Nitrate induction in spruce: an approach using compartmental analysis

Using ¹³NO ₃ ^– -efflux analysis, the induction of nitrate uptake by externally supplied nitrate was monitored in roots of intact Picea glauca (Moench) Voss. seedlings over a 5-d period. In agreement with our earlier studies, efflux analysis revealed three compartments, which have been identified as surface adsorption, apparent free space, and cytoplasm. While induction of nitrate uptake was pronounced, NO ₃ ^– fluxes in induced plants were decidedly lower and the induction response was slower than in other species. Influx rose from 0.1 mol·g^–1·h^–1 (measured at 100 M [NO ₃ ^– _o) in uninduced plants to a maximum of 0.5 mol·g^–1h^–1 after 3 d of exposure to 100 M [NO ₃ ^– _o and declined to 0.3–0.4 mol·g^–1h^–1 at the end of the 5-d period. Efflux remained relatively constant around 0.02-0.04 mol·g^–1h^–1, but its percentage with respect to influx declined from initially high values (around 30%) to steady-state values of 4–7%. Cytoplasmic [NO ₃ ^– ] ranged from the low micromolar in uninduced plants to a maximum of 2 mM in plants fully induced at 100 M [NO ₃ ^– ]_o. In-vivo root nitrate reductase activity (NRA) was measured over the same time period, and was found to follow a similar pattern of induction as influx. The maximum response in NRA slightly preceded that of influx. It increased from 25 nmol·g^–1·h^–1 without prior exposure to NO ₃ ^– to peak values around 150 nmol· g^–1h^–1 after 2 d of exposure to 100 M [NO ₃ ^– ]_o. Subsequently, NRA declined by about 50%. The dynamics of flux partitioning to reduction, to the vacuole, the xylem, and to efflux during the induction process are discussed.The research was supported by an Natural Sciences and Engineering Research Council, Canada, grant to Dr. A.D.M. Glass and by a University of British Columbia Graduate Fellowship to Herbert J. Kronzucker. Our thanks go to Dr. M. Adam and Mr. P. Culbert at the particle accelerator facility TRIUMF on the University of British Columbia campus for providing ¹³N, to Drs. R.D. Guy and S. Silim for providing plant material, and to Dr. M.Y. Wang, Mr. J. Bailey, Mr. J. Mehroke and Mr. J. Vidmar for essential assistance in experiments.     

Compartmentation and flux characteristics of ammonium in spruce

Using ¹³NH ₄ ⁺ as a tracer, compartmental analyses for NH ₄ ⁺ were performed in non-mycorrhizal roots of intact Picea glauca (Moench) Voss. seedlings at four different concentration regimes of external NH ₄ ⁺ ([NH ₄ ⁺ ]_o), i.e. 0, 10, 100, and 1500 M. Three kinetically distinct compartments were identified, with half-lives of exchange of approximately 2 s, 30 s, and 14 min, assumed to represent surface adsorption, Donnan free space, and cytoplasm, respectively. No significant differences were found in half-lives of exchange with changes in [NH ₄ ⁺ ]_o. Influx was calculated to be 0.96 mol·g^–1·h^–1 in N-deprived plants (measured at 10 M [NH ₄ ⁺ ]_o), while under steady-state conditions it was 0.21 mol·g^–1h^–1 at 10 M [NH ₄ ⁺ ]_o, 1.96 mol·g^–1h·^–1 at 100 M [NH ₄ ⁺ ]_o, and 6.45 mol·g^–1·h^–1 at 1.5 mM [NH ₄ ⁺ ]_o. Efflux measured over the same range constituted approximately 9% of influx in N-deprived plants, 10% at 10 M, 28% at 100 M, and 35% at 1.5 mM [NH ₄ ⁺ ]_o. Cytoplasmic [NH ₄ ⁺ ] was estimated at 6 m M in N-deprived plants, 2 mM at 10 M [NH ₄ ⁺ ]_o, 14 mM at 100 M, and 33 mM at 1.5 mM. Free-space [NH ₄ ⁺ ] was 84 M, 50 M, 700 M, and 8 mM, respectively. In comparison with previously published data on fluxes and compartmentation of NO ₃ ^– in white-spruce seedlings, results of this study identify a pronounced physiological preference of this species for NH ₄ ⁺ over NO ₃ ^– as an inorganic N source in terms of uptake and intracellular accumulation. The significant ecological importance of this N-source preference is discussed.The research was supported by a Natural Sciences and Engineering Research Council, Canada, grant to Dr. A.D.M. Glass and a University of British Columbia Graduate Fellowship to Herbert J. Kronzucker. Our thanks go to Dr. M. Adam and Mr. P. Culbert at the particle accelerator facility TRIUMF on the University of British Columbia campus for providing ¹³N, to Drs. R.D. Guy and S. Silim for providing plant material, and to Dr. M.Y. Wang, Mr. J. Bailey, Mr. J. Mehroke and Mr. P. Poon for essential assistance in experiments.     

