Iron Mediated Nitrate Reductase Activity in Different Parts of Young Maize Seedlings

Incubation of 5-d-old maize seedlings in the half-strength Hoagland's nutrient solution containing 10 mM KNO₃ with FeCl₃ or FeSO₄ (0.5 or 2.0 mM) caused a significant increase in nitrate reductase (NR) activity and slightly increased total protein content in root, shoot and scutellum. In case of root, NADPH:NR activity was inhibited contrary to the NADH:NR activity. In spite of NR activity, nitrate uptake was inhibited from 13 to 37 % by the iron. The results presented demonstrate an isoform specific, organ specific, and to some extent salt specific responses of NR to iron.     

Stomatal Reactions of Two Different Maize Lines to Osmotically Induced Drought Stress

Two maize lines differing in drought resistance were grown at different drought stress induced by polyethylene glycol (PEG 10 000) solutions with osmotic potentials of –0.20, –0.40 and –0.80 MPa in the semipermeable membrane system. During the five days soil water content decreased (from 0.43 to 0.29, 0.25 and 0.23 g cm^–3 for three PEG solutions, respectively) as well as leaf water potentials (_w; from – 0.54 to –0.76, –1.06 and –1.46 MPa). These values were not significantly different between the investigated lines, indicating that a controlled and consistent soil moisture stress was achieved. Soil drying induced an increase in the ABA content of leaves and xylem of both lines and the effects on stomatal conductance were greater in drought susceptible line (B-432) compared to drought resistant line (ZPBL-1304). To test possible difference in stomatal sensitivity to xylem ABA between lines and to assess any ABA vs. _w interaction, roots were fed with 10, 50 and 100 mmol m^–3 ABA solutions in another set of experiments. These results showed that manipulation of xylem ABA affected stomata of both lines similarly. Comparison of stomatal sensitivity to drought-induced and applied ABA demonstrated that drought treatment affected stomata of investigated lines by differentially increasing their sensitivity to xylem ABA, thus confirming an interaction between chemical signalling and hydraulic signalling.     

Thermoluminescence Investigation of Low Temperature Stress in Maize

The thermoluminescence (TL) emission of photosynthesising materials originates from the recombination of charge pairs created by a previous excitation. Using a recently described TL set-up the effect of chilling stress on TL bands occurring at positive temperatures (AG, B, and HTL) was investigated in intact leaves. The far-red irradiation of leaves at low, but non-freezing temperatures induced a TL band peaking at around 40–45 °C (AG band), together with a B band peaking between 20 and 35 °C. Low temperature stress first caused a downshift and a temporary increase in the AG band after 4 h at 0 °C in the light, then a decrease in the AG and B TL bands after 1 d at 0 °C in the light. This decrease was less pronounced in cold-tolerant genotypes and in those grown at acclimating temperatures. Furthermore, an additional band appeared above 80 °C after severe cold stress. This band indicates the presence of lipid peroxides. Thus TL is a useful technique for studying the effects of low temperature stress.     

Introduction of Alcohol Dehydrogenase in Lettuce Seedlings by Flooding Stress

Alcohol dehydrogenase (ADH), its isozyme profiles and ethanol concentration in lettuce (Lactuca sativa L.) seedlings subjected to flooding stress were determined. Flooding stress caused increases in ADH activity and ethanol concentration. By 48 h, ADH activity and ethanol concentration in the flooded seedlings increased 3.2- and 7.0-fold, respectively, in comparison with those in non-stressed seedlings. Five electrophoretically separable ADH bands were found in extract of the flooded seedlings, whereas only two or three ADH bands were found in extract of non-stressed seedlings. These results indicate that lettuce ADH may have a system of three-gene and six-isozyme, and the increase in ADH activity in the flooded seedlings may be due to increased synthesis of the enzyme.     

