De novo arginine biosynthesis in leaves of phosphorus-deficient citrus and poncirus species

Young, fully expanded leaves from 7-month-old P-deficient citrus rootstock seedlings had levels of nonprotein arginine that were 10- to 50-fold greater than those from P-sufficient control plants. Arginine content of the protein fraction increased 2- to 4-fold in P-deficient leaves. Total arginine content, which averaged 72 ± 6 micromoles per gram dry weight of P-sufficient leaf tissue (mean ± se, n = the four rootstocks) was 207, 308, 241, and 178 micromoles in P-deficient leaves from Citrus limon cv rough lemon, Poncirus trifoliata × C. sinensis cv Carrizo citrange and cv Troyer citrange, and P. trifoliata cv Australian trifoliate orange, respectively. For each rootstock, the accumulation of arginine paralleled an increase in the activity of the pathway for the de novo biosynthesis of arginine. The ratio of the nanomoles NaH¹⁴CO₃ incorporated into the combined pool of arginine plus urea per gram fresh weight intact leaf tissue during a 3-hour labeling period for P-deficient to P-sufficient plants was 91:34, 49:11, 35:11, and 52:41, respectively. When P-deficient plants were supplied with P, incorporation of NaH¹⁴CO₃ into arginine plus urea was reduced to the level observed for the P-sufficient control plants of the same age and arginine ceased to accumulate. Arginase and arginine decarboxylase activity were either unaffected or slightly increased during phosphorus deficiency. Taken together, these results provide strong evidence that arginine accumulation during phosphorus deficiency is due to increased activity of the de novo arginine biosynthetic pathway.     

Metabolic Pathway of alpha-Ketoglutarate in Citrus Leaves as Affected by Phosphorus Nutrition

The uptake and metabolism of α-[5-¹⁴C]ketoglutarate by phosphorus-deficient and full nutrient (control) lemon (Citrus limon) leaves were studied over various time intervals. After 45 minutes in P-deficient leaves, the bulk of incorporated ¹⁴C appeared in organic acids and much less in amino acids, while in the control leaves, the ¹⁴C contents of organic and amino acids were equal. In P-deficient leaves, after longer incubation times the ¹⁴C content of organic acids and amino acids increased, while that of CO₂ and residue fractions remained low. In full nutrient leaves the ¹⁴C content of amino acids and organic acids decreased after longer incubation time and increased in the insoluble residue and CO₂. In full nutrient leaves the organic and amino acid metabolism were closely related and accompanied by protein synthesis and CO₂ release, while in P-deficient leaves an accelerating accumulation of arginine and citric acid was linked together with inhibition of protein synthesis and CO₂ liberation.     

Diurnal starch accumulation and utilization in phosphorus-deficient soybean plants

The effects of phosphorus deficiency on carbohydrate accumulation and utilization in 34-day-old soybean (Glycine max L. Merr.) plants were characterized over a diurnal cycle to evaluate the mechanisms by which phosphorus deficiency restricts plant growth. Phosphorus deficiency decreased the net CO₂ exchange rate throughout the light period. The decrease in the CO₂ exhange rate was associated with a decrease in stomatal conductance and an increase in the internal CO₂ concentration. These observations indicate that phosphorus deficiency increased mesophyll resistance. Assimilate export rate from the youngest fully expanded leaves was decreased by phosphorus deficiency, whereas starch concentrations in these leaves were increased. Higher starch concentrations in phosphorus-deficient youngest fully expanded leaves resulted from a longer period of net starch accumulation and a shorter period of net starch degradation relative to those for phosphorus-sufficient controls. Phosphorus deficiency decreased sucrose-P synthase activity by 27% (averaged over the diurnal cycle), and essentially eliminated diurnal variation in sucrose-P-synthase activity. Diurnal variations in nonstructural carbohydrate concentrations in leaves and stems were also less pronounced in phosphorus-deficient plants than in controls. In phosphorus-deficient plants, only 30% of the whole plant starch present at the end of a light phase was utilized during the subsequent 12-hour dark phase as compared with 68% for phosphorus-sufficient controls. Although phosphorus deficiency decreased the CO₂ exchange rate and whole plant leaf area, accumulation of high starch concentrations in leaves and stems and restricted starch utilization in the dark indicate that growth processes (i.e. sink activities) were restricted to a greater extent than photosynthetic capacity. Further experimentation is required to determine whether decreased starch utilization in phosphorus-deficient plants is the cause or the result of restricted growth.     