Photoautotrophic micropropagation of Russet Burbank Potato

The photoautotrophic micropropagation of potato cv. Russet Burbank was investigated. Single node microcuttings were grown for four weeks on Murashige and Skoog (MS) medium with or without sucrose (30 g l^–1) in the growth room at 21/19 °C day/night temperature, with 16-h photoperiod at 150 mol m^–2 s^–1, with or without supplemental CO₂ at 1500 l l^–1. A 20% increase in the number of nodes per stem (from 7.5 to 9.4) and a 50% increase in stem dry weight were observed in cultures grown on media with sucrose and in CO₂ enriched atmosphere comparing to the conventionally micropropagated cultures or the cultures grown photoautotrophically on media without sucrose but in air supplemented with 1500 l l^–1CO₂. Stems of these cultures (from media with sucrose in CO₂ enriched air) almost doubled in length the stems of cultures from the other two treatments. No significant differences were observed between Control (MS medium supplemented with sucrose, 30 g l^–1) and photoautotrophic cultures coming from MS medium with no sucrose grown under 1500 l l^–1 of CO₂. Photoautotrophic cultures produced stems averaging 43.3 mm, with 7 nodes and weighing 9.2 mg (dry weight), similar to conventionally grown in vitro cultures (47.9 mm with 7.5 nodes, 9.7 mg dry weight). Growers may consider photoautotrophic culturing of potato in areas where the high sterility levels are difficult to maintain. Supplementing air in the growth room with 1500 l l^–1 of CO₂ could be beneficial for potato plantlet production even on media containing sucrose since it significantly improved quality, size and biomass of produced plantlets, speeding up the multiplication.     

Analysis of oxygen evolution during photosynthetic induction and in multiple-turnover flashes in sunflower leaves

Exchange of CO₂ and O₂ and chlorophyll fluorescence were measured in the presence of 360 1 · 1^–1 CO₂ in nitrogen in Helianthus annuss L. leaves which had been preconditioned in the dark or at a photon flux density (PFD) of 24 mol · m^–2 · s^–1 either in 21 or 0% O₂. An initial light-dependent O₂ outburst of 6 mol · m^–2 was measured after aerobic dark incubation. It was attributed to the reduction of electron carriers, predominantly plastoquinone. The maximum initial rate of O₂ evolution at PFD 8000 mol · m^–2 · s^–1 was 170 mol · m^–2 · s^–2 or about four times the steady CO₂-and light-saturated rate of photosynthesis. Fluorescence measurements showed that the rate was still acceptor-limited. Fast O₂ evolution ceased after electron carriers were reduced in the dark-adapted leaf, but continued for a short time at the lower rate of 62 mol · m^–2 · s^–1 in the light-adapted leaf. The data are interpreted to show that enzymes involved in 3-phosphoglycerate reduction are dark-inhibited, but were fully active in low light. In a dark-adapted leaf, respiratory CO₂ evolution continued under nitrogen; it was partially inhibited by illumination. Prolonged exposure of a leaf to anaerobic conditions caused reducing equivalents to accumulate. This was shown by a slowly increasing chlorophyll fluorescence yield which indicated the reduction of the PSII acceptor Q_A in the dark. When the leaf was illuminated, no O₂ evolution was detected from short light pulses, although transient O₂ production was appreciable during longer light pulses. This indicates that an electron donor (pool size about 2–3 e/PSII reaction center) became reduced in the dark and the first photons were used to oxidise this donor instead of water.Abbreviations Chl chlorophyll - CRC carbon reduction cycle - GAPDH NADP-glyceraldehyde-phosphate dehydrogenase - PFD photon flux density - PGA 3-phosphoglycerate - RuBP ribulose bisphosphate - TCA tricarboxylic acid cycle To whom correspondence should be addressedThis work received support by the Estonian Academy of Sciences, the Gottfried-Wilhelm-Leibniz Program of the Deutsche For-schungsgemeinschaft and the Sonderforschungsbereich 251 of the University of Würzburg.     

