Solute Heterogeneity and Osmotic Adjustment in Different Leaf Structures of Semi-Leafless Pea (Pisum sativum L.) Subjected to Water Stress

Abstract: Semi-leafless varieties of pea have considerable agronomic importance and it has been suggested that they may have a superior response to water deficits than conventional varieties. However, these varieties are poorly characterized from a physiological point of view and there is lack of a physiological basis for their supposed better performance under conditions of water deficit. Here, we describe the solute distribution in the different leaf structures of a semi-leafless pea variety ( Pisum sativum L.) under non-limiting water conditions and under water stress. A conventional variety was subjected to the same conditions for comparative purposes. A detailed study was carried out both at the tissue level and at the single cell level. In control conditions, epidermal vacuoles of tendrils showed a different ion distribution of those of the laminar leaf structures. However, under water deficit, only stipules of the semi-leafless variety showed a significantly higher capability to increase osmolarity. This occurred by accumulating potassium, magnesium and chloride to a higher extent than other leaf structures. The inability of performing an adequate osmotic adjustment in tendrils may be the cause of the lack of a better response to water deficit.     

Effect of nodulation on the nitrate assimilation in vegetative soybean plants

Summary Nitrate assimilation in the first trifoliate leaf of vegetative soybean plants (Glycine max L. Merr, cv Hodgson) was studied in relation to nodulation. Nodulated and non-nodulated plants were grown in a nitrate medium (4 mM). As a control nodulated plants were grown in a nutrient medium without combined nitrogen. This study included measurements of the acetylene reduction activity of the whole plant and of thein vitro nitrate reductase, glutamine synthetase and glutamate dehydrogenase activities in the first leaf and of the nitrate concentration. Nitrate accumulation and nitrate reductase activity were depressed in nodulated plants; root growth was decreased in the presence of nitrate. The relationships between nitrate assimilation and nodulation are discussed.     

Nitrate reductase activity and protein concentration of two populas clones

Nitrate reductase activity and protein percentage of various tree parts of two Populus clones were determined in relation to nitrate ion activity. Nitrogen was supplied as NH(4)NO(3) in a nutriculture system. Wisconsin-5 had significantly greater nitrate reductase activity than Tristis No. 1. Protein percentages of leaf plastochron index 10 leaves (tenth leaf below first leaf lamina exceeding 20 mm in length), bottom leaves, and roots in relation to nitrate ion activity were not appreciably different between clones. The nitrate reductase activity and protein percentage of Tristis No. 1 apex started to level off at the same nitrate ion activity, about 0.09 mm. In Wisconsin-5 apex protein percentage continued to increase at nitrate ion activities where nitrate reductase activity decreases sharply, suggesting that protein nitrogen was being supplied by ammonium ion. The difference in nitrate reductase activity between clones was probably due to genetically determined ability to synthesize nitrate reductase in response to nitrate ion. The expression of nitrate reductase activity was not an index of nitrogen assimilation ability but may be a useful index of growth potential when nitrate ion does not limit nitrate reductase synthesis.     

The Mechanism of Nitrate Accumulation in Pakchoi [Brassica Campestris L.ssp. Chinensis(L.)]