Aspartate Carbamyltransferase : Site of End-Product Inhibition of the Orotate Pathway in Intact Cells of Cucurbita pepo

Lovatt et al. (1979 Plant Physiol 64: 562-569) have previously demonstrated that end-product inhibition functions as a mechanism regulating the activity of the orotic acid pathway in intact cells of roots excised from 2-day-old squash plants (Cucurbita pepo L. cv Early Prolific Straightneck). Uridine (0.5 millimolar final concentration) or one of its metabolites inhibited the incorporation of NaH¹⁴CO₃, but not [¹⁴C]carbamylaspartate or [¹⁴C]orotic acid, into uridine nucleotides (ΣUMP). Thus, regulation of de novo pyrimidine biosynthesis was demonstrated to occur at one or both of the first two reactions of the orotic acid pathway, those catalyzed by carbamylphosphate synthetase (CPSase) and aspartate carbamyltransferase (ACTase). The results of the present study provide evidence that ACTase alone is the site of feedback control by added uridine or one of its metabolites. Evidence demonstrating regulation of the orotic acid pathway by end-product inhibition at ACTase, but not at CPSase, includes the following observations: (a) addition of uridine (0.5 millimolar final concentration) inhibited the incorporation of NaH¹⁴CO₃ into ΣUMP by 80% but did not inhibit the incorporation of NaH¹⁴CO₃ into arginine; (b) inhibition of the orotate pathway by added uridine was not reversed by supplying exogenous ornithine (5 millimolar final concentration), while the incorporation of NaH¹⁴CO₃ into arginine was stimulated more than 15-fold when both uridine and ornithine were added; (c) incorporation of NaH¹⁴CO₃ into arginine increased, with or without added ornithine when the de novo pyrimidine pathway was inhibited by added uridine; and (d) in assays employing cell-free extracts prepared from 2-day-old squash roots, the activity of ACTase, but not CPSase, was inhibited by added pyrimidine nucleotides.     

The physiological mechanism of enhanced oxidizing capacity of rice (Oryza sativa L.) roots induced by phosphorus deficiency

Plants show various responses to phosphorus (P) deficiency. Root oxidizing capacity enhancement is one of adaptive mechanisms for rice (Oryza sativa L.) to P deficiency. However, it remains unclear how P deficiency enhances the root oxidizing capacity. In this study, rice seedlings were treated in P-deficient nutrient solution for different periods. Variations of reactive oxygen species (ROS), antioxidant enzyme activity, root lignin content, root porosity, root oxygen release, total oxidative substances and root structural changes in rice roots in response to P-sufficient and P-deficient treatments were investigated. Results indicated that P deficiency induced the production of H₂O₂ and O ₂ ^·? in roots significantly, which reached their maximum after 1- to 2-day P-deficient treatment. Interestingly, the endogenous total oxidative substances kept stable in rice roots. P deficiency increased the activities of peroxidase and superoxide dismutase by 89.5 and 51.8 % after 4-day P-deficient treatment, respectively. Moreover, one-day P deficiency elevated lignin accumulation. Root porosity of rice seedling under 2-day P-deficient treatment was 19.8 % higher than that under P-sufficient treatment. P deficiency also enhanced the release of both O₂ and total oxidative substances after 1- to 4-day P deficiency. In addition, results from electronic microscopy indicated that the thickness of root cell wall tended to increase after 2-day P-deficient treatment. Taken together, our results suggested that P-deficiency-induced enhancement of root oxidizing capacity in rice roots was probably associated with ROS production, antioxidant enzyme activity increment in root tissues, and the release of O₂ and oxidative substances from root inside to rhizosphere.     