Some effects of long-term ozone fumigations on Norway spruce

Summary Potted cuttings of a 12-year-old, and grafts of an 80-year-old, Norway spruce (Picea abies (L.) Karst.) were subjected to 100 or 300 g O₃·m^–3 for 1215 h (45 h of daylight per week) during the growing season of 1985. At the end of the fumigation the plants did not exhibit any visible signs of injury. Whereas in the fumigation with 100g O₃·m^–3 we did not detect any significant change in gas exchange, 300 g O₃·m^–3 did alter the CO₂ uptake after 27 weeks, and in one clone transpiration was also altered. Stomatal reaction to a change of light suggested sluggishness, but the change was not statistically significant.     

Growth,photosynthesis and photorespiration of Lemna gibba: response to variations in CO2 and O2 concentrations and photon flux density

Dry weight and Relative Growth Rate of Lemna gibba were significantly increased by CO₂ enrichment up to 6000 l CO₂ l^–1. This high CO₂ optimum for growth is probably due to the presence of nonfunctional stomata. The response to high CO₂ was less or absent following four days growth in 2% O₂. The Leaf Area Ratio decreased in response to CO₂ enrichment as a result of an increase in dry weight per frond. Photosynthetic rate was increased by CO₂ enrichment up to 1500 l CO₂ l^–1 during measurement, showing only small increases with further CO₂ enrichment up to 5000 l CO₂ l^–1 at a photon flux density of 210 mol m^–2 s^–1 and small decreases at 2000 mol m^–1 s^–1. The actual rate of photosynthesis of those plants cultivated at high CO₂ levels, however, was less than the air grown plants. The response of photosynthesis to O₂ indicated that the enhancement of growth and photosynthesis by CO₂ enrichment was a result of decreased photorespiration. Plants cultivated in low O₂ produced abnormal morphological features and after a short time showed a reduction in growth.     

Product inhibition during the feeding phasein fed-batch ethanol fermentation of sugar-cane blackstrap molasses

Summary The following equations represent the influence of the ethanol concentration (E) on the specific growth rate of the yeast cells () and on the specific production rate of ethanol () during the reactor filling phase in fed-batch fermentation of sugar-cane blackstrap molasses: = ₀ - k · E and v = v ₀ · K/(K +E) Nomenclature E ethanol concentration in the aqueous phase of the fermenting medium (g.L^–1) - E_m value of E when = 0 or = 0 (g.L^–1) - F medium feeding rate (L.h^–1) - k empirical constant (L.g^–1.h^–1) - K empirical constant (g.L^–1) - M_as mass of TRS added to the, reactor (g) - M_cs mass of consumed TRS (g) - M_e mass of ethanol in the aqueous phase of the fermenting medium (g) - M_s mass of TRS in the aqueous phase of the fermenting medium (g) - M_x mass of yeast cells (dry matter) in the fermenting medium (g) - r correlation coefficient - S TRS concentration in the aqueous phase of the fermenting medium (g.L^–1) - S_m TRS concentration of the feeding medium (g.L^–1) - t time (h) - T temperature (° C) - TRS total reducing sugars calculated as glucose - V volume of the fermenting medium (L) - V₀ volume of the inoculum (L) - X yeast cells concentration (dry matter) in the fermenting medium (g.L^–1) - filling-up time (h) - specific growth rate of the yeast cells (h^–1) - ₀ value of when E=0 - specific production rate of ethanol (h^–1) - ₀ value of when E=0 - density of the yeast cells (g.L^–1) - dry matter content of the yeast cells     

Effects of constitutive expression of nitrate reductase in transgenic Nicotiana plumbaginifolia L. in response to varying nitrogen supply

