Flower Induction by Suppression of Nitrate Assimilation in Lemna paucicostata 6746

In vitro activity of nitrate reductase was studied in Lemnapaucicostata 6746 grown on modified Hoagland medium supplementedwith 1% sucrose, containing various inhibitors. Copper, silver,tungstate or cyanide which induces daylength-independent flowering,inhibited the nitrate reductase activity, but azide which doesnot induce daylength-independent flowering did not. Molybdate-deficientmedium induced flowering, and inhibited nitrate reductase activity.Lowering of nitrate level of the medium also induced daylength-independentflowering. These results suggest that the suppression of nitrate assimilationcauses daylength independent flowering in Lemna paucicostata6746, and that one of the flower-inducing actions of the copper,silver, tungstate, cyanide or the deletion of molybdate is tosuppress the nitrate assimilation. (Received June 26, 1985; Accepted October 30, 1985)     

Inhibition of Flowering in the Long-Day Plant Lemna gibba G3 by Hutner's Medium and its Reversal by Medium Modification

The long-day plant Lemna gibba G3 flowers normally in E medium(Hoagland-type medium plus 30 µM EDTA) but in 0.5 H mediumthere is no flowering. Ammonium is present in 0.5 H medium andis known to inhibit flowering in L. gibba G3, but even in NH₄⁺-free0.5 H medium there is virtually no flowering under continuouslight. Increasing the phosphate concentration of the NH₄⁺-free0.5 H medium from 1.15 ITIM to 12 or 16 mM results in substantialflowering. Decreasing the EDTA concentration from 850 µIMto 250 µM, or raising the nitrate concentration from 4mM to 12 mM, results in only a small increase in flowering.If the decrease in EDTA and increase in nitrate are combinedwith the increase in phosphate, however, the flowering responseis nearly as good as that obtained using E medium. Thus, withthese three changes the inhibitory effect of NH₄⁺free 0.5 Hmedium for flowering in L. gibba G3 is almost completely reversed In the above studies flowering was not limited by daylength.When plants were grown on E medium under an 11 hour daylengthwhere flowering is limited by daylength, decreasing the phosphateconcentration in the medium reduced flowering, but increasingthe phosphate concentration in the medium did not stimulateflowering. Thus, when flowering is limited by daylength, highphosphate will not cause flowering, but a certain level of phosphateappears to be necessary for the expression of photoinductionunder long days. (Received January 14, 1986; Accepted June 24, 1986)     

Xylem exudation is related to nitrate assimilation pathway in detopped maize seedlings: use of nitrate reductase and glutamine synthetase inhibitors as tools

The xylem exudation of detopped 7-d-old seedlings of Zea maysL. doubled when KCI was present in the root medium comparedto seedlings maintained on water. It was further enhanced whenKCI was replaced by nitrogen compounds such as nitrate, ammoniumand glutamine. The role of the nitrate assimilation pathwayon the enhancement of xylem exudation rate was investigatedusing tungstate, an inhibitor of nitrate reductase (NR) activity,and phosphinothricin or methionine sulphoximine, inhibitorsof glutamine synthetase (GS) activity. The sap levels of NO₃^–,NH₄⁺, glutamine, and asparagine was used to ascertain the invivo inhibition of both enzymes. The tungstate effects werealso checked by measuring leaf in vitro NA activity and NR proteincontent. Xylem exudation rate of detopped seedlings fed withKNO₃ decreased when the nitrate assimilation pathway was blockedeither at the NR or at GS sites. This decrease was preventedwhen urea (acting as NH₄⁺ supply) was given simultaneously withtungstate. KNO₃ does not act directly on exudation, but throughthe involvement of NH₄⁺. The involvement of glutamine was alsoshown since GS inhibition resulted in a cancellation of theenhancing effect of KNO₃ on exudation. As change of exudationrate was not linked to change in sap osmolarity, it is assumedthat the assimilation chain could modify root water conductance.The role of glutamine was discussed. Key words: Exudation, maize, nitrate, conductance, NR, GS     