Effect of Osmotic Stress on Abscisic Acid Efflux and Compartmentation in the Roots of Two Maize Lines Differing in Drought Susceptibility

Roots of two Zea mays L. lines (drought-resistant Polj 17, and drought-susceptible F-2) were exposed to osmotic stress induced by sorbitol (osmotic potential –1.0 MPa). The following parameters were determined in cortex cells: membrane permeability for abscisic acid (ABA), ABA fluxes across membranes, pH values and ABA content in cytoplasm and vacuole. Osmotic stress induced different distribution of ABA within cell compartments in the investigated lines. ABA transport in the F-2 line occurred according to the intracellular pH gradient and the anion trap concept. In Polj 17, however, osmotic stress did not cause any significant effect on pH gradient and compartmental ABA content, but had a stimulating effect on ABA efflux from cytoplasm to apoplast and than via xylem to the leaf. These findings indicate different mechanisms of ABA transport between the investigated lines in response to osmotic stress.     

外源一氧化氮对镉胁迫下玉米幼苗根生长及氧化伤害的影响

以玉米幼苗为材料,通过在镉处理的同时补充外源一氧化氮(NO)供体硝普钠(SNP)及其类似物[K3Fe(CN)6]、以及NO消除剂,分析NO对植物耐镉性的影响,探讨NO在植物逆境胁迫响应中的作用及其机理。结果显示:添加20μmol·L-1 SNP能显著降低镉引发的玉米幼苗根生长抑制及根尖内源镉的积累,减少电解质的渗漏以及超氧化物自由基(O2.-)和过氧化氢(H2O2)的上升幅度,抑制超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、抗坏血酸过氧化物酶(APX)活性的增加,进一步提高镉胁迫下谷胱甘肽还原酶(GR)的活性。SNP的上述效应可被NO消除剂2-(4-羧基-2-苯基)-4,4,5,5-四甲基咪唑-1-氧-3-氧化物(cPTIO)所逆转,而SNP类似物K3Fe(CN)6的应用对上述反应几乎无影响,说明该反应具有NO特异性。研究表明,外源NO能够显著缓解镉胁迫对玉米幼苗生长造成的伤害,该缓解作用主要是通过降低植株体内内源镉积累和减轻镉诱发的氧化伤害来实现的。     

Production of Polyamines Is Enhanced by Endogenous Abscisic Acid in Maize Seedlings Subjected to Salt Stress

It is known that salt stress and exogenously applied abscisic acid （ABA） can enhance the polyamine content in plants and that salt stress itself can lead to an increase in endogenous ABA production. In the present study, the relationships between salt-induced ABA and polyamine accumulation were inves- tigated using ABA-deficient mutant （vp5/vp5） maize （Zea mays L.） seedlings and ABA and polyamine biosynthesis inhibitors. The results show that reduced endogenous ABA levels, as a result of either the mutation or by using a chemical inhibitor （sodium tungstate）, also reduced the accumulation of polyamines in salt-stressed leaves of maize seedlings. The polyamine synthesis inhibitors D-arginine and α- difluoromethylornithine also reduced the polyamine content of the leaves of maize seedling under salt stress. Both ABA and polyamine enhanced the dry weight accumulation of salt-stressed seedlings and also increased the activities of the two dominant tonoplast membrane enzymes, H^＋-ATPase and H^＋-PPase, when plants were under salt stress. The results suggest that salt stress induces an increase in endogenous ABA levels, which then enhances polyamine synthesis. Such responses may increase a plant＇s tolerance to salt.     

Biomass Enhancement in Maize and Soybean in Response to Glutamate Dehydrogenase Isomerization

The relationship between nutrient composition, crop biomass, and glutamate dehydrogenase (GDH) isoenzyme pattern was investigated in soybean (Glycine max) and maize (Zea mays) by monitoring the nutrient induced isomerization of the enzyme from the seedling stage to the mature crop. GDH was extracted from the leaves of the plants, and the isoenzymes were fractionated by isoelectric focusing followed by native polyacrylamide gel electrophoresis. The isomerization V_max values for soybean GDH, similar to maize GDH increased curvilinearly from 200 – 400 μmol mg⁻¹ min⁻¹ as the inorganic phosphate nutrient applied to the soil decreased from 50 − 0 mM. In soybean, combinations of N and K, P, or S nutrients induced the acidic and neutral isoenzymes, and gave biomass increases 25 – 50 % higher than the control plant. GDH isoenzymes were suppressed in soybean that received nutrients without N, K, or P and accordingly the biomass was about 30 % lower than the control. Treatment of maize with NPK nutrients increased the GDH V_max values from 138.9 at the vegetative to 256.4 μmol mg⁻¹ min⁻¹ at the reproductive phase, and suppressed the basic isoenzymes, but induced both the acidic and neutral isoenzymes thereby inducing seed production (27.0 ± 1.4 g per plant); whereas both the acidic and basic isoenzymes were suppressed in the control maize, and seeds did not develop. Simultaneous induction of the acidic, neutral, and basic isoenzymes of GDH indicated the occurrence of senescence. Therefore in maize and soybean, the induction of the acidic and basic isoenzymes of GDH led to the enhancement of biomass. This revised version was published online in July 2006 with corrections to the Cover Date.     