Transformed Nicotiana plumbaginifolia plants with constitutive expression of nitrate reductase (NR) activity were grown at different levels of nitrogen nutrition. The gradients in foliar NO ₃ ^– content and maximum extractable NR activity observed with leaf order on the shoot, from base to apex, were much decreased as a result of N-deficiency in both the transformed plants and wild type controls grown under identical conditions. Constitutive expression of NR did not influence the foliar protein and chlorophyll contents under any circumstances. A reciprocal relationship between the observed maximal extractable NR activity of the leaves and their NO ₃ ^– content was observed in plants grown in nitrogen replete conditions at low irradiance (170 mol photons·m^–2 ·s^–1). This relationship disappeared at higher irradiance (450 mol photons·m^–2·S^–1) because the maximal extractable NR activity in the leaves of the wild type plants in these conditions increased to a level that was similar to, or greater than that found in constitutive NR-expressors. Much more NO ₃ ^– accumulated in the leaves of plants grown at 450 mol photons·m^–2·s^–1 than in those grown at 170 mol photons·m^–2·s^–1 in N-replete conditions. The foliar NO ₃ ^– level and maximal NR activity decreased with the imposition of N-deficiency in all plant types such that after prolonged exposure to nitrogen depletion very little NO ₃ ^– was found in the leaves and NR activity had decreased to almost zero. The activity of NR decreased under conditions of nitrogen deficiency. This regulation is multifactoral since there is no regulation of NR gene expression by NO ₃ ^– in the constitutive NR-expressors. We conclude that the NR protein is specifically targetted for destruction under nitrogen deficiency. Consequently, constitutive expression of NR activity does not benefit the plant in terms of increased biomass production in conditions of limiting nitrogen.Abbreviations Chl chlorophyll - N nitrogen - NR NADH-nitrate reductase - WT wild type     

Glycolate photoproduction by free and alginate-entrapped cells of Chlamydomonas reinhardtii

Summary Chlamydomonas reinhardtii cells provide an effective system for glycolate photoproduction, operative during 30 h when they are growing under low CO₂, in the presence of 1 mM aminooxyacetate and 50 M acetazolamide. Glycolate excretion by the cells can proceed for about 4 days if every other 12 h a high CO₂ level is restored in the culture in the absence of inhibitors. The immobilized system in alginate beads has about a twofold higher glycolate photoproduction rate (23 mol·mg chlorophyll (Chl)^–1·h^–1) than free-living cells (12 mol · mg Chl^–1 · h^–1). Offprint requests to: C. Vílchez     

Elevated CO2 and temperature effects on soil carbon and nitrogen cycling in ryegrass/white clover turves of an Endoaquept soil

Effects of elevated CO₂ (700 L L^–1) and a control (350 L L^–1 CO₂) on the productivity of a 3-year-old ryegrass/white clover pasture, and on soil biochemical properties, were investigated with turves of a Typic Endoaquept soil in growth chambers. Temperature treatments corresponding to average winter, spring, and summer conditions in the field were applied consecutively to all of the turves. An additional treatment, at 700 L L^–1 CO₂ and a temperature 6°C higher throughout than in the other treatments, was included.Under the same temperature conditions, overall herbage yields in the 700 L L^–1 CO₂ treatment were ca. 7% greater than in the control at the end of the summer period. Root mass (to ca 25 cm depth) in the 700 L L^–1 CO₂ treatment was then about 50% greater than in the control, but in the 700 L L^–1 CO₂+6°C treatment it was 6% lower than in the control. Based on decomposition results, herbage from the 700 L L^–1+6°C treatment probably contained the highest proportion of readily decomposable components.Elevated CO₂ had no consistent effect on soil total C and N, microbial C and N, or extractable C concentrations in any of the treatments. Under the same temperature conditions, it did, however, enhance soil respiration (CO₂-C production) and invertase activity. The effects of elevated CO₂ on rates of net N mineralization were less distinct, and the apparent availability of N for the sward was not affected. Under elevated CO₂, soil in the higher-temperature treatment had a higher microbial C:N ratio; it also had a greater potential to degrade plant materials.Data interpretation was complicated by soil spatial variability and the moderately high background levels of organic matter and biochemical properties that are typical of New Zealand pasture soils. More rapid cycling of C under CO₂ enrichment is, nevertheless, indicated. Futher long-term experiments are required to determine the overall effect of elevated CO₂ on the soil C balance.     