Effect of nitrate concentration in the medium on the flowering of Lemna paucicostata 6746

Addition of copper or tungstate to or exclusion of molybdenumfrom M-sucrose medium induced long-day flowering in Lemna paucicostata6746 provided the medium contained sufficient nitrate. By contrast,ferricyanide, cyanide or silver induced long day flowering evenin nitrate-deficient, M-sucrose medium. (Received August 26, 1977; )     

A Nitrate-Inducible Plasma Membrane Protein of a Marine Alga, Heterosigma akashiwo

The rate of nitrate uptake by Heterosigma akashiwo cells thathad been cultured in medium with nitrate or ammonium ions asthe source of nitrogen was measured using¹⁵NO_3– The ratioof ¹⁵N/¹⁴N increased dramatically in nitrate-grown cells. Inammonium-grown cells, the ratio of ¹⁵N/¹⁴N did not increasefor 3 h but then it began to increase. Even when nitrate reductaseactivity was inhibited by tungstate, nitrate-grown cells couldtake up nitrate. Plasma membranes from nitrate-grown and ammonium-grown cellswere purified by the silica-microbead method, and polypeptidesassociated with the membranes were analyzed by SDS-PAGE andimmunostaining. A major polypeptide with a molecular mass of26 kDa appeared 3 h after the transfer of ammonium-grown cellsto nitrate-containing medium, and it disappeared 2 d after thetransfer of nitrate-grown cells to ammonium-containing medium.The 26 kDa polypeptide also appeared when cell growth shiftedfrom the logarithmic phase to the stationary phase and the ammoniumcontent of the medium decreased, even when the cells were culturedin ammonium-containing medium. (Received April 10, 1992; Accepted July 30, 1992)     

Flower-inducing Activity of Vitamin K in Lemna paucicostata

Vitamins K₁ K₃ and K₅ induced flowering in Lemna paucicostata151, a short-day plant, cultured in 1/10 strength M medium (1/10M medium) under continuous light, and their activity was greatlyintensified by simultaneous application of benzyladenine. Themost active of these was vitamin K₅ L. paucicostata 6746 ismore sensitive to vitamin K₅ than strain 151, but the effectof vitamin K₅ on strain 6746 was not intensified by benzyladenine.The flower-inducing activity of vitamin K₅ was intensified bythe addition of benzoic acid in both strains and by the additionof copper or ferricyanide in Strain 6746, when these chemicalswere added at such low concentrations that they would scarcelyinduce flowering. In strain 6746, vitamin K₅ added to 1/10 M had little effecton flowering under a subcritical photoperiod, while it clearlyinduced flowering under continuous light. In this strain, vitaminK₅ added to full strength M medium, in which this plant wasmore sensitive to short photoperiods than in 1/10 M medium,did not induce flowering even under continuous light, and wasrather inhibitory under short photoperiods. (Received August 14, 1984; Accepted October 16, 1984)     

Suppression of long-day flowering by nitrogenous compounds in Lemna perpusilla 6746

The long-day flowering of Lemna perpusilla 6746 on an SH inhibitor-containingmedium was inhibited by the application of ammonium ion to themedium. Ammonium ion not only suppressed long-day flowering,but relieved the inhibition of vegetative growth caused by theinhibitors. Nitrite, casamino acids, glutamine and asparaginehad a similar effect, suggesting that the inhibition of long-dayflowering by ammonium ion is not a direct effect of the ion.Most amino acids, with the exception of glutamate and aspartate,also prevented long-day flowering, but their effects on vegetativegrowth varied. No qualitative differences in amino acid compositionwere observed among plants cultured on media containing nitrate,nitrite or NH4₄NO₃as the sole nitrogen source. However, theamounts of free and total amino acids werehigher in plants fedwith nitrite or NH₄NO₃ than in those fed with nitrate. Thissuggests that the inhibition of long-day flowering by ammoniumand nitrite can be ascribed to increased nitrogen metabolism. Though decreased activity by SH inhibitors of nitrate reductase(SH enzyme) is assumed to result in long-day flowering by loweringthe nitrogen metabolism, lowering the nitrogen level in M mediumdid not bring about floral initiation in the absence of SH inhibitors. (Received January 7, 1975; )     

