Nitrite Uptake Patterns in Wheat Seedlings as Influenced by Nitrate and Ammonium

Nitrite and nitrate uptake by wheat (Triticum vulgare) from 0.5 mM potassium solutions both showed an apparent induction pattern characterized by a slow initial rate followed by an accelerated rate. The accelerated phase was more rapid for nitrate uptake, was initiated earlier, and was seriously restricted by the presence of equimolar nitrite. The accelerated phase of nitrite uptake was restricted by nitrate to a lesser extent. The two anions seem not to be absorbed by identical mechanisms. Ammonium pretreatments or prior growth with ammonium had relatively little influence on the pattern of nitrite uptake. However, prior growth with nitrate eliminated the slow initial phase and induced development of the accelerated phase of nitrite uptake. A beneficial effect was noted after 3 h nitrate pretreatment and full development had occurred by 12 h nitrate pretreatment. The evidence suggests that a small amount of tissue nitrite, which could be supplied either by absorption or by nitrate reduction, was specifically required for induction of the accelerated phase of nitrite uptake. Cycloheximide (2 μg ml^?1) seriously restricted development of the accelerated phase of nitrite uptake, but its effect was not as severe when it was added after the accelerated phase had been induced by prior exposure to nitrite or nitrate. However, translocation of ¹⁵N from the absorbed nitrite was sharply decreased under the latter conditions, indicating a difference in sensitivity of the uptake and translocation processes to cycloheximide. Potassium uptake was greater from KNO₃ than from KNO₂ and in both instances it was enhanced during the early stages of the accelerated phase of anion uptake. Moreover, addition of NaNO₃ to KNO₂ substantially increased potassium uptake. A coupling between anion and potassium uptake was therefore evident, but the coupling was not obligatory because the accelerated phase of nitrite uptake could occur in absence of rapid potassium uptake.     

Accelerated Nitrate Uptake in Wheat Seedlings: Effects of Ammonium and Nitrite Pretreatments and of 6-Methylpurine and Puromycin

The experiments reported herein had two objectives. One was to determine if the slow rate of nitrate uptake which occurs upon initial exposure of nitrogen-depleted wheat (Triticum vulgare cv. Knox) plants to nitrate was the result of insufficient reduced nitrogen. The second was to determine the impact of restrictions in ribonucleic acid or protein synthesis on both nitrate uptake and nitrate reduction. Pretreatments of 14-day-old seedlings for a few hours in ammonium or nitrite did not result in an enhancement of the initial slow rate of nitrate uptake. Growth for two additional weeks in ammonium also failed to eliminate the induction period. The evidence indicates that the presence of nitrate, rather than a product of its reduction, was required to initiate development of the accelerated rate of nitrate uptake. Puromycin (400 μg ml^?1) and 6-methylpurine (0.5 mM) prevented development of the accelerated phase of nitrate uptake. With both compounds, the relative restriction of nitrate uptake was greater than that of nitrate reduction as revealed by incorporation of ¹⁵N from labeled nitrate into reduced forms. The proportion of reduction which occurred in the root system under the imposed treatments could not be delineated precisely, preventing an unequivocal determination of the extent to which the two processes are coupled in the root system. The data nevertheless indicate nitrate reduction was closely associated with nitrate uptake. Accumulation of nitrate in the shoots was markedly restricted in presence of 6 methylpurine. This effect most likely was a result of a severe restriction in the translocation of nitrate into the xylem, rather than an increase in the reduction rate in the shoots.     

Nitrate Uptake and Assimilation by Wheat Seedlings during Initial Exposure to Nitrate

Nitrate uptake, reduction, and translocation were examined in intact, 14-day-old, nitrogen-depleted wheat (Triticum vulgare var. Knox) seedlings during a 9-hour exposure to 0.2 mm Ca (NO(3))(2). The nitrate uptake rate was low during the initial 3-hour period, increased during the 3- to 6-hour period, and then declined. By the 3rd hour, 14% of the absorbed nitrate had been reduced, and this increased to 36% by the 9th hour. Shoots accumulated reduced (15)N more rapidly than roots and the ratio of reduced (15)N to (15)N-nitrate was higher in the shoots. A significant proportion of the total reduction occurred in the root system under these experimental conditions. Accumulation of (15)N in ethanol-insoluble forms was evident in both roots and shoots by the 3rd hour and, after 4.5 hours, increased more rapidly in shoots than in roots.An experiment in which a 3-hour exposure to 0.2 mm Ca ((15)NO(3))(2) was followed by a 12-hour exposure to 0.2 mm Ca ((14)NO(3))(2) revealed a half-time of depletion of root nitrate of about 2.5 hours. A large proportion of this depletion, however, was due to loss of (15)N-nitrate to the ambient (14)N-nitrate solution. The remaining pool of (15)N-nitrate was only slowly available for reduction. Total (15)N translocation to the shoot was relatively efficient during the first 3 hours after transfer to Ca ((14)NO(3))(2) but it essentially ceased after that time in spite of significant pools of (15)N-nitrate and alpha-amino-(15)N remaining in the root tissue.     