Photosynthesis and apparent affinity for dissolved inorganic carbon by cells and chloroplasts of Chlamydomonas reinhardtii grown at high and low CO2 concentrations

Chloroplasts with high rates of photosynthetic O₂ evolution (up to 120 mol O₂· (mg Chl)^-1·h^-1 compared with 130 mol O₂· (mg Chl)^-1·h^-1 of whole cells) were isolated from Chlamydomonas reinhardtii cells grown in high and low CO₂ concentrations using autolysine-digitonin treatment. At 25° C and pH=7.8, no O₂ uptake could be observed in the dark by high- and low-CO₂ adapted chloroplasts. Light saturation of photosynthetic net oxygen evolution was reached at 800 mol photons·m^-2·s^-1 for high- and low-CO₂ adapted chloroplasts, a value which was almost identical to that observed for whole cells. Dissolved inorganic carbon (DIC) saturation of photosynthesis was reached between 200–300 M for low-CO₂ adapted chloroplasts, whereas high-CO₂ adapted chloroplasts were not saturated even at 700 M DIC. The concentrations of DIC required to reach half-saturated rates of net O₂ evolution (K_m(DIC)) was 31.1 and 156 M DIC for low- and high-CO₂ adapted chloroplasts, respectively. These results demonstrate that the CO₂ concentration provided during growth influenced the photosynthetic characteristics at the whole cell as well as at the chloroplast level.Abbreviations Chl chlorophyll - DIC dissolved inorganic carbon - K_m(DIC) coneentration of dissolved inorganic carbon required for the rate of half maximal net O₂ evolution - PFR photon fluence rate - SPGM silicasol-PVP-gradient medium     

Effects of <Emphasis Type="Italic">in vitro</Emphasis>rooting environments and irradiance on growth and photosynthesis of strawberry plantlets during acclimatization

Strawberry (Fragaria ananassaDuch. cv. Fengxiang) plantlets were cultured under two in vitroenvironments for rooting, and then acclimatized under two ex vitroirradiance conditions. At the end of rooting stage plant height, fresh weight and specific leaf area of T1-plants grown under high sucrose concentration (3 sucrose), low photosynthetic photon flux density (30 mol m^–2 s^–1) and normal CO₂ concentration (350–400 l l^–1) were significantly higher than those of T2-plantlets grown under low sucrose concentration (0.5), high photosynthetic photon flux density (90 mol m^–2 s^–1) and elevated CO₂ concentration (700–800 l l^–1). But T2-plantlets had higher net photosynthetic rate (Pn), effective photochemical quantum yield of PSII (_PSII), effective photosynthetic electron transport rate (ETR), photochemical quenching (q_P) and ratio of chlorophyll fluorescence yield decrease (Rfd). After transfer, higher irradiance obviously promoted the growth of plantlets and was beneficial for the development of photosynthetic functions during acclimatization. T2-plantlets had higher fresh weight, leaf area, _PSII and ETR under higher ex vitroirradiance condition.     

The effect of carbon dioxide on the growth kinetics of fructose-limited chemostat cultures of Acetobacterium woodii DSM 1030