Stimulation of root acid phosphatase by phosphorus deficiency is regulated by ethylene in Medicago falcata

Plants have developed numerous strategies to cope with phosphorus (P) deficiency resulting from low availability in soils. Evolution of ethylene and up-regulation of root secreted acid phosphatase activity are common for plants in response to P deficiency. To determine the role of ethylene in response of plants to P deficiency, we investigated the effects of ethylene precursor (1-amino cyclopropane-1-carboxylic acid, ACC) and ethylene synthesis antagonists (aminoethoxyvinylglycine AVG, cobalt, Co²⁺) on P concentrations in roots and shoots of Medicago falcata seedlings grown in P-sufficient (500 μM H₂PO₄⁻) and P-deficient (5 μM H₂PO₄⁻) solution. After transferring M. falcata seedlings from P-sufficient to P-deficient solution for 2 days, root P concentration was significantly reduced. The reduction in root P concentration was reversed by AVG and Co²⁺, and a similar reduction in root P concentration of seedlings exposed to P-sufficient solution was observed by ACC. Expression of high-affinity phosphate transporters (MfPT1, MfPT5) was enhanced by P-deficiency and this process was reversed by AVG and Co²⁺. There was a marked increase in activity of root acid phosphatase (APase) and expression of gene encoding APase (MfPAP1) under P-deficient conditions, and the increase in APAse activity and expression of MfPAP1 was inhibited by AVG and Co²⁺. APase activity and expression of MfPAP1 expression in seedlings grown in P-sufficient solution were enhanced by ACC. Root and shoot P concentrations were increased when organic phosphorus was added to the P-deficient solution, and the increase in P concentration was significantly inhibited by AVG and Co²⁺. These results indicate that ethylene plays an important role in modulation of P acquisition by possibly mobilizing organic P via up-regulating root APase activity and high-affinity phosphate transporters.     

Growth of Salvinia molesta as affected by water temperature and nutrition II. Effects of phosphorus level

The growth of Salvinia molesta D.S. Mitchell, as affected by phosphorus supply and water temperature, was studied in a greenhouse using controlled water temperature baths at 16, 19 and 22°C. For significant responses to the phosphorus treatments to be obtained it was found necessary to use P-deficient plant propagules (containing <0.01% P on a dry matter basis). For these plants the highest relative growth rate and dry matter production occurred at 22°C when they received 10.01 mg PO₄P l^?1, but this was not significantly different from that of plants receiving 1.01 mg PO₄-P l^?1. Over a period of 21 days for plants receiving 0.02 mg PO₄ l^?1 the biomass increased 4-fold at 19°C and 6-fold at 22°C. In contrast, for plants receiving 10.01 mg PO₄-P l^?1 biomass was increased 20-fold at 19°C and 32-fold at 22°C. At the latter temperature, when receiving 60.01 mg PO₄-P l^?1, plants concentrated up to 1.3% of phosphorus on a dry matter basis, suggesting a possible use as a biological filter and purifier of contaminated waters.     

Plant leaf area expansion and assimilate production in wheat (Triticum aestivum L.) growing under low phosphorus conditions