Uptake of Nitrite by Neurospora crassa

Like the nitrate transport system, the nitrite uptake system in Neurospora crassa is induced by either nitrate or nitrite. This induction is prevented by cycloheximide, puromycin, or 6-methyl purine. The K(m) for nitrite of the induced nitrite uptake system is 86 muM, and the V(max) is 100 mumol of nitrite per g (wet weight) per h. Nitrite uptake is inhibited by metabolic poisons such as arsenate, dinitrophenol, cyanide, and antimycin A. No repression or inhibition of the nitrite transport system by ammonia, nitrate, or Casamino Acids was observed.     

Active Silicon Uptake by Wheat

The absorption of Si by wheat, Triticum aestivum L. ‘Yecora Rojo,’ followed Michaelis–Menten kinetics over a concentration range of 0.004–1.0 mM. K_m and V_max were determined using linear transformations and the calculated curve fitted the data closely. The absorption resulted in accumulation ratios of 200/1 or more. In keeping with that finding, this study also demonstrated that Si uptake by wheat is under metabolic control, being severely restricted by dinitrophenol (DNP) and potassium cyanide (KCN). Silicon uptake by wheat was not significantly affected by phosphate ions, but the chemical analog Ge exerted a direct competitive effect on Si uptake, and vice versa.     

Comparative Induction of Nitrate and Nitrite Uptake and Reduction Systems by Ambient Nitrate and Nitrite in Intact Roots of Barley (Hordeum vulgare L.) Seedlings

The induction by ambient NO3- and NO2- of the NO3- and NO2- uptake and reduction systems in roots of 8-d-old intact barley (Hordeum vulgare L.) seedlings was studied. Seedlings were induced with concentrations of NaNO3 or NaNO2 ranging from 0.25 to 1000 [mu]M. Uptake was determined by measuring the depletion of either NO3- or NO2- from uptake solutions. Enzyme activities were assayed in vitro using cell-free extracts. Uptake and reduction systems for both NO3- and NO2- were induced by either ion. The Km values for NO3- and NO2- uptake induced by NO2- were similar to those for uptake induced by NO3-. Induction of both the uptake and reduction systems was detected well before any NO3- or NO2- was found in the roots. At lower substrate concentrations of both NO3- and NO2- (5-10 [mu]M), the durations of the lag periods preceding induction were similar. Induction of uptake, as a function of concentration, proceeded linearly and similarly for both ions up to about 10 [mu]M. Then, while induction by NO3- continued to increase more slowly, induction by NO2- sharply decreased between 10 and 1000 [mu]M, apparently due to NO2- toxicity. In contrast, induction of NO3- reductase (NR) and NO2- reductase (NiR) by NO2- did not decrease above 10 [mu]M but rather continued to increase up to a substrate concentration of 1000 [mu]M. NO3- was a more effective inducer of NR than was NO2-; however, both ions equally induced NiR. Cycloheximide inhibited the induction of both uptake systems as well as NR and NiR activities whether induced by NO3- or NO2-. The results indicate that in situ NO3- and NO2- induce both uptake and reduction systems, and the accumulation of the substrates per se is not obligatory.     

The Uptake of Nitrite by the Diatom Phaeodactylum: Interactions between Nitrite and Nitrate

Freshly harvested ammonium-grown cells of the diatom Phaeodactylumtricornutum cannot take up nitrite but they acquire the abilityto do so after a period (about 3 h) of nitrogen deprivationunder conditions that allow photosynthesis. Addition of cycloheximide(1.0 µg ml^–1) prevents development as does incubationin Na⁺ or K⁺-free medium. Nitrite uptake is dependent on thepresence of Na⁺ in the medium and is inhibited by ammonium andnitrite uptake but it is concluded that the inhibition of nitriteuptake by nitrate is not of the competitive type.     