Effects of some SH-inhibitors and EDTA on flowering in Lemna perpusilla 6746

Lemna perpusilla 6746, a short-day duckweed, flowered undercontinuous illumination on M-sucrose medium containing CuSO₄,AgNO₃ and HgCl₂, which are SH-inhibitors. The optimum concentrationsof CuSO₄, AgNO₃ and HgCl₂ were 5, 1 and 20 µM, respectively.Other metal ions tested were ineffective, but at least two otherSHinhibitors, potassium ferricyanide and iodoacetamide, alsoinduced long-day flowering at the concentrations of 0.1-1 µM. Adding 50 µM EDTA to the medium prevented the effect ofcupric ion, but not that of other SH-inhibitors. EDTA at 200µM induced some long-day flowering when added to a mediumwith no SH-inhibitors. It also permitted some flowering whenadded together with cupric ion, and accelerated flowering inthe presence of the other SHinhibitors listed above. EDTA andSH-inhibitor effects appeared to be additive. (Received May 25, 1973; )     

The In Vitro Flowering of Kalanchoe blossfeldiana Poellniz: I. ROLE OF CULTURE CONDITIONS AND NUTRIENTS

Dickens, C. W. S. and Van Staden, J. 1988. The in vitro floweringof Kalanchoe blossfeldiana Poellniz. 1. Role of culture conditions.—J.exp. Bot. 39: 461—471. Nodal explants of Kalanchöe blossfeldiana Poellniz. werecultured in vitro on a low nutrient hormone-free medium. Floweringwas achieved in response to short-day inductive cycles. Thissystem was used to test the influence, on the flowering response,of a variety of culture conditions and media. Reduced vesseland medium volume both inhibited flowering, as did renderingthe vessel impervious to gasses. Nitrogen in the form of NH₄NO₃and KNO₃ promoted flowering and vegetative growth in differentways. Increasing sucrose content in the medium caused some increasein the flowering response and in leaf anthocyanin production,but inhibited most aspects of vegetative growth. All of theseaspects are discussed in relation to the induction and evocationof flowering. Key words: Kalanche, flowering, in vitro     

Flowering responses of Lemna perpusilla and L. gibba in relation to nitrate concentration in the culture medium

Flowering responses of Lemna perpusilla strain 6746, a short-dayplant, and L. gibba strain G3, a long-day plant, to nitrateconcentration in Hoagland's type medium with or without EDTA,were compared. Maximum flowering of L. perpusilla under SD occurredat higher nitrate concentrations than did colony proliferation.Even under CL, L. perpusilla grown at sub-optimal nitrate concentrationsfor colony proliferation, flowered irrespective of the presenceof EDTA which reduces flowering. Unlike L. perpusilla, L. gibba failed to flower under SD atany nitrate concentration whether or not EDTA was added. UnderCL, however, L. gibba flowered at almost any nitrate concentrationwith or without EDTA. Double optima for nitrate concentrationwas exhibited in the presence of EDTA; optimal concentrationfor colony proliferation came between the two optima for flowering. We concluded that the nitrogen level of the medium is importantin regulating flowering of duckweeds, and that the effect ofEDTA, if any, may primarily be on colony proliferation and onlysecondarily or antagonistically on flowering. ¹ Present address: Institute for Agricultural Research, TohokuUniversity, Sendai 980, Japan. (Received September 25, 1971; )     

Soluble Dissimilatory Nitrate Reductase Containing Cytochrome c from a Photodenitrifier, Rhodopseudomonas sphaeroides forma sp. denitrificans

