these proteins contain~100 amino acid sequences theoretically addressing each isoenzyme to different subcellular compartments. RT-PCR analysis indicated that PvNADH-GOGAT-II expression is higher than PvNADH-GOGAT-I during nodule development. Expression analysis by RT-PCR also revealed that both of these genes are differentially regulated by sucrose. On the other hand, the expression of PvNADH-GOGAT-I, but not PvNADH-GOGAT-II, was inhibited with nitrogen compounds. In situ hybridization and promoter expression analyses demonstrated that the NADH-GOGAT-I and-II genes are differentially expressed in bean root and nodule tissues. In silico analyses of the NADH-GOGAT promoters revealed the presence of potential cis elements in them that could mediate differential tissue-specific, and sugar and amino acid responsive expression of these genes.
Soybean (Glycine max L.) is an important source of protein for human and animal nutrition, as wel... more Soybean (Glycine max L.) is an important source of protein for human and animal nutrition, as well as a major source of vegetable oil. The soybean crop requires adequate water all through its growth period to attain its yield potential, and the lack of soil moisture at critical stages of growth profoundly impacts the productivity. In this study, utilizing 1H NMR-based metabolite analysis combined with the physiological studies we assessed the effects of short-term water stress on overall growth, nitrogen fixation, ureide and proline dynamics, as well as metabolic changes in drought tolerant (NA5009RG) and sensitive (DM50048) genotypes of soybean in order to elucidate metabolite adjustments in relation to the physiological responses in the nitrogen-fixing plants towards water limitation. The results of our analysis demonstrated critical differences in physiological responses between these two genotypes, and identified the metabolic pathways that are affected by short-term water limit...
ABSTRACT Leaf segments of 8-day-old oat plants were incubated with 100μM glycolate or xanthine in... more ABSTRACT Leaf segments of 8-day-old oat plants were incubated with 100μM glycolate or xanthine inthe dark or with 100μM paraquat in the light. These treatments were designed to enhance the internalproduction of activated oxygen species and to allow us to study the possible role of such species in the regulation of the activity of NO 3 -induced nitrate reductase (NR) in illuminated green tissues. Each of the treatments inhibited NO 3 -induced NR activity. HPMS (an inhibitor of glycolate oxidase) and allopurinol (an inhibitor of xanthine oxidase) protected NR from inactivation by glycolate and by xanthine. Free radical scavengers also protected NR from inactivation by xanthine and paraquat but they had no effect on the inhibition by glycolate. The activities of catalase and super oxide dismutase did not increase in response to the various treatments. Thus, while the production of active oxygen species appears to be stimulated under such conditions, the rates of the enzymatic destruction and “scavenging” of these species seem to remain unchanged. Hence, the endogenous levels of these active oxygen species can be expected to increase in the treated leaves. We interpret our results as evidence that inhibition by glycolate of NR is caused by the H 2 O 2 generated during the oxidation of glycolate while the effects of xanthine and paraquat depend on the enhanced production of free radicals. It is concluded that activated oxygen species that are formed endogenously may play a role in the regulation of NR in green tissues.
... Sucrose-Modulated Morphogenesis in Anagallis arvensis L. Sonia T. Silvente and Victorio S. Tr... more ... Sucrose-Modulated Morphogenesis in Anagallis arvensis L. Sonia T. Silvente and Victorio S. Trippi ... pp. 1-116. Reveuni, O. and I. Adata (1974) Endogenous carbohydrates, root promoters and root inhibitors in easy and difficult to root date palm {Phoenix dactylifera L.) of shoots. ...
Uploads
Papers by Sonia Silvente