The growth of the anaerobic acetogenic bacterium Acetobacterium woodii DSM 1030 was investigated in fructose-limited chemostat cultures. A defined medium was developed which contained fructose, mineral salts, cysteine · HCl and Ca pantothenate (1 mg · 1^–1) supplied in a vitamin supplement. Growth at high dilution rates was dependent on the presence of CO₂ in the gas phase. The ^max was found to be 0.16 h^–1 and the fructose maintenance requirement was 0.1 to 0.13 mmol fructose · (g dry wt)^–1 · h^–1. A growth yield of 61 g dry wt · (mol fructose)^–1, corrected for the cell maintenance requirement and for incorporation of fructose carbon into cell biomass, was determined from the fructose consumption. A corresponding growth yield of 69 g dry wt · (mol fructose)^–1 was calculated from the acetate production assuming that fructose fermentation was homoacetogenic. A Y_ATP of 12.2 to 13.8 g dry wt · (mol ATP)^–1 was calculated from these growth yields using a value of 5 mol ATP · (mol fructose)^–1 as an estimate of the amount of ATP synthesised from fructose fermentation. The addition of yeast extract (0.5 g · 1^–1) to the medium did not influence the ^max or cell yield. After prolonged growth under fructose-limited conditions the requirement of the culture for CO₂ in the gas phase was reduced.Abbreviations YE yeast extract - IC inorganic carbon - D fermenter dilution rate : h^–1 - M_X maintenance requirement for X: mmol X · (g dry wt)^–1 · h^–1 - X may be fructose (Fruct), fructose consumed in energy metabolism (Fruct [E]), acetate (Ac) - ATP CO₂, NH _inf4 ^sup+ or Pi - qX specific rate of utilisation or consumption of X: mmol X · (g dry wt)^–1 · h^–1 - V fermenter volume: litre - rC · Cell, fermenter cell carbon production: mmol C · h^–1 - Y_X yield of cells on X: g dry wt · (mol X)^–1 - Y _infx ^supmax the yield corrected for cell maintenance: g dry wt · (mol X)^–1 - S_ATP stoichiometry of ATP synthesis from fructose: mol ATP · (mol frucose)^–1 - x cell concentration: g dry wt · 1^–1 - specific growth rate : h^–1 - ^max maximum specific growth rate: h^–1     

Procedures for the axenic isolation of conchocelis and monospores from the red seaweed Porphyra yezoensis

In order to maintain axenic seedstock cultures axenically of thecommercially important red seaweed, Porphyra yezoensis, aprocedure was developed for axenic isolation and culture of conchocelis andmonospores. For axenic isolation of the conchocelis, contaminated microalgaewere most effectively removed by filtering contaminated samples through a100-m mesh after sonication. Removal of bacteria and otheralgaewas accomplished using a mixture of 5 agents (0.02% chitosan, 100 gml^–1 GeO₂, 10 gml^–1 ampicillin, 40 gml^–1 kanamycin and 200 gml^–1 streptomycin). Axenic single colonies wereisolatedfrom a semi-solid medium prepared from 1% transfer gel. After collectingmonospores from the 40–50% density layer on a percoll-gradient, removalofbacteria and fungi from the monospores was accomplished using a mixture of 5antibiotics (3.5 g ml^–1 nystatin, 2 mgml^–1 ampicillin, 400 gml^–1 kanamycin, 50 gml^–1 neomycin and 800 gml^–1 streptomycin). Axenic single juvenile blades wereisolated from a semi-solid medium prepared from 0.5% transfer gel.     

Transient state characteristics of adaptation to changes in light conditions for the cyanobacterium Oscillatoria agardhii

Transitions in growth irradiance level from 92 to 7 Em^-2 s^-1 and vice versa caused changes in the pigment contents and photosynthesis of Oscillatoria agardhii. The changes in chlorophyll a and C-phycocyanin contents during the transition from high to low irradiance (HL) were reflected in photosynthetic parameters. In the LH transition light utilization efficiencies of the cells changed faster than pigment contents. This appeared to be related to the lowering of light utilization efficiencies of photosynthesis. As a possible explanation it was hypothesized that excess photosynthate production led to feed back inhibition of photosynthesis. Time-scales of changes in the maximal rate of O₂ evolution were discussed as changes in the number of reaction centers of photosystem II in relation to photosynthetic electron transport. Parameters that were subject to change during irradiance transitions obeyed first order kinetics, but hysteresis occurred when comparing HL with LH transients. Interpretation of first order kinetic analysis was discussed in terms of adaptive response vs changes in growth rate.Non-standard abbreviations Chla chlorophyll a - CPC C-phycocyanin - PS II photosystem II - PS I photosystem I - RC II reaction center of photosystem II - P photosynthetic O₂-evolution - I irradiance, Em^-2 s^-1 - light utilization efficiency of cells, mmol O₂·mg dry wt^-1·h^-1/Em^-2 s^-1 - light utilization efficiency of photosynthetic apparatus, mol O₂·mol Chla ^-1·h^-1/Em^-2 s^-1 - P_max maximal rate of O₂ evolution by cells, mol O₂·mg dry wt^-1·h^-1 - P_max maximal rate of O₂ evolution by photosynthetic apparatus, mol O₂·mol·Chla ^-1·h^-1 - LL low light, E m^-2 s^-1 - HL high light, E m^-2 s^-1 - LH low to high light transition - HL high to low light transition - k specific rate of adaptation, h^-1 - specific growth rate, h^-1 - Q pool size of cell constituent, mol·mg dry wt^-1 - q net synthesis rate of cell constituent, mol·mg dry wt^-1·h^-1     