Dissimilatory nitrate reductase [nitrite: (acceptor) oxidoreductase.EC 1.7.99.4 [EC] ] from a denitrifying photosynthetic bacterium, Rhodopseudomonassphaeroides forma sp. denitrificans proved to be a soluble enzymethat could be purified 47-fold. It was labile, and containedcytochrome c, based on the results of specific staining forheme on polyacrylamide gel electrophoresis and on its absorptionspectrum. Its physiological molecular weight was determinedto be 112k, although heterogeneous molecular weights of 112k,100k, 73k and 60k were found for different preparations. Theoptimum for enzyme activity was about pH 6, and the K_m for thenitrate was 1.6 mM. As an electron donor, benzyl viologen wasvery good; but NADH, NADPH, FAD, FMN, cytochromes b₂ and c₂,dichlorophenolindophenol and phenazine methosulfate were noteffective. Bathophenanthroline and thiocyanate inhibited enzymaticactivity. The addition of 1 mM tungstate to the growing culturein place of molybdate decreased the nitrate reductase in thecells, but a further addition of 1 mM molybdate stopped it.This nitrate reductase is believed to be a molybdo-iron proteinsimilar to the enzymes from other bacteria with a nitrate respiratingability. (Received February 29, 1980; Accepted January 29, 1981)     

Seedling Nitrate Reductase Activity and Nitrate Partitioning in Maize (Zea mays L.) Strains Divergently Selected For Post-anthesis Leaf-Lamina Nitrate Reductase Activity

Two experiments were conducted to evaluate the effects of phenotypicrecurrent selection for high and low post-anthesis leaf-laminain vivo NRA on nitrate uptake, nitrate partitioning and in vitroNRA of seedling roots and leaves. In Experiment 1, intact plantsof cycle 0, 4, and 6 of the high and low NRA strains were grownon NH₄-N for 11 d, then exposed to 1.0 mol m^–3 KNO₃, andcultures sampled at 6 h and 28 h (induction and post-inductionperiods). Nitrate uptake, tissue nitrate concentration and invitro NRA were determined. The pattern of response to selectionin seedling leaf NRA was similar to that observed for in vivoNRA of field grown plants. Leaf NRA increased between 6 h and28 h. Root NRA was not affected by selection or sampling time.Treatments differed in total fresh weight but not in reductionor uptake of nitrate per unit weight, indicating a lack of correspondencebetween NRA and reduction and supporting the idea that concomitantreduction by NR is not obligatorily linked to nitrate influxin the intact plant. In Experiment 2, dark-grown plants of cycle 0, and 6 of thehigh and low NRA strains were cultured without N, detopped onday 6, transferred the following day to 0-75 mol m^–3 KNO₃and sampled at 6 h and 28 h. In contrast to Experiment 1, selectionpopulations differed in nitrate reduction and root NRA, whichby 28 h reached higher average levels than root NRA of intactplants. Translocation and reduction were inversely related amongstrains within each sampling time. The high level of translocationin detopped plants of the low NRA strain was difficult to reconcilewith its low leaf NRA level of Experiment 1. It is suggestedthat nitrate transport in detopped roots is altered relativeto the intact system in a way which permits greater NRA inductionand nitrate reduction. The results indicate that nitrate partitioningby detopped root systems should be interpreted with caution. Key words: Zea, nitrate reductase activity, nitrate uptake, nitrate reduction, nitrate partitioning, selection     

Flower-inducing Effect of Benzoic and Salicylic Acids in Various Strains of Lemna paucicostata and L. minor

The flower-inducing activities of benzoic and salicylic acidsadded to the medium differ with the species (Lemna paucicostataand L. minor), and even with the strains used. The type andpH of the medium used, full or 1/10 strength M medium at pH3.8, 4.4 or 5.1, or 1/2 or 1/20 strength NH₄⁺-free Hutner'smedium at pH 5.0, 6.0 or 7.0, also modify their activity. L.paucicostata, strain 151 is the most sensitive of the strainsused to both benzoic and salicylic acids followed by strain381. Such dramatic flowering responses were not obtained withthe other strains, but even strain 321, reportedly insensitiveto benzoic acid, could be induced to flower by adding benzoicacid to a modification of the medium. Benzoic acid is more effectivethan salicylic acid for all strains of L. paucicostata, butthe contrary is true for two L. minor strains tested. A higherpercentage of flowering is obtained in L. paucicostata in 1/2strength NH₄⁺-free Huter'sn medium than in M medium, exceptfor strain 151. When diluted, both media enhance flowering inall L. paucicostata strains. Generally, a lower concentrationof benzoic acid or salicylic acid is enough to induce floweringwhen the pH of the medium is lower. (Received March 30, 1981; Accepted May 16, 1981)     