Under phosphorus deficiency reductions in plant leaf area have been attributed to both direct effects of P on the individual leaf expansion rate and to a reduced availability of assimilates for leaf growth. In this work we use experimental and simulation techniques to identify and quantify these processes in wheat plants growing under P-deficient conditions. In a glasshouse experiment we studied the effects of soil P addition (0–138 kg P₂O₅ ha^-1) on tillering, leaf emergence, leaf expansion, plant growth, and leaf photosynthesis of wheat plants (cv. INTA Oasis) that were not water stressed. Plants were grown in pots containing a P-deficient (3 mg P g^-1 soil) sandy soil. Sowing and pots were arranged to simulate a crop stand of 173 plants m^-2. Experimental results were integrated in a simulation model to study the relative importance of each process in determining the plant leaf area during vegetative stages of wheat. Phosphorus deficiency significantly reduced plant leaf area and dry weight production. Under P-deficient conditions the phyllochron (PHY) was increased up to a 32%, compared to that of high-P plants. In low-P plants the rate of individual leaf area expansion during the quasi-linear phase of leaf expansion (LER) was significantly reduced. The effect of P deficiency on LER was the main determinant of the final size of the individual leaves. In recently expanded leaves phosphorus deficiency reduced the photosynthesis rate per unit leaf area at high radiation (AMAX), up to 57%. Relative values of AMAX showed an hyperbolic relationship with leaf P% saturating at 0.27%. Relative values of the tillering rate showed an hyperbolic relationship with the shoot P% saturating at values above 0.38%. The value of LER was not related to the concentration of P in leaves or shoots. A morphogenetic model of leaf area development and growth was developed to quantify the effect of assimilate supply at canopy level on total leaf area expansion, and to study the sensitivity of different model variables to changes in model parameters. Simulation results indicated that under mild P stress conditions up to 80% of the observed reduction in plant leaf area was due to the effects of P deficiency on leaf emergence and tillering. Under extreme P-deficient conditions the simulation model failed to explain the experimental results indicating that other factors not taken into account by the model, i.e. direct effects of P on leaf expansion, must have been active. Possible mechanisms of action of the direct effects of P on individual leaf expansion are discussed in this work.     

Changes Induced by Low Oxygen Concentration in Photosynthetic and Respiratory CO2 Exchange in Phosphate-Deficient Bean Leaves

Effect of phosphorus deficiency on photosynthetic and respiratory CO₂ exchanges were analysed in primary leaves of 2-week-old bean (Phaseolus vulgaris L. cv. Golden Saxa) plants under non-photorespiratory (2 % O₂) and photorespiratory (21 % O₂) conditions. Low P decreased maximum net photosynthetic rate (P_Nmax) and increased the time necessary to reach it. In the leaves of P-deficient plants the relative decrease of P_Nmax at 2 % O₂ was larger than at 21 % O₂. The results suggested the influence of photorespiration in the cellular turnover of phosphates.     

Role of potassium and malate in nitrate uptake and translocation by wheat seedlings

Wheat seedlings (Triticum vulgare) treated with 1 mm KNO₃ or NaNO₃, in the presence of 0.2 mm CaSO₄, were compared during a 48-hour period with respect to nitrate uptake, translocation, accumulation and reduction; cation uptake and accumulation; and malate accumulation. Seedlings treated with KNO₃ absorbed and accumulated more nitrate, had higher nitrate reductase levels in leaves but less in roots, accumulated 17 times more malate in leaves, and accumulated more of the accompanying cation than seedlings treated with NaNO₃. Within seedlings of each treatment, changes in nitrate reductase activity and malate accumulation were parallel in leaves and in roots. Despite the great difference in malate accumulation, leaves of the KNO₃-treated seedlings had only slightly greater levels of phosphoenolpyruvate carboxylase than leaves of NaNO₃-treated seedlings. NADP-malic enzyme levels increased only slightly in leaves and roots of both KNO₃- and NaNO₃-treated seedlings. The effects of K⁺ and Na⁺ on all of these parameters can best be explained by their effects on nitrate translocation, which in turn affects the other parameters. In a separate experiment, we confirmed that phosphoenolpyruvate carboxylase activity increased about 2-fold during 36 hours of KNO₃ treatment, and increased only slightly in the KCl control.     