Compartmentation and flux characteristics of nitrate in spruce

The radiotracer¹³N was used to undertake compartmental analyses for NO ₃ ^– in intact non-mycorrhizal roots ofPicea glauca (Moench) Voss. seedlings. Three compartments were defined, with half-lives of exchange of 2.5 s, 20 s, and 7 min. These were identified as representing surface adsorption, apparent free space, and cytoplasm, respectively. Influx, efflux, and net flux as well as cytoplasmic and apparent-free-space nitrate concentrations were estimated for three different concentration regimes of external nitrate. After exposure to external NO ₃ ^– for 3 d, influx was calculated to be 0.09 mol·g^–1·h^–1 (at 10 M [NO ₃ ^– ]_o), 0.5mol·g^–1·h^–1 (at 100 M [NO _inf3 ^sup– ]_o), and 1.2 mol · g^–1· h^–1 (at 1.5 mM [NO ₃ ^– ]_o). Efflux increased with increasing [NO ₃ ^– ]_o, constituting 4% of influx at 10 M, 6% at 100 M, and 21% at 1.5 mM. Cytoplasmic [NO ₃ ^– ] was estimated to be 0.3 mM at 10 uM [NO ₃ ^– ]_o, 2mM at 100 M [NO ₃ ^– ]_o, and 4mM at 1.5 mM [NO ₃ ^– ]_o, while free-space [NO ₃ ^– ] was 16 M, 173 M, and 2.2 mM, respectively. A series of experiments was carried out to confirm the identity of the compartments resolved by efflux analysis. Pretreatment at high temperature or application of 2-chloro-ethanol, sodium dodecyl sulphate or hydrogen peroxide made it possible to distinguish the metabolic (cytoplasmic) phase from the remaining two (physical) phases. Likewise, varying [Pi] of the medium altered efflux and thereby [NO ₃ ^– ]_cyt, but did not affect [NO ₃ ^– ]_{free space}.Abbreviations and Symbols [NO ₃ ^– ]_cyt cytoplasmic NO ₃ ^– concentration - [NO ₃ ^– ]_{free space} apparent-free-space NO ₃ ^– concentration - [NO ₃ ^– ]_o concentration of NO ₃ ^– in the external solution - NO ₃ ^– flux - _co efflux from the cytoplasm - _oc influx to the cytoplasm - _net net flux - _xylem flux to the xylem - _red/vac combined flux to reduction and the vacuole The research was supported by a Natural Sciences and Engineering Research Council, Canada, grant to Dr. A.D.M. Glass and by a University of British Columbia Graduate Fellowship to Herbert J. Kronzucker. Our thanks go to Dr. M. Adam and Mr. P. Culbert at the particle accelerator facility TRIUMF on the University of British Columbia Campus for providing¹³NO ₃ ^– , Drs. R.D. Guy and S. Silim for providing plant material, and Dr. M.Y. Wang, Mr. J. Mehroke and Mr. P. Poon for assistance in experiments and for helpful discussions.     

Light response of D1 turnover and photosystem II efficiency in the seagrassZostera capricorni

Biochemical and biophysical parameters, including D1-protein turnover, chlorophyll fluorescence, oxygen evolution activity and zeaxanthin formation were measured in the marine seagrassZostera capricorni (Aschers) in response to limiting (100 mol·m^–2·^–1), saturating (350 mol·m^–2·s^–1) or photoinhibitory (1100 mol·m^–2·s^–1) irradiances. Synthesis of D1 was maximal at 350 mol·m^–2·s^–1 which was also the irradiance at which the rate of photosynthetic O₂ evolution was maximal. Degradation of D1 was saturated at 350 mol·m^–2·s^–1. The rate of D1 synthesis at 1100 mol·m^–2·s^–1 was very similar to that at 350 mol·m^–2·s^–1 for the first 90 min but then declined. At limiting or saturating irradiance little change was observed in the ratio of variable to maximal fluorescence (Fv/Fm) measured after dark adaptation of the leaves, while significant photoinhibition occurred at 1100 mol·m^–2·s^–1. The proportion of zeaxanthin in the total xanthophyll pool increased with increasing irradiance, indicative of the presence of a photoprotective xanthophyll cycle in this seagrass. These results are consistent with a high level of regulatory D1 turnover inZostera under non-photoinhibitory irradiance conditions, as has been found previously for terrestrial plants.We would like to thank Professor Peter Böger (Department of Plant Biochemistry, University of Konstanz, Germany) for the kind gift of D1 antibodies. This work was partly supported by a University of Queensland Enabling Grant to CC.     