Kinetic Characterization of Nitrite Uptake and Reduction by Chlamydomonas reinhardtii

Kinetics of nitrite uptake and reduction by Chlamydomonas reinhardtii cells growing phototrophically has been studied by means of progress curves and the Michaelis-Menten integrated equation. Both uptake and reduction processes exhibited hyperbolic saturation kinetics, the nitrite uptake system lacking a diffusion component. Nitrite uptake and reduction showed significant differences in K_s for nitrite at pH 7.5 (1.6 versus 20 micromolar, respectively), optimal pH, activation energy values, and sensitivity toward reagents of sulfhydryl groups. K_s values for nitrite uptake were halved in cells subjected to darkness or to nitrogen-starvation. Nitrate inhibited nitrite uptake by a partially competitive mechanism. The same inhibition pattern was found for nitrite uptake by C. reinhardtii mutant 305 cells incapable of nitrate assimilation. The results demonstrate that C. reinhardtii cells take up nitrite via a highly specific carrier, probably energy-dependent, kinetically responsive to environmental changes, distinguishable from the enzymic nitrite reduction and endowed with an active site for nitrite not usable for nitrate transport.     

Kinetics of Manganese Uptake by Intact Citrus Seedlings

Uptake of manganese by intact citrus seedlings can be represented by three phases of a single, multiphasic isotherm in the range 10^?8M–2× 10^?4M. The phases are separated by marked jumps and the kinetic constants increase upon transition to higher phases.     

稀土元囊对小麦幼苗氮素吸收及同化的影响

稀土元素浸种能够促进小麦(Triticum aestivum L.)幼苗对No3-的吸收,提高硝酸还原酶活力。这些效应与稀土浓度有关,低浓度有促进作用,高浓度则有抑制作用。稀土元素处理还能促进小麦幼苗体内No3-的同化还原,使硝态氮含量降低,氨基氮含量增加,促进了氮素代谢过程。     

Influence of Nitrogen Supply on Uptake and Translocation of Strontium and Calcium in Wheat Seedlings

Supplying nitrate to N-depleted wheat seedlings (Triticum vulgare cv. Knox) stimulated the uptake and translocation of both ⁸³Sr and ⁴⁵Ca. Since the increase in ⁴⁵Ca accumulation was greater, the ⁸⁵Sr/⁴⁵Ca ratio in the plant tissue was decreased. Nitrate had relatively little influence on the amount of the divalent cations and ⁸⁵Sr/⁴⁵Ca ratio in the exchangeable fraction on the root surfaces, whereas it greatly increased the uptake into root tissue and translocation to shoots. The increase in percent transported to shoots occurred largely in the period of most rapid nitrate uptake. A split root study indicated that nitrate was ineffective when it was supplied to a different portion of the root system than that exposed to ⁸⁵Sr and ⁴⁵Ca. Nevertheless, ammonium and urea also increased the translocation of the two cations, indicating that the effects of nitrate could not entirely be ascribed to a direct effect of the nitrate anion.     

Potassium-Ammonium Uptake Interactions in Tobacco Seedlings

Short-term (< 12 h) uptake experiments were conducted with6–7-week-old tobacco (Nicotiana tabacum L. cv. Ky 14)seedlings to determine absorption interactions between K⁺ andNH₄⁺. At equal solution concentrations (0.5 mol m^–3) netK⁺ uptake was inhibited 30–35% by NH₄⁺ and NH₄⁺ uptakewas decreased 9–24%. Removal of NH₄⁺ resulted in completerecovery in K⁺ uptake rate, but NH⁴⁺ uptake rate did not recoverwhen K⁺ was removed. In both cases, inhibition of the uptakerate of one cation saturated as the concentration of the othercation was increased up to 0.5 mol m^–3. The relative effectof K⁺-NH₄⁺ interactions was not altered when Cl- was replacedwith SO₄^2–, but the magnitudes of the uptake rates wereless in the absence of Cl-. The V_max for NH₄⁺ uptake was reducedfrom 128 to 105 µmol g^–1 dry wt. h^–1 in thepresence of 0.5 mol m^–3 K⁺ and the K_m for NH₄⁺ doubledfrom 12 to 27 mmol m^–3 in the presence of K⁺. The resultsof these K⁺-NH₄⁺ experiments are interpreted as mixed-noncompetitiveinteractions. However, an enhanced efflux of K⁺ coupled to NH₄⁺influx via an antiporter cannot be ruled out as contributingto the decrease in net K⁺ uptake. Key words: Nicotiana tabacum, K⁺, NH₄⁺, Uptake interactions     

Nitrate Reductase Regulates Expression of Nitrite Uptake and Nitrite Reductase Activities in Chlamydomonas reinhardtii