Phosphorus deficiency-induced reduction in root hydraulic conductivity in Medicago falcata is associated with ethylene production

Plants grown in phosphorus-deficient solutions often exhibit disruption of water transport due to reduction in root hydraulic conductivity (Lpr) and enhanced ethylene production. To uncover the relationship between the reduction in Lpr and increase in ethylene production, we investigated effect of phosphorus (P) deficiency on ethylene production and Lpr in legume plants of Medicago falcata L. There was an increase in ethylene production and a reduction of Lpr of M. falcata roots when M. falcata seedlings grown in P sufficient solutions (0.5 mM H₂PO₄^2?) were transferred to P-deficient solutions (5 μM H₂PO₄^2?). Antagonists of ethylene biosynthesis, CoCl₂ and aminoethoxyvinylglycine (AVG), abolished the P deficiency-induced ethylene production. Root hydraulic conductivity of M. falcata seedlings grown in P-sufficient solutions was insensitive to CoCl₂ and AVG, while the two chemicals enhanced Lpr for those grown in P-deficient solutions, suggesting that P deficiency-induced decrease in Lpr can be reversed by inhibiting ethylene production. Ethylene precursor 1-amino cyclopropane-1-carboxylic acid (ACC) and ethylene donor ethephon had greater inhibitory effect on Lpr of P-sufficient seedlings than that of P-deficient seedlings. Root hydraulic conductivity of P-sufficient seedlings was more sensitive to HgCl₂ than that of P-deficient seedlings. Taken together, these findings suggest that ethylene induced by P deficiency may play an important role in modulation of root hydraulic conductivity by affecting aquaporins in plants.     

Plant pathotoxins from Alternaria citri: The major toxin specific for rough lemon plants

A pathotype of the fungus Alternaria citri that attacks rough lemon plants produces several toxins in culture which specifically damage rough lemon and Rangpur lime plants. The major toxin produced, Toxin I, was by far the most potent compound (ED₅₀ = 30 ng/ml). Five other minor toxins were active at ED₅₀ levels greater than 1 μg/ml. On the basis of mass, ¹H and ¹³C NMR spectra and decoupling studies of Toxin I and derivative, Toxin I is a 19 carbon polyalcohol with an α-dihydropyrone ring. The γ-dihydropyrone tautomer was less predominant. Culture filtrates of A. citri also contained a biologically inactive, partially analogous, component possessing a tetrahydropyran ring. It probably arises from decarboxylation of Toxin I. Toxin I was highly specific and did not affect nonhost plants at 10 000 times the concentrations affecting rough lemon.     

Activity of ascorbate-dependent H2O2-scavenging enzymes and leaf chlorosis are enhanced in magnesium- and potassium-deficient leaves, but not in phosphorus-deficient leaves

The effect of phosphorus (P), potassium (K), and magnesium (Mg)deficiency on the development of leaf symptoms (chlorosis andnecrosis) and activities of ascorbate-dependent H₂O₂ scavengingenzymes (ascorbate peroxidase, monodehydroascorbate reductase,dehydroascorbate reductase, and glutathione reductase) was studiedin bean (Phaseolus vulgans) plants over a 12 d period of growthin nutrient solution. With increasing plant age Mg- and K-deficientleaves developed severe interveinal chlorosis and, accordingly,chlorophyll concentrations were reduced. However, in P-deficientleaves neither chlorosis nor necrosis appeared; the leaves remaineddark green and even at an advanced stage of P deficiency, chlorophyllconcentrations were still higher than those of control plants.In K- and, particularly, Mg-deficient leaves with an increasein severity of leaf chlorosis, activity of ascorbate-dependentH₂O₂- scavenging enzymes was progressively increased. In contrast,in P-deficient leaves, as in leaves of the control plants, activityof H₂O₂-scavenging enzymes remained at a low level over the12 d period. Accordingly, compared with P-deficient and controlplants, Mg- and K-deficient leaves with elevated anti-oxidativepotential showed much higher resistance to chlorophyll destructionby the herbicide paraquat. Elevated levels of H₂O₂-scavengingenging enzymes in Mg- and K-deficient leaves indicate a higherproduction of H₂O₂ and related toxic O₂ species. It Is suggestedthat in Mg- and K-deficient leaves, utilization of photoreductantsin CO₂ fixation is restricted because of impaired export andthus accumulation of photosynthates. This disturbance mightlead to enhanced photoreduction of molecular O₂ to toxic O₂species causing chlorophyll destruction (chlorosis), a processwhich is not important in P-deficient leaves where export ofsucrose is not affected. Key words: Bean, hydrogen peroxide detoxification, leaf chlorosis, magnesium nutrition, oxygen activation, phosphorus nutrition, potassium nutrition     