Decreased nitrate in vegetables can improve crop nitrogen utilization efficiency and lessen the human health risk caused by the reduction of nitrate to nitrite in vegetables. This paper studied the mechanisms of differences in nitrate accumulation and distribution within organs of two cultivars of pakchoi (Brassica campestris L.ssp. Chinensis (L.) previously screened in hydroponic experiments from 12 cultivars popularly grown in China at present. The two typical cultivars used in this experiment were Shanghaiqing with low nitrate accumulation and Liangbaiye 1 with high nitrate accumulation. There was no significant difference of total nitrate uptake but a significant difference in nitrate content existed between the two cultivars. Compared with Liangbaiye 1, Shanghaiqing showed a significantly higher photosynthetic rate and nitrate reductase activity. Determination of nitrate concentration (activity) in vacuoles with double-barrelled nitrate-selective microelectrodes showed that Shanghaiqing had lower vacuolar nitrate activity than Liangbaiye 1. Two putative nitrate reductase genes, nia1 and nia2, were amplified from the leaf blades of these two cultivars. Nia1 mRNA fragments (887 bp, accession numbers DQ082868 and DQ082869) were amplified using degenerate primer and nia2 mRNA fragment was amplified using one pair of generate primers designed according to DQ001901. Sequence analysis of DQ082868 and DQ082869 both showed 97% and 87% similarity with two nitrate reductase mRNA sequences of Brassica napus, accession numbers D38219 and D38220, respectively. The results of real time PCR to compare the relative expression of the putative nitrate reductase genes (nia1 and nia2) showed that Shanghaiqing had significantly higher expression level than Liangbaiye 1 and nia2 was significantly higher than nia1 in leaf blade and petiole. Both the nitrate reductase activity and the relative expression level of nia1 were in the order of leaf blade > root > petiole, while that of nia2 was leaf blade > petiole > root. There was no statistically significant difference of nitrate activity stored in vacuoles between the different organs of the two cultivars. It can be concluded that Shanghaiqing took up slightly less nitrate, but had significantly higher nitrate reductase activity in cytosol and had a higher relative expression of the putative nitrate reductase genes than Liangbaiye 1; this leads to the fact that Shanghaiqing has a lower nitrate content than Liangbaiye 1.     

Nitrogen and Nitrate Accumulation in Species Having Different Relationships Between Nitrate Uptake and Reduction

Species differ in the relationship of nitrate reductase activityto nitrate uptake. In Capsicum annuum different diurnal patternsof leaf nitrate reductase activity and nitrate uptake have beenreported. As a consequence, the relationship of free nitratein the plant to nitrate supplied has a higher level of significancethan has reduced nitrogen to nitrate supplied. In Zea mays ithas been reported that leaf nitrate reductase activity respondsdirectly to nitrate translocation to the leaf and in this speciesthe relationship of greatest significance is reduced nitrogencontent to nitrate supplied. In both species, and also in Cucumis melo, the proportion oftotal plant free nitrate and reduced nitrogen in the roots decreases,and in the stem increases, with increasing nitrate supplied. The accumulation of free nitrate in leaves is accompanied bya quantitatively different relationship between reduced nitrogenand dry weight compared to leaves not accumulating nitrate. Capsicum annuum. L., Cucumis melo L., melon, Zea mays L., maize, sweet corn, nitrate reductase, nitrate uptake     

半无叶型菜豌豆革质膜、甜度性状遗传研究及利用

对半无叶型菜豌豆革质膜、甜度性状的遗传规律进行了研究,结果表明：半无叶型菜豌豆革质膜受两对相对基因控制,F1表现大块革质膜,F2分离出大块革质膜、小块革质膜和无革质膜三种类型,分离比例为9：6：1,并经F2~3试验验证,属于基因互作的积加作用。糖度为数量性状遗传,F2单株间呈连续变异,符合正态分布。选育出的半无叶型菜豌豆新品种“须菜1号”高产、优质,其嫩荚鲜食,嫩卷须作为龙须菜,嫩茎叶也是一种优质蔬菜。     

The Effects of Irradiance on Uptake and Assimilation of Nitrate by Young Barley Seedlings

The nitrogen economy of barley plants growing in a range ofirradiances from full shade (less than 0·5 W m^–2)to 119 W ^m–2 has been examined by analysing levels oftotal, organic and nitrate nitrogen, and by determining nitratereductase activity in leaf extracts. It has been confirmed thatroot growth is reduced in low irradiances which are also associatedwith a lower level of total nitrogen in the plant, and hencewith a lower uptake of nitrate. In all parts of the plant thelevel of organic nitrogen is higher in high light intensitybut nitrate-nitrogen as a proportion of the total is greatestin low irradiances. In the first leaf accumulation of free nitrateis substantially greater in low irradiances. The data indicate a higher level of nitrate assimilation inhigh irradiances and nitrate reductase activity in leaf extractsis higher in such conditions. When the first leaf is shadednitrate reductase activity falls to undetectable levels afterabout 4 days, but in the case of the second leaf, where thisis shaded, some reductase activity is always found, althoughthis is substantially less than that in unshaded conditions. It is concluded that in vitro rates of nitrate reduction mayover-estimate nitrate assimilation determined as increase inorganic nitrogen.     