In Chlamydomonas reinhardtii mutants defective at the structural locus for nitrate reductase (nit-1) or at loci for biosynthesis of the molybdopterin cofactor (nit-3, nit-4, or nit-5 and nit-6), both nitrite uptake and nitrite reductase activities were repressed in ammonium-grown cells and expressed at high amounts in nitrogen-free media or in media containing nitrate or nitrite. In contrast, wild-type cells required nitrate induction for expression of high levels of both activities. In mutants defective at the regulatory locus for nitrate reductase (nit-2), very low levels of nitrite uptake and nitrite reductase activities were expressed even in the presence of nitrate or nitrite. Both restoration of nitrate reductase activity in mutants defective at nit-1, nit-3, and nit-4 by isolating diploid strains among them and transformation of a structural mutant upon integration of the wild-type nit-1 gene gave rise to the wild-type expression pattern for nitrite uptake and nitrite reductase activities. Conversely, inactivation of nitrate reductase by tungstate treatment in nitrate, nitrite, or nitrogen-free media made wild-type cells respond like nitrate reductase-deficient mutants with respect to the expression of nitrite uptake and nitrite reductase activities. Our results indicate that nit-2 is a regulatory locus for both the nitrite uptake system and nitrite reductase, and that the nitrate reductase enzyme plays an important role in the regulation of the expression of both enzyme activities.     

低磷条件下不同基因型小麦幼苗对磷的吸收和利用效率及水分的影响

 选用在土壤磷水平为5～7mgP·kg-1土的条件下筛选出来的不同磷效率的4个冬小麦品种,采用盆栽试验研究了有效磷为3．2mgP·kg-1土时的磷效率、磷吸收效率、磷利用效率及土壤水分对这些指标的影响。结果表明：在有效磷很低的土壤上,“磷高效”品种小偃54和81(85)5—3—3—3在幼苗期并未表现出较高的磷效率。尽管这两个品种的磷吸收效率显著地高于NC37和京411,但由于它们的磷利用效率相对低于京411,从而使磷效率并未显著地提高。土壤水分对4个品种的磷吸收效率和利用效率均有显著影响。     

Nitrate Uptake and the Induction of Nitrate Reductase Activity in Wheat (Triticum aestivum L.) Seedlings

Nitrate uptake and the subsequent induction of in vivo nitratereductase activity in wheat were studied by investigating aeuploid and certain ditelosomic stocks which exhibited in vivoactivity significantly greater than that of the euploid. Thekinetics of nitrate uptake were investigated, but the high activitiesof the ditelosomics were not caused by increased uptake of nitrate,although ditelo-7B^L exhibited unusual uptake dynamics. Analysisof the induction of nitrate reductase activity revealed a biphasicgeneral pattern, with an initial rapid phase being followedby a slower but longer period of induction. The induction rateover the second period, although responsible for only a minorproportion of the total activity induced, was positively correlatedwith the final nitrate reductase level, unlike the rate overthe first induction period. Several stocks exhibited high inductionrates over one or other of the two phases, while ditelo- 1 A^sshowed an abnormal monophasic induction pattern. At the endof the second period of induction, nitrate reductase activitybecame more or less steady, except for activity fluctuationsassociated with the time of application of induction stimuli.     

The Influence of Arginine-specific Reagents on Nitrate Uptake by Corn Seedlings

The effects of three arginine-specific reagents on uptake were studied using corn seedlings (Zea mays L., GoldenCross Bantam). In the presence of borate, 0.25 mM 2, 3-butanedione(BD) and 1.0 mM 1, 2-cyclohexanedione (CHD) inhibited uptake by 76% and 68%, respectively, compared tothe controls. However, in the absence of borate, only 18% and38% inhibition was observed for 0.25 mM BD and 1.0 mM CHD, respectively.Similarly, 0.5 mM phenylglyoxal (PGO) resulted in 75% inhibition.The degree of inhibition of nitrate uptake exhibited a concentration-dependencewith respect to the reagents. Corn seedlings are 2- or 3-foldmore sensitive to BD than to PGO and CHD, respectively, presumablydue to the unfavourable steric effects of the benzal ring. Uptakeof was partially restored after removal of BD, CHD, and PGO from the uptake medium. No significant differenceswere observed for the ATPase and plasma membrane-associatedvanadate-sensitive H⁺-ATPase or K⁺-stimulated ATPase activityin homogenates and microsomal fractions prepared from corn seedlingswhich had been incubated for 2 h in the presence or absenceof 0.5 mM BD or 1.0 mM PGO. This suggests that inhibition ofnitrate uptake by the arginine-specific reagents was not causedby the indirect effect of their binding and inhibiting H⁺-ATPase.The fact that the arginine-specific reagents strongly inhibit uptake indicates that the transport system has arginine residues at or near the activesite. Key words: Arginine-specific reagents, borate effect, nitrate uptake, reversibility     