Photosynthate metabolism in the source leaves of n(2)-fixing soybean plants

Soybean plants (Glycine max [L.] Merr. cv Williams), which were symbiotic with Bradyrhizobium japonicum, and which grew well upon reduced nitrogen supplied solely through N₂ fixation processes, often exhibited excess accumulation of starch and sucrose and diminished soluble protein in their source leaves. Nitrate and ammonia, when supplied to the nodulated roots of N₂-fixing plants, mediated a reduction of foliar starch accumulation and a corresponding increase in soluble protein in the source leaves. This provided an opportunity to examine the potential metabolic adjustments by which NO₃⁻ and NH₄⁺ (N) sufficiency or deficiency exerted an influence upon soybean leaf starch synthesis. When compared with soybean plants supplied with N, elevated starch accumulation was focused in leaf palisade parenchyma tissue of N₂-fixing plants. Foliar activities of starch synthesis pathway enzymes including fructose-1,6-bisphosphate phosphatase, phosphohexoisomerase, phosphoglucomutase (PGM), as well as adenosine diphosphate glucose pyrophosphorylase (in some leaves) exhibited highest activities in leaf extracts of N₂-fixing plants when expressed on a leaf protein basis. This was interpreted to mean that there was an adaptation of these enzyme activities in the leaves of N₂-fixing plants, and this contributed to an increase in starch accumulation. Another major causal factor associated with increased starch accumulation was the elevation in foliar levels of fructose-6-phosphate, glucose-6-phosphate, and glucose-1-phosphate (G1P), which had risen to chloroplast concentrations considerably in excess of the K_m values for their respective target enzymes associated with starch synthesis, e.g. elevated G1P with respect to adenosine diphosphate glucose pyrophosphorylase (ADPG-PPiase) binding sites. The cofactor glucose-1,6-bisphosphate (G1,6BP) was found to be obligate for maximal PGM activity in soybean leaf extracts of N₂-fixing as well as N-supplemented plants, and G1,6BP levels in N₂-fixing plant leaves was twice that of levels in N-supplied treatments. However the concentration of chloroplastic G1,6BP in illuminated leaves was computed to be saturating with respect to PGM in both N₂-fixing and N-supplemented plants. This suggested that the higher level of this cofactor in N₂-fixing plant leaves did not confer any higher PGM activation and was not a factor in higher starch synthesis rates. Relative to plants supplied with NO₃⁻ and NH₄⁺, the source leaf glycerate-3-phosphate (3-PGA) and orthophosphate (Pi) concentrations in leaves of N₂-fixing plants were two to four times higher. Although Pi is a physiological competitive inhibitor of leaf chloroplast ADPG-PPiase, and hence, starch synthesis, elevated chloroplast 3-PGA levels in N₂-fixing plant leaves apparently prevented interference of Pi with ADPG-PPiase catalysis and starch synthesis.     