The Role of Benzoic Acid and Plant Hormones in Flowering of Lemna gibba G3

The level of benzoic acid was measured in Lemna gibba G3 grownon M and E media under inductive and non-inductive daylengths.Benzoic acid was slightly higher in plants grown on M mediumbut there was no difference in the benzoic acid levels in floweringand vegetative plants. When L. gibba G3 was grown under continuouslight on 1/10 M medium or 1/2 H medium there was virtually noflowering, but addition of benzoic acid to either medium ledto a substantial flowering response. In both cases this floweringresponse was inhibited by the plant hormones IAA, GA₃, ABA andzeatin, with IAA and GA₃ being the least inhibitory and ABAbeing the most inhibitory. This same pattern of inhibition wasseen when L. gibba G3 was grown on M medium under continuouslight, conditions that lead to photoinduction of flowering.These results leave open the possibility that endogenous benzoicacid may interact with other factors to influence the floweringresponse in L. gibba G3. (Received November 13, 1984; Accepted February 27, 1985)     

Effects of Interactions between Low-temperature Treatments, Gibberellin (GA3) and Photoperiod on Flowering and Stem Height of Spring Rape (Brassica napus var. annua)

Exogenous gibberellin A₃(GA₃) reduced the number of leaf nodesat flowering and time to flowering and increased the stem heightat flowering in three genotypes of spring rape (Brassica napusvar.annua L.). The responses to GA₃were similar to those forlong days (LD) and low-temperature treatments, suggesting thatthe effect of photoperiod and the vernalization response areprobably mediated through gibberellins. The response to exogenousGA₃was greatest in non-cold-treated plants in short days (SD)suggesting that endogenous GAs are limiting in these conditions.CCC, an inhibitor of gibberellin biosynthesis, caused a smallincrease in the number of leaf nodes at flowering and time toflowering and a small decrease in the stem height at flowering,but unexpectedly, its effect was hardly influenced by the applicationof exogenous GA₃. Genotypes that showed the clearest responsesto the treatments with regard to the number of leaf nodes atflowering and time to flowering did not show the clearest responseswith regard to the stem height at flowering; the pattern ofresponses of the number of leaf nodes at flowering and timeto flowering was distinct from that of stem height at flowering.This indicates that flower formation and stem elongation areseparable developmental processes which may be controlled bydifferent endogenous gibberellins, different levels of a specificendogenous gibberellin, or different responses to gibberellin.Copyright 1999 Annals of Botany Company Brassica napus var. annua, gibberellin, photoperiod, spring rape, vernalization.     

Nitrate Uptake and Reduction of Aseptically Cultivated Spruce Seedlings, Picea abies (L.) Karst

Spruce (Picea abies (L.) Karst.) seedlings were asepticallycultivated and the effects of different N-nutrition on net uptakeand reduction of nitrate were investigated. The characteristicsof nitrate uptake were calculated, K_s as 0?2 mol m^–3 andV_max as 18 µmol g^–1 d^–1. Low pH, and Al³⁺ in the medium caused adecrease in nitrate uptake rate. An in vivo assay was set upwhich allowed the measurement of NRA in both roots and needlesof spruce seedlings. The in vivo nitrate reductase activitywas repressed by ammonium and stimulated by nitrate. Nitratereduction was similar to nitrate uptake, negatively affectedby low pH and ammonium. Therefore, a limited N-supply to spruceseemed to occur when pH was low in the rhizosphere combinedwith the presence of Al³⁺ and . Key words: Spruce, nitrate uptake, nitrate reduction     