Effect of Water Deficit and Membrane Destruction on Water Diffusion in the Tissues of Maize Seedlings

We investigated diffusion of water in maize seedlings (Zea mays L. cv. Dnepropetrovskaya) following addition of polyethylene glycol (PEG) 6000 (osmotic potential –0.1 and –0.3 MPa) to the root medium by NMR method with pulsed gradient of magnetic field. Diffusion coefficients of different water phases in plant tissues (water of apoplast and vacuoles, water transported through the membranes) have been estimated from multicomponent decays of echo amplitude. Different signs of changes of water diffusion coefficients of fast and slow components of diffusional echo decay in roots and leaves under the influence of PEG-induced water deficits were shown. It has been supposed that under water deficit a sharing of water flows takes places through the different pathways (apoplastic, symplastic and transmembrane). In roots, 1-h water deficit increased the rate of fast diffusing water (water of apoplasm, vacuoles and, perhaps, water contained in intercellular endoplasm system), and decreased the rate of slowly diffusing water (water passing across the membranes). A long-term water deficit increased to a small extent the rate of water transmembrane transfer in root tissue. Leaf response to water stress was in the intensification of rate of transmembrane water transport that could be connected with the expression of water channels, and in the decrease of apoplastic water flow and flow along endoplasm. The possibility of estimation of plant tissue (membrane) integrity on the basis of diffusional data has been demonstrated.     

Nitrogen Deprivation Induces Changes in the Leaf Elongation Zone of Maize Seedlings

The influence of nitrogen deprivation on leaf development and the biomechanics of leaf growth were studied using maize (Zea mays L.) seedlings grown under low irradiance. Although the nitrogen deprivation had no significant effect on photosynthesis, the leaf length, the leaf area, and the total assimilation area of plants decreased. The mature size of the epidermal cells was not altered, while the cells of nitrogen-deprived plants reached their final length closer to the leaf base than the epidermal cells of control plants. Decreases in the length of the growing zone (from 50 to 30 mm) and in the maximum value of relative elemental growth rate (from 0.08 to 0.06 mm mm^–1 h^–1) were observed in the nitrogen deprived plants. The maximal value of growth velocity in the control treatment was higher along the elongation zone, except for the basal 20 mm, where there was no significant difference between the control and the N-deprived plants. The net deposition rates of water and dry matter were also affected by nitrogen deprivation: the values of these features decreased and the spatial position of the maximum of the deposition rates shifted towards the leaf base.     

Osmotic induced stimulation of the reduction of the viability dye 2,3,5-triphenyltetrazolium chloride by maize roots and callus cultures

Live cells can reduce colorless 2,3,5-triphenyltetrazolium chloride (TTC) to a red insoluble compound, formazan. Maize (Zea mays) callus, when osmotically stressed by 0.53 mol/L mannitol, produced 7-times or more formazan than untreated control callus. This result was seen with all osmotica tested and could not be attributed to differences in TTC uptake rate or accumulation, increased respiration rate as measured by O2 uptake, or to de novo protein synthesis. Increased formazan production could be detected after 2.5 h of exposure to osmotic stress and leveled off after 48 h of exposure. The increased formazan production was only detected when callus was moved from high osmotic medium to low osmotic, TTC-containing medium. Abscisic acid increased TTC reduction only when added in combination with 0.53 mol/L mannitol. Incubation of maize seedling roots with 0.53 mol/L mannitol also increased formazan production as seen visually. Further studies are needed to determine the cause of the increased formazan production. These results show that TTC viability measurements must be carefully evaluated with appropriate controls to confirm their validity.     