De Novo Purine Biosynthesis in Intact Cells of Cucurbita pepo

The capacity of intact cells of roots excised from summer squash plants (Cucurbita pepo L. cv Early Prolific Straightneck) to synthesize purine nucleotides de novo was investigated. Evidence that purine nucleotides are synthesized de novo included: (a) demonstration of the incorporation of [1-¹⁴C]glycine, [2-¹⁴C]glycine, NaH¹⁴CO₃, and H¹⁴COONa into total adenine nucleotides; (b) observation that the addition of azaserine or aminopterin, known inhibitors of de novo purine synthesis in other organisms, blocked the incorporation of these precursors into adenine nucleotides; and (c) demonstration that the purine ring synthesized from these precursors was labeled in a manner consistent with the pathway for de novo purine biosynthesis found in microorganisms and animal tissues. Under optimal conditions, the activity of this pathway in roots excised from 2-day-old squash plants was 244 ± 13 nanomoles (mean ± standard error, n = 17) NaH¹⁴CO₃ incorporated into ∑Ade (the sum of the adenine nucleotides, nucleoside and free base) per gram tissue during the 3-hour incubation period.

The possible occurrence of alternative enzymic reactions for the first steps of de novo purine biosynthesis was also investigated. No conclusive evidence was obtained to support the operation of alternative enzymic reactions in the intact cell of C. pepo.

     

Host-Specific Effects of Toxin from the Rough Lemon Pathotype of Alternaria alternata on Mitochondria

Host-specific toxin from the rough lemon (Citrus jambhiri Lush) pathotype of Alternaria alternata (ACR toxin) was tested for effects on mitochondria isolated from several citrus species. The toxin caused uncoupling of oxidative phosphorylation and changes in membrane potential in mitochondria from leaves of the susceptible host (rough lemon); the effects differed from those of carbonylcyanide-m-chlorophenylhydrazone, a typical protonophore. ACR toxin also inhibited malate oxidation, apparently because of lack of NAD⁺ in the matrix. In contrast, the toxin had no effect on mitochondria from citrus species (Dancy tangerine and Emperor mandarin [Citrus reticulata Blanco], and grapefruit [Citrus paradisi Macf.]) that are not hosts of the fungus. The effects of the toxin on mitochondria from rough lemon are similar to the effects of a host-specific toxin from Helminthosporium maydis (HMT) on mitochondria from T-cytoplasm maize. Both ACR and HMT toxins are highly selective for the respective host plants. HMT toxin and methomyl had no effect (toxic or protective) on the activity of ACR toxin against mitochondria from rough lemon.     

Decrease of Nitrate Reductase Activity in Spinach Leaves during a Light-Dark Transition

In leaves of spinach plants (Spinacia oleracea L.) performing CO₂ and NO₃⁻ assimilation, at the time of sudden darkening, which eliminates photosystem I-dependent nitrite reduction, only a minor temporary increase of the leaf nitrite content is observed. Because nitrate reduction does not depend on redox equivalents generated by photosystem I activity, a continuation of nitrate reduction after darkening would result in a large accumulation of nitrite in the leaves within a very short time, which is not observed. Measurements of the extractable nitrate reductase activity from spinach leaves assayed under standard conditions showed that in these leaves the nitrate reductase activity decreased during darkening to 15% of the control value with a half-time of only 2 minutes. Apparently, in these leaves nitrate reductase is very rapidly inactivated at sudden darkness avoiding an accumulation of the toxic nitrite in the cells.     

Oxidative stress in the root nodules of Phaseolus vulgaris is induced under conditions of phosphorus deficiency