Decreased ribulose-1,5-bisphosphate carboxylase-oxygenase in transgenic tobacco transformed with “antisense” rbcS

Tobacco (Nicotiana tabacum L.) plants transformed with antisense rbcS to decrease the expression of ribulose-1,5-bisphosphate carboxylase-oxygenase (Rubisco) have been used to investigate the contribution of Rubisco to the control of photosynthesis in plants growing at different irradiances. Tobacco plants were grown in controlled-climate chambers under ambient CO₂ at 20°C at 100, 300 and 750 mol·m^–2·s^–1 irradiance, and at 28°C at 100, 300 and 1000 mol·m^–2·s^–1 irradiance. (i) Measurement of photosynthesis under ambient conditions showed that the flux control coefficient of Rubisco (C _infRubisco ^supA ) was very low (0.01–0.03) at low growth irradiance, and still fairly low (0.24–0.27) at higher irradiance. (ii) Short-term changes in the irradiance used to measure photosynthesis showed that C _infRubisco ^supA increases as incident irradiance rises, (iii) When low-light (100 mol·m^–2·s^–1)-grown plants are exposed to high (750–1000 mol·m^–2·s^–1) irradiance, Rubisco is almost totally limiting for photosynthesis in wild types. However, when high-light-grown leaves (750–1000 mol·m^–2·s^–1) are suddenly exposed to high and saturating irradiance (1500–2000 mol·m^–2·s^–1), C _infRubisco ^supA remained relatively low (0.23–0.33), showing that in saturating light Rubisco only exerts partial control over the light-saturated rate of photosynthesis in sun leaves; apparently additional factors are co-limiting photosynthetic performance, (iv) Growth of plants at high irradiance led to a small decrease in the percentage of total protein found in the insoluble (thylakoid fraction), and a decrease of chlorophyll, relative to protein or structural leaf dry weight. As a consequence of this change, high-irradiance-grown leaves illuminated at growth irradiance avoided an inbalance between the light reactions and Rubisco; this was shown by the low value of C _infRubisco ^supA (see above) and by measurements showing that non-photochemical quenching was low, photochemical quenching high, and NADP-malate dehydrogenase activation was low at the growth irradiance. In contrast, when a leaf adapted to low irradiance was illuminated at a higher irradiance, Rubisco exerted more control, non-photochemical quenching was higher, photochemical quenching was lower, and NADP-malate dehydrogenase activation was higher than in a leaf which had grown at that irradiance. We conclude that changes in leaf composition allow the leaf to avoid a one-sided limitation by Rubisco and, hence, overexcitation and overreduction of the thylakoids in high-irradiance growth conditions, (v) Antisense plants with less Rubisco contained a higher content of insoluble (thylakoid) protein and chlorophyll, compared to total protein or structural leaf dry weight. They also showed a higher rate of photosynthesis than the wild type, when measured at an irradiance below that at which the plant had grown. We propose that N-allocation in low light is not optimal in tobacco and that genetic manipulation to decrease Rubisco may, in some circumstances, increase photosynthetic performance in low light.Abbreviations A rate of photosynthesis - C _infRubisco ^supA flux control coefficient of Rubisco for photosynthesis - c_i internal CO₂ concentration - qE energy-dependent quenching of chlorophyll fluorescense - qQ photochemical quenching of chlorophyll fluorescence - NADP-MDH NADP-dependent malate dehydrogenase - Rubisco ribulose-1,5-bisphosphate carboxylase-oxygenase - RuBP ribulose-1,5-bisphosphate This work was supported by the Deutsche Forschungsgemeinschaft (SFB 137).