DNA methylation in the Alcohol dehydrogenase-1 gene of maize

Using a battery of methylation-sensitive restriction enzymes, cytosine methylation at 23 sites in a 7.6 kb region surrounding the Alcohol dehydrogenase-1 (Adh1) gene was measured in DNA prepared from immature maize cobs. Both the 5 upstream region and the entire coding region were hypomethylated in the two alleles examined. Methylation in Adh1 is independent of changes in Mutator transposable element methylation. The role of DNA methylation in Adh1 gene regulation is discussed.     

Characterization of topoisomerase I and II activities in nuclear extracts during callogenesis in immature embryos of Zea mays

We have characterized the topoisomerase I and II activities in nuclear extracts from immature embryos of Zea mays and the effect of the treatment with 2,4-dichlorophenoxyacetic acid (2,4-D) and abscisic acid (ABA). These extracts were shown to be essentially devoid of protease and nuclease activities and they were tested for their ability to relax supercoiled DNA, unknotting P4 DNA and catenate circular duplex DNA under catalytic conditions. Unknotting and catenation reactions are strictly magnesium- and ATP-dependent, but not the relaxation of circular supercoiled DNA allowing the detection of both topoisomerase I and II activities. Two cytotoxic drugs, camptothecin, a plant alkaloid that inhibits cukaryotic topoisomerase I, and epipodophyllotoxin VM-26 (teniposide) that inhibits topoisomerase II, have been assayed in our extracts showing similar inhibitory effects on topoisomerase enzymes. Alkaline phosphatase treatment of nuclear extracts abolishes both topoisomerase activities. Nuclear extracts from embryos treated with 2,4-D showed 200% increase on topoisomerase II activity as compared with untreated ones, but only residual activity was detected in ABA-treated embryos. Nuclear extracts from hormone-treated and untreated embryos showed similar topoisomerase I activity with deviations of less than 25%. These differences are discussed in terms of possible post-translational modifications of the enzymes associated with the increase in proliferation activity of calli.     

Proton Pumping in Growing Part of Maize Root: Its Correlation with 14-3-3 Protein Content and Changes in Response to Osmotic Stress

The spatial pattern of mitotic activity, cell elongation, rate of H+ fluxes, and 14-3-3 protein content were determined in Zea mays roots. We found that the regions along the apical part of the growing root conversely differ in their proton pumping activity. Higher rate of H+ efflux coincides with higher growth rate and correlates with increased 14-3-3 protein content in membrane preparations. The segment consisting of the root cap and the apical part of the meristem exerts net inward proton pumping, which can be inverted under fusicoccin treatment or osmotic stress. In the latter case, this inversion is accompanied by accumulation of 14-3-3 protein in plasma membranes. The results obtained highlight 14-3-3 protein as an obvious candidate for the fine regulation of plasma membrane H+-ATPase in root apex.     

Preferential Induction of Alcohol Dehydrogenase in Coleoptiles of Rice Seedlings Germinated in Submergence Condition

Difference in the growth response to submergence between coleoptiles and roots of rice (Oryza sativa L.) was investigated in 9-d-old rice seedlings. The coleoptile length in the submergence condition was much greater than that in aerobic condition, whereas the root length in the submergence condition was less than that in the aerobic condition. Alcohol dehydrogenase (ADH) activity in the coleoptiles in the submergence condition was much greater than that in the aerobic condition, but ADH activity in the roots in the submergence condition increased slightly. These results suggest that the preferential ADH induction in rice seedlings may contribute to the difference in the growth response between the coleoptiles and roots under low oxygen conditions.     