Copper Uptake by Ryegrass Seedlings; Contribution of Cell Wall Adsorption

Net copper uptake by cellulose discs, isolated root cell walls,and by live and dead roots of whole ryegrass seedlings, werestudied using ⁶⁴Cu as a tracer. Uptake by cellulose discs stoppedafter around 10 h while uptake by isolated root cell walls continuedfor up to 50 h. An initial fast phase of uptake consisting predominantlyof cell wall adsorption was similar in live and dead tissuefor up to 19 h. A slower phase of uptake continued for up to50 h, greater in live than in dead tissue, the slower phaseof uptake in live tissue consisting of both a living and a deadcomponent. Based on these results, an alternative to the desorptionmethod for estimating the apoplastic contribution to total copperuptake is presented. Time-course studies with seedlings givena variety of growing solution/uptake solution regimes, and therelationship between copper uptake and external copper concentration,for short (4.8 h) and long (42.4 h) term uptakes, suggest thatdiffering contributions of cell wall adsorption and symplasmicabsorption may be responsible for differing effects of externalcopper concentration on uptake being expressed by the same tissue.Water flux had little effect on total uptake of copper althougha possible effect on absorption could not be ruled out. Key words: Copper uptake, cell wall adsorption, ryegrass seedlings     

External Nitrogen and Carbon Source-Mediated Response on Modulation of Root System Architecture and Nitrate Uptake in Wheat Seedlings

Nitrogen uptake efficiency is an important component trait that could be targeted for improving nitrogen use efficiency of crop plants. To understand the responses of different nitrate transport systems and the influence of root system architecture on nitrate uptake under limited nitrate conditions in wheat (Triticum aestivum L.) at the seedling stage, we studied nitrate uptake, root system architecture, and expression of different nitrate transporter genes in induced and non-induced wheat seedlings. Further, effects of inclusion of sucrose and two amino acids (glutamine and asparagine) in induction medium on these parameters were also studied. We observed that the induced wheat root system took up more nitrate as compared to non-induced root system in a dose-dependent manner. Gene expression of both high- and low-affinity nitrate transporter gene showed differential expression in the induced root tissues, as compared to non-induced tissues, depending on the concentration of nitrate present in induction medium. External nutrient media containing sucrose, glutamine, and asparagine reduce nitrate concentration in both root and shoot tissues and also influence the gene expression of these transporters. Our observations indicate that upon induction with milder external nitrate concentrations, the root architecture is modulated by changing overall lateral root size and 1st order lateral root numbers along with activation of nitrate transporters which acquire and transport nitrate in roots and shoots, respectively, depending on the carbon and nitrogen source available to seedlings.

     

Role of Nitrate and Nitrite in the Induction of Nitrite Reductase in Leaves of Barley Seedlings

The role of NO₃⁻ and NO₂⁻ in the induction of nitrite reductase (NiR) activity in detached leaves of 8-day-old barley (Hordeum vulgare L.) seedlings was investigated. Barley leaves contained 6 to 8 micromoles NO₂⁻/gram fresh weight × hour of endogenous NiR activity when grown in N-free solutions. Supply of both NO₂⁻ and NO₃⁻ induced the enzyme activity above the endogenous levels (5 and 10 times, respectively at 10 millimolar NO₂⁻ and NO₃⁻ over a 24 hour period). In NO₃⁻-supplied leaves, NiR induction occurred at an ambient NO₃⁻ concentration of as low as 0.05 millimolar; however, no NiR induction was found in leaves supplied with NO₂⁻ until the ambient NO₂⁻ concentration was 0.5 millimolar. Nitrate accumulated in NO₂⁻-fed leaves. The amount of NO₃⁻ accumulating in NO₂⁻-fed leaves induced similar levels of NiR as did equivalent amounts of NO₃⁻ accumulating in NO₃⁻-fed leaves. Induction of NiR in NO₂⁻-fed leaves was not seen until NO₃⁻ was detectable (30 nanomoles/gram fresh weight) in the leaves. The internal concentrations of NO₃⁻, irrespective of N source, were highly correlated with the levels of NiR induced. When the reduction of NO₃⁻ to NO₂⁻ was inhibited by WO₄²⁻, the induction of NiR was inhibited only partially. The results indicate that in barley leaves NiR is induced by NO₃⁻ directly, i.e. without being reduced to NO₂⁻, and that absorbed NO₂⁻ induces the enzyme activity indirectly after being oxidized to NO₃⁻ within the leaf.