Inorganic nitrogen assimilation in plants of Austrlian rainforest communities

In a study of the plant communities of two Australian rainforests, it was found that pioner species had high levels of nitrate reductase (EC 1.6.6.1) and were predominantly leaf nitrate assimilators. Under- and over-storey species had low levels of shoot and root nitrate reductase activity, and many of them showed little capacity for nitrate reduction even when nitrate ions were freely available. Although closed-forest species have lower levels of nitrate reductase than those of gaps and forest margins, their total nitrogen contents were similar, suggesting the former utilize nitrogen sources other than nitrate ions. Glutamine synthetase (EC 6.3.1.2) was present in the leaves of all species examined. In the leaves of pioneer species the chloroplastic isoform of glutamine synthetase predominted, while in most of the species typical of closed-forest the cytosolic isoform accounted for at least 40% of total leaf activity. Low levels of chloroplastic glutamine synthetase were correlated with a low capacity for leaf nitrate reduction, and both are characteristic of many species that regenerate and grow for some time in shade. Low levels of chloroplastic glutamine synthetase imply that, in some of these woody plants, photorespiratory ammonia is re-assimilated via cytosolic glutamine synthetase.     

Growth and Photosynthesis of Mutant Pea Seedlings

The recessive of gene, producing tendrils in place of leaves,and the recessive st gene, reducing stipule size, produce phenotypesof pea that are termed leafless (afafstst) and semi-leafless(afafStSt). Photosynthesis and growth of these two types werecompared with the conventional phenotype (AfAfStSt) during thefirst 9 days of post-emergent growth. The conventional seedlingshowed faster net photosynthesis per unit dry weight than theleafless phenotype, whilst the semi-leafless seedlings wereintermediate. Differences in dark respiration were small butleafless seedlings had significantly higher rates soon afteremergence. Where the three phenotypes used were isogenic, except for ofand st, the rates of shoot growth were in the same ranking orderas net CO₂ uptake. With three other genotypes, representingthe three phenotypes, more similar shoot growth was found betweenthe conventional and semi-leafless phenotype, possibly becauseof compensating differences in embryonic axis size. The ratesof growth of roots and the rates of dry weight loss from thecotyledons showed no consistent differences between phenotypes. The results are discussed in relation to the potential for thesemi-leafless phenotype as an alternative to the conventionalphenotype for the dried pea crop. Pea seedling, Pisum sativum, leafless pea, photosynthesis, seedling growth     

豌豆半无叶突变体性状的遗传及在育种上的利用

以半无叶类型、普通类型豌豆为试验材料,对卷须性状的特征特性、遗传规律及在育种上的利用进行了研究。结果表明：半无叶类型豌豆品种卷须极其发达,能够在株间相互缠绕,形成棚架结构,显著地提高了品种的抗倒伏能力,同时改善了群体通风透光性能,显著提高了新品种的产量,是豌豆抗倒伏育种的重要原始材料。半无叶类型属单基因质量性状遗传,显隐性完全,卷须基因af和子叶颜色基因i表现连锁,位于1号染色体上,交换值为5.72%。 Abstract:Using semi-leafless pea and common pea,the authors studied tendril character,its inheritance law and how to use it in pea breeding.The results were as follows:Semi-leafless pea had well developed tendrils;They twined with each other and formed an arbor structure.This ideal structure had greatly increased lodging resistance capability,improved canopy′s air and light level,and remarkably increased new varieties yield.So,semi-leafless pea was one of the most important materials in lodging resistance breeding.Tendril gene,af,and seed color gene i,in semi-leafless pea was linked on chromosome Ⅰ,and cross-over value was 5.72%.     