Heterogeneity of Maize Root Mitochondria from Plants Grown in the Presence of Ammonium

Mitochondria isolated from root tissue of maize plants grown on a modified Knop solution containing 10.9 mM nitrate ± 7.2 mM ammonium were purified on the discontinuous Percoll density gradient with polyvinylpyrrolidone (PVP) added. The presence of PVP allowed separation of several mitochondrial fractions of a different density. Contrary to mitochondria isolated from plants grown in the presence of nitrate alone, revealing only two fractions, the mitochondria from NH₄ ⁺/NO₃ ^–-plants were distributed in four fractions. Total amount of mitochondria, as well as specific activities of some nitrogen metabolism enzymes and tricarboxylic acid (TCA) cycle enzymes of all mitochondrial fractions, and respiratory activities of two lower density fractions isolated from plants grown on mixed nitrogen were higher in comparison to mitochondria from nitrate-grown plants.     

外源ABA对楸树幼苗NaCl胁迫的缓解效应及其生长生理响应特征

以2年生楸树(苏楸1号和008-1)扦插苗为材料,采用盆栽试验法,分析盐胁迫(0.5%NaCl)处理下楸树幼苗生长、生理的变化,并分析不同浓度外源ABA(15、25、35 mg/L)对盐胁迫(30 d)楸树幼苗的缓解效应及其生理生化特性,以探索重度盐胁迫下适合楸树幼苗生长的适宜外源ABA浓度,为增强盐碱地楸树的耐盐性、提高盐碱地的利用提供理论依据。结果显示:(1)0.5%NaCl胁迫下,两品种楸树幼苗叶片表现出不同程度的盐害症状,且‘苏楸1号’叶片盐害症状较‘008-1’严重;随胁迫时间延长,两品种楸树幼苗的相对电导率(REC)均呈先上升后下降的变化趋势,叶绿素(Chl)、相对含水量(RWC)均呈降低趋势,可溶性糖(SS)、可溶性蛋白(SP)、脯氨酸(Pro)以及超氧化物歧化酶(SOD)活性均呈先上升后下降趋势,但‘008-1’的REC显著低于‘苏楸1号’,Chl、RWC、SS、SP、Pro、SOD均显著高于苏楸1号,表明‘008-1’的耐盐性较‘苏楸1号’更强。(2)喷施外源ABA使得盐胁迫下‘008-1’楸树的苗高显著增加、新叶提前萌发,表明外源ABA在一定程度上能够缓解盐胁迫对楸树生长的影响;喷施外源ABA降低了盐胁迫下‘008-1’楸树幼苗叶片的REC,提高了Chl、RWC、SS、SP、Pro、SOD、过氧化物酶(POD)以及过氧化氢酶(CAT)活性,促进了内源激素生长素(IAA)、脱落酸(ABA)、赤霉素(GA3)以及玉米素核苷(ZR)的积累。研究表明,楸树品种‘008-1’的耐盐性更强;外源喷施适宜浓度ABA能够缓解盐胁迫对楸树幼苗生长的影响,降低幼苗叶片细胞膜透性,促进幼苗渗透调节物质的积累,增强渗透调节能力,并提高盐胁迫下幼苗的抗氧化酶活性,促进植物对内源激素含量的调节,从而提高楸树的耐盐性,且以25 mg/L ABA处理的效果最好。     

Colchicine Induced Embryogenesis in Maize

The present study involves in vitro androgenesis of Zea mays L. using anther culture. We tested combinations of single factors and their influence on microspore induction. Embryogenic induction of microspores within anthers in in vitro conditions was the best when combination of cold treatment, TIBA (0.1 mg l^–1) in media and colchicine (0.02% during first 3 days of culture) was applied, but colchicine alone can be factor, which can stimulate or initiate embryogenesis in anther culture of maize.     

Isozymes as Genetic Markers in Maize Breeding

The major objective of the research is to identify and locate quantitative trait loci (QTL_s) in the Yugoslav maize population. The plants (F2) were selected for the analysis at seedling stage and were selfed to obtain F3 generation. The analysis covered about 15 enzymes controlled by about 30 loci. The seeds of F3 family planted in the greenhouse for measuring some quantitative traits, recorded tasselling and silking during vegetation. At the end of vegetation grain yield, and some other quantitative traits of grain in F3 family were assessed. The relationship between marker loci and the loci for quantitative traits (QTL_s) were estimated by computerized statistical method.