Effect of Nitrogen Supply on the Grass and Clover Components of Simulated Mixed Swards Grown under Favourable Environmental Conditions II. Nitrogen Fixation and Nitrate Uptake

Simulated mixed swards of Perennial Ryegrass (Lolium perenneL.) cv. S23 and White clover (Trifolium repens L.) cv. S100were grown from seed under a constant 20 °C day/15 °Cnight temperature regime and harvested at intervals over and88 d growht period. The swards received a nutrient solutiondaily, which was either High (220 mg l^–1) or Low (10 mgl^–1) in nitrate N. The nitrate was labelled with the ¹⁵Nisotope. An acetylene reduction assay was carried out on eachsward just prior to harvest. Rates of acetylene reduction agreed qualitatively with the ^l5Nanalyses but absolute values did not match (assuming a 4:1 C₂H₄:N₂ratio) and errors in the acetylene assay are discussed. In theLow-N swards clover relied almost entirely on symbioticallyfixed N₂, fixing more than ten times as much as the High-N cloverplants. In the Low-N treatment the grass was N-deficient despiteobtaining much more nitrate per unit root dry weight than clover.In the High-N swards, however, clover took up more nitrate perunit root weight than grass. The High-N clover plants also fixedsome N₂ and maintained a higher total-N content than grass throughoutthe period. There was no evidence of transfer of symbioticallyfixed N from the clover to the grass in either treatment. Trifolium repens, Lolium perenne, nitrate, nitrogen fixation, ¹⁵N, acetylene reduction     

Changes in Crassulacean Acid Metabolism of Kalanchoe blossfeldiana by Different Nitrogen Sources

Kalanchoe blossfeldiana Poelln. cv. Hikan (a Crassulacean acidmetabolism (CAM) plant) was grown in pots containing soil for6 months and then cultured in nutrient solution containing 10mM nitrate or ammonium as a sole nitrogen source for 2 or 3months, under a long-day (16 h) condition. Plant growth was better in the nitrate medium. Leaves of thenitrate-grown plants showed greater diurnal fluctuations intitratable acidity and malate content than those of the ammonium-grownplants. The diurnal patterns in CO₂ exchange of nitrate-grownplants were basically similar for both groups, but the amountof net CO₂ uptake at night was twice as large in the nitrate-grownplants. The leaves of the nitrate-grown plants had 1.3 to 2.5times higher activities of phosphoenolpyruvate carboxylase (PEPC),phosphofructokinase (PFK) and NAD glycelaldehyde-3-phosphatedehydrogenase (G3PDH). These results indicate that K. blossfeldianagrown in nitrate medium showed more CAM activity than thosein ammonium medium. (Received August 13, 1987; Accepted February 22, 1988)     

Specific interactions in the physiology of flowering and gibbosity of Lemna gibba G3

Fronds of Lemna gibba G3 became conspicuously gibbous when ethrel,an ethylenereleasing compound, was added to the nutrient medium.Maximal gibbosity was obtained at ethrel concentrations of 1µg/ml and higher. Unlike the chelating agent, EDDHA, whichcauses profuse flowering and markedly gibbous fronds under long-dayconditions, ethrel did not affect flowering. In the presenceof an optimal concentration of EDDHA (10 µ/ml), ethreleven significantly inhibited flowering and caused developmentof excessively gibbous fronds. Autoclaved gibberellic acid specifically negated the ethreleffect as it does that of EDDHA. Three decomposition productsof GA₃, allogibberic acid, epiallogibberic acid and gibbericacid, also nullified flowering and gibbosity in the presenceof EDDHA. A fourth decomposition product of GA₃, epigibbericacid, inhibited gibbosity but hardly affected flowering. Salicylic acid was confirmed to affect flowering and gibbosityin L. gibba G3. However, contrary to an earlier report, it didnot induce flowering under short-day conditions. (Received January 10, 1976; )