Specificity for nicotinamide adenine dinucleotide by nitrate reductase from leaves

Preliminary work revealed that nitrate reductase in crude extracts prepared from leaves of certain corn genotypes as well as soybeans could utilize NADPH as well as NADH as the electron donor. Isoelectric focusing and diethylaminoethyl cellulose chromatography confirmed previous findings that NADH and NADPH activities could not be separated, which suggests the involvement of a single enzyme. Nitrate reduction with both cofactors varies with plant species, plant age, and assay conditions. The ability of the nitrate reductase from a given genotype to utilize NADPH was associated with the amount of NADPH-phosphatase in the extract. While diethylaminoethyl cellulose chromatography of plant extracts separated nitrate reductase from the bulk (90%) of the phosphatase and caused a decrease in the NADPH activity, the residual level of phosphatase was sufficient to account for the apparent NADPH nitrate reductase activity. Addition of KH₂PO₄ and KF, inhibitors of NADPH-phosphatase activity in in vitro assays, caused a drastic reduction or abolishment of NADPH-mediated nitrate reductase activity but were without effect on NADH nitrate reductase activity. It is concluded that NADPH-nitrate reduction, in soybean and certain corn genotypes, is an artifact resulting from the conversion of NADPH to NADH by a phosphatase and that the enzyme in leaf tissue is NADH-dependent (E.C.1.6.6.1).     

Nitrate reduction in ramets of a clonal plant Eichhornia crassipes responding to nitrate availability during clonal growth stage

Effect of nitrate availability on nitrate reduction was examined in inter-connected ramets of invasive clonal plant Eichhornia crassipes grown with two nitrate supply regimes during different clonal growth stage. Increase of nitrate availability accelerated nitrate reductase activity (NRA) in parent and offspring ramets of E. crassipes, and there was greatly different pattern in inter-connected ramets during clonal growth stage. Leaf NRA was lower in offspring than that in parent ramets in phase 1, while significantly higher leaf NRA in offspring ramets was detected during phase 2. The results indicated NRA in inter-connected ramets of E. crassipes was highly dependent on nitrate availability and growth stage.     

Characterization of Hormonal Complex in Pea Phenotypes Differing in Leaf Morphology

Two genotypes of the pea (Pisum sativum L.) with wild-type leaves (variety Orlovchanin, Af/Af genotype) and the afila morphotype (aphyllous variety Nord, af/af genotype) were compared in terms of growth performance and hormonal characteristics of different leaf parts and the whole plant. The replacement of leaflets by tendrils in the afila variety led to a reduction in total dry weight and the area of photosynthesizing surfaces. The loss of leaflets was partly compensated for by rapid expansion of stipules at early stages of plant development and by the hypertrophy of tendrils at later stages. The excessive development of stipules in afila plants was paralleled by the increase in IAA and cytokinin level in their tissues. The hypertrophied development of tendrils and chlorophyll accumulation in tendrils of afila plants was correlated with a high IAA and cytokinin content at a low ABA background level. The elevated content of ABA in tissues of wild-type plants was associated with the preferential development of leaflets and a larger transpiratory surface compared with those in the afila form. It is assumed that this feature ensures the turgescence of wild-type plants. The possible involvement of phytohormones in growth and morphogenesis of pea mutants is discussed.     

Nitrate content and nitrate reductase activity in Rumex obtusifolius L.