One of the most adverse effects of phosphorus (P) deficiency on N₂-fixing legumes is the generation of harmful active oxygen species which cause oxidative stress. And although oxidative stress has been widely studied in roots and shoots of various plant species, it has not yet sufficiently been documented in bean nodules so far. In this study, two recombinant inbred lines RIL115 (P-deficiency tolerant) and RIL147 (P-deficiency sensitive) of common bean and Concesa (local variety) were inoculated separately with the reference strain R. tropici CIAT899, RhM₁₁ (R. gallicum) or RhM₁₄ (R. tropici); two local strains of the Marrakesh region of Morocco. Nodulated plants were grown under semi-hydroponic conditions with sufficient or deficient P supply and analyzed for their oxidative responses at the flowering stage. The results indicated that P-deficiency decreased the growth of shoots (48 %) and nodules (32 %), particularly with RhM₁₄ exhibiting the highest decrease (52 %) of nodulation. This constraint increased electrolyte leakage of nodules (40 %) as compared to leaves (20 %), especially for plants inoculated with RhM₁₄ and CIAT899. Moreover, high H₂O₂ and malondialdehyde contents were noticed in P-deficient nodules of RhM₁₄ and RhM₁₁. These variations were associated with peroxidase activity stimulation in P-deficient nodules induced by CIAT899 and RhM₁₄. In symbiosis with RIL115, these last strains exhibited the highest nodule phenol content. Overall, phenol content was mainly enhanced in P-deficient nodules (35 %) as compared to the leaves (16 %). It was concluded that the genotypes inoculated with CIAT899 and RhM₁₁ are relatively P-deficiency tolerant combinations as compared to those inoculated with RhM₁₄. Increase of oxidative stress in nodules rather than in leaves points to the need for further investigations of mechanisms that improve the root-nodule efficiency under adverse conditions.     

Effect of carbon dioxide on nitrate accumulation and nitrate reductase induction in corn seedlings

Exposure of the leaf canopy of corn seedlings (Zea mays L.) to atmospheric CO₂ levels ranging from 100 to 800 μl/l decreased nitrate accumulation and nitrate reductase activity. Plants pretreated with CO₂ in the dark and maintained in an atmosphere containing 100 μl/l CO₂ accumulated 7-fold more nitrate and had 2-fold more nitrate reductase activity than plants exposed to 600 μl/l CO₂, after 5 hours of illumination. Induction of nitrate reductase activity in leaves of intact corn seedlings was related to nitrate content. Changes in soluble protein were related to in vitro nitrate reductase activity suggesting that in vitro nitrate reductase activity was a measure of in situ nitrate reduction. In longer experiments, levels of nitrate reductase and accumulation of reduced N supported the concept that less nitrate was being absorbed, translocated, and assimilated when CO₂ was high. Plants exposed to increasing CO₂ levels for 3 to 4 hours in the light had increased concentrations of malate and decreased concentrations of nitrate in the leaf tissue. Malate and nitrate concentrations in the leaf tissue of seven of eight corn genotypes grown under comparable and normal (300 μl/l CO₂) environments, were negatively correlated. Exposure of roots to increasing concentrations of potassium carbonate with or without potassium sulfate caused a progressive increase in malate concentrations in the roots. When these roots were subsequently transferred to a nitrate medium, the accumulation of nitrate was inversely related to the initial malate concentrations. These data suggest that the concentration of malate in the tissue seem to be related to the accumulation of nitrate.     

Effect of salt stress on antioxidant system of plants as related to nitrogen nutrition

In plants of wheat (Triticum aestivum L.) grown in the media with nitrate (NO ₃ ^? plants), ammonium (NH ₄ ⁺ plants), and without nitrogen (N-deficient plants), the response to oxidative stress induced by the addition of 300 mM NaCl to the nutrient solution was investigated. Three-day-long salinization induced chlorophyll degradation and accumulation of malondialdehyde (MDA) in the leaves. These signs of oxidative stress were clearly expressed in NO ₃ ^? and N-deficient plants and weakly manifested in NH ₄ ⁺ plants. In none of the treatments, salinization induced the accumulation of MDA in the roots. Depending on the conditions of N nutrition, salt stress was accompanied by diverse changes in the activity of antioxidant enzymes in the leaves and roots. Resistance of leaves of NH ₄ ⁺ plants to oxidative stress correlated with a considerable increase in the activities of ascorbate peroxidase and glutathione reductase. Thus, wheat plants grown on the NH ₄ ⁺ -containing medium were more resistant to the development of oxidative stress in the leaves than those supplied with nitrate.