Summary With Rumex obtusifolius L., the influence of some environmental conditions on nitrate uptake and reduction were investigated. Nitrate concentrations of plant material were determined by HPLC, the activity of nitrate reductase by an in vivo test. As optimal incubation medium, a buffer containing 0.04 M KNO₃; 0.25 M KH²PO₄; 1.5% propanol (v/v); pH 8.0 was found. Vacuum infiltration caused an increase of enzyme activity of up to 40%.High nitrate concentrations were found in roots and leaf petioles. Nitrate reductase activity of these organs, however, was low. On the other hand, the highest nitrate reductase activity was observed in leaf laminae, which contained lowest nitrate concentrations.In leaves, nitrate content and nitrate reductase activity exhibited inverse diurnal fluctuations. During darkness, decreasing activities of the enzyme were followed by increasing nitrate concentrations, while during light the contrary was true. In petioles diurnal fluctuations in nitrate content were observed, too. No significant correlations with illumination, however, could be found.Our results prove that Rumex obtusifolius is characterized by an intensive nitrate turnover. Theoretically, internal nitrate content of the plant would be exhausted within a few hours, if a supply via the roots would be excluded.     

Foliar application of fungicide-opera alleviates negative impact of water stress in soybean plants

The modulatory effect of opera was investigated on the physiological and morphological aspects in soybean thriving in water stress environment. The data procured from current investigation indicated that water stress significantly declined the plant growth, leaf area in addition to photosynthetic efficiency, nitrate reductase activity and crop yield at various stages of growth such as vegetative (VS), flowering (FS) and pod filling stage (PFS). However, foliar application of opera (0.15%) was effective to enhance the the leaf area (42%), rate of photosynthesis (194%), and nitrate reductase activity (68%) at FS stage while the maximum enhancement in biomass accumulation (92%) and yield (119%) was observed at PFS stage as compared to their control plants. The opera is applied as foliar spray in field experiments to augment the assimilation of nitrogen and carbon in soybean which contributes to increased crop development and productivity under water stress conditions.     

Metabolism of Urtica dioica as dependent on the supply of mineral nutrients

Plants of Urtica dioica L., a very nitrophilous species, were grown in a nutrient solution containing either high (100%) or low (2%) nutrient supply. Part of these plants were subjected to a sudden switch from 100% to 2% or vice versa. Plant weight, sugar and organic nitrogen (both soluble and insoluble) and nitrate content were measured during growth. The activities of two nitrogen assimilating enzymes, nitrate reductase (NR) and glutamine synthetase (GS) were determined.
Growth of Urtica dioica was retarded at low nutrient supply. Root growth was limited by another factor than nitrogen. This was shown by a higher protein content. In the first period after a switch from 100% to 2%, redistribution of nitrogen from shoot to root could be demonstrated, and leakage from the root into the nutrient solution. It is suggested that in these conditions GS in the root reacted to this downward flux. Comparison with earlier findings on the less nitrophilous Plantago lanceolata showed that at 100% nutrient supply a correlation occurs between nitrate reduction and glutamine synthetase activity in that plant part which exported reduced nitrogen: the root in P. lanceolata and the shoot in U. dioica. In the importing plant part, glutamine synthetase was influenced by nitrate reduction as well as by imported reduced nitrogen.     

Influence of NO3 - and NH4 + nutrition on fermentation,nitrate reductase activity and adenylate energy charge of roots of Carex pseudocyperus L. and Carex sylvatica Huds. exposed to anaerobic nutrient solutions

The adenylate energy charge, production of ethanol and lactate, and nitrate reductase activity were determined in order to study the influence of different nitrogen sources on the metabolic responses of roots of Carex pseudocyperus L. and Carex sylvatica HUDS. exposed to anaerobic nutrient solutions. Determination of adenylates was carried out by means of a modified HPLC technique. Total quantity of adenylates was higher in Carex pseudocyperus than in Carex sylvatica under all conditions. In contrast, the adenylate energy charge was only slightly different between the species and decreased more or less in relation to the applied nitrogen source under oxygen deficiency. The adenylate energy charge in roots of plants under nitrate nutrition showed a smaller decrease under anaerobic environmental conditions than plants grown with ammonium or nitrate/ammonium. Roots of nitrate-fed plants showed a lower ethanol and lactate production than ammonium/nitrate- and ammonium-fed plants. Ethanol production was higher in C. pseudocyperus, formation of lactate was lower compared to that in Carex sylvatica. The activity of enzymes involved in fermentation processes (ADH, LDH and PDC) was enhanced significantly after 24 hours of exposure to anaerobic nutrient solutions in roots of both species. The induction of these enzymes was only slightly influenced by different nitrogen supply. In vivo nitrate reductase activity increased almost 3-fold compared to the aerobic treatment in both species and overcompensated loss of NADH reoxidation capacity caused by decrease of ethanol and lactate development. Induction of in vitro nitrate reductase activity was enhanced 313% in C. pseudocyperus and 349% in C. sylvatica under anaerobic environmental conditions and nitrate supply. These results indicate that nitrate may serve as an alternative electron acceptor in anaerobic plant root metabolism and that the nitrate-supported energy charge may be due to an accelerated glycolytic flux resulting from a more effective NADH reoxidation capacity by nitrate reduction plus fermentation than by fermentation alone.Abbreviations ADH alcohol dehydrogenase - AEC adenylate energy charge - DMSO dimethyl sulfoxide - EDTA ethylen diamine tetraacetic acid - HPLC high performance liquid chromatography - LDH lactate dehydrogenase - NRA nitrate reductase activity - PCA perchloric acid - PDC pyruvate decarboxylase - PVP polyvinylpyrrolidone - PVPP polyvinylpolypyrrolidone - TCA trichloroacetic acid, Tris-tris(hydroxymethyl)aminomethane     

Loci controlling nitrate reductase activity in maize: ultraviolet‐B signaling in aerial tissues increases nitrate reductase activity in leaf and root when responsive alleles are present

Environmental factors, such as ultraviolet‐B (UV‐B) irradiation, have the ability to affect pathways such as nitrogen metabolism. As fixed nitrogen is the keystone mineral nutrient that controls grain crop yield, any alteration in this cycle can be detrimental to plant productivity. Nitrate reductase enzyme activity is responsible for the reduction of nitrate to nitrite, and nitrate is the major form of nitrogen assimilated in plants. In maize (Zea mays L.) production, nitrate assimilation kinetics are important for both high‐ and low‐input agricultural systems. Nitrate reductase protein activity is controlled by phosphatases and kinases. Nitrate reductase activity is responsive to environmental signals such as light–dark cycles and UV‐B radiation, although the regulatory controls are not yet fully understood. We have determined the location of maize genetic factors that control nitrate reductase activity and the extent of contribution of each of these factors, both locally in the leaf tissue and via long‐distance signaling loci that affect root nitrate reductase activity upon leaf UV irradiation. In the IBM94 recombinant inbred mapping population, the loci controlling regulation of nitrate reductase activity under UV‐B map to different positions than the loci controlling nitrate reductase activity in unexposed plants.     

Nitrate Reductase Activity and Polyribosomal Content of Corn (Zea mays L.) Having Low Leaf Water Potentials

Desiccation of 8- to 13-day-old seedlings, achieved by withholding nutrient solution from the vermiculite root medium, caused a reduction in nitrate reductase activity of the leaf tissue. Activity declined when leaf water potentials decreased below −2 bars and was 25% of the control at a leaf water potential of −13 bars. Experiments were conducted to determine whether the decrease in nitrate reductase activity was due to reduced levels of nitrate in the tissue, direct inactivation of the enzyme by low leaf water potentials, or to changes in rates of synthesis or decay of the enzyme.     

Immunological comparisons of nitrate reductase of different plant species using monoclonal antibodies

Six monoclonal antibodies against different epitopes of maize leaf nitrate reductase were used to compare plant nitrate reductases in enzyme linked immunosorbent assay and enzyme activity inhibition tests. The number of cross-reacting antibodies was shown to vary with species according to phylogenetic classification, ranging from five (sugarcane) to one (dicotyledonous species). Cross-reactions were restricted to higher plant nitrate reductases.