Phosphate Deficiency in Maize. I. Leaf Phosphate Status, Growth, Photosynthesis and Carbon Partitioning

Maize plants (Zea mays L.) were cultured with nutrient solutioncontaining 0.001 or 0.5 mM orthophosphate (Pi). Effects of lowphosphate (low-P) nutrition on growth, leaf phosphate status,photosynthesis, and carbon partitioning were investigated. Withlow-P treatment, the fresh weight of aerial parts decreasedby about 40% by 24 days after planting. Detailed studies ofthe effects of low-P treatment on the other characteristicsof maize leaves-were done using the middle part of the thirdleaf, counting from the base. Low-P treatment had almost noeffect on specific leaf weight or soluble protein content measured13 to 21 days after planting. Low-P treatment decreased Chicontent slightly (by 15% 19 days after planting). Twenty onedays after planting, low-P treatment had greatly decreased thelevels of leaf acid extractable Pi (by 77%) and photosynthesisrates (by 68%). The detrimental effects of low-P treatment onthe rates of photosynthesis and the amounts of acid extractablePi became progressively greater with time. There was a strongcorrelation between levels of leaf acid extractable Pi and ratesof photosynthesis. The minimum level of Pi necessary to sustainthe maximum photosynthesis rate was 0.6 mmol m^–2. Belowthis minimum content of Pi, the rate of photosynthesis decreasedsharply with decreasing Pi. To investigate the direct effectof Pi depletion on photosynthate partitioning at equivalentrates of photosynthesis, the rates in controls were reducedto almost the same as those in 18 or 19 day old low-P plants(about 50% of those in controls) by lowering light intensityand/ or ambient CO₂ concentration. The data clearly indicatesthat low-P treatment had a direct effect in lowering photosynthatepartitioning into starch. Starch mobilization during the nightwas also inhibited under low-P conditions. (Received January 7, 1991; Accepted March 5, 1991)     

A possible role for C4 photosynthetic enzymes in tolerance of Zea mays to NaCl

Treatment of 14-day-old maize cultivars (Hybrid351 and Giza2) with 250 mM NaCl significantly reduced shoot fresh and dry weights and protein content during the subsequent 12 days. The magnitude of reduction was more pronounced in Giza than Hybrid. Both cultivars contained converging levels of protein for the enzymes phosphoenolpyruvate carboxylase (PEPC), malate dehydrogenase (MDH), pyruvate phosphate dikinase (PPDK) and ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco) under normal conditions; however, NaCl led to increase these levels in Hybrid and decrease them in Giza. Moreover, NaCl significantly inhibited the activities of PEPC, MDH and PPDK in both cultivars during the first 2 days, thereafter the inhibition nullified only in Hybrid; nonetheless, Rubisco was the least affected enzyme in both cultivars. In addition, NaCl slightly increased V _max of PEPC, MDH and PPDK in Hybrid with no change in K _m; nevertheless V _max dropped in Giza with an increase in K _m of only PEPC and MDH. Also K _cat, K _cat/K _m and V _max/K _m of all enzymes were lower in treated Giza than in treated Hybrid. The increased V _max of all enzymes in only Hybrid by NaCl confirms that they were synthesised more in Hybrid than in Giza. However, the decreased V _max in Giza concomitant with the increased K _m points to an interference of salinity with synthesis of enzymes and their structural integrity. This would lead to a noncompetitive inhibition for the enzymes. These findings declare that maize tolerance to NaCl was larger in Hybrid compared to Giza due to a role for C4 enzymes.     

Kinetic Properties of Phosphoenolpyruvate Carboxylase in Peperomia camptotricha

Kinetic characteristics of phosphoenolpyruvate carboxylase (PEPC) from the epiphytic C₃ or C₄: CAM intermediate plant, Peperomia camptotricha, were investigated. Few day versus night differences in V_max,K_m(PEP)), or malate inhibition were observed, even in extracts from water-stressed plants which characteristically perform CAM, regardless of efforts to stabilize day/night forms. The PEPC extracted from plants during the light period remained stable, without much of an increase or decrease in activity for at least 22 hours at 0 to 4°C. Extracts from mature, fully developed leaves had slightly greater PEPC activity than from very young, developing leaves. Generally, however, the kinetic properties of PEPC extracted from mature leaves of plants grown under short day (SD), long day (LD), or 1-week water-stress conditions, as well as from young, developing leaves, were similar. The PEPC inhibitor, l-malate, decreased the V_max and increased the K_m(PEP) for all treatments. Under specific conditions, malate did not inhibit PEPC rates in the dark extracts as much as the light. The PEPC activator, glucose-6-phosphate (G-6-P), lowered the K_m(PEP) for all treatments. At saturating PEP concentrations, PEPC activity was independent of pH in the range of 7.5 to 9.0. At subsaturating PEP concentrations, the pH optimum was 7.8. The rates of PEPC activity were lower in the light period extracts than the dark, at pH 7.1, but day/night PEPC was equally active at pH 7.8. At pH 7.5 and a subsaturating PEP concentration, G-6-P significantly activated PEPC. At pH 8, however, only slight activation by G-6-P was observed. The lower pH of 7.5 combined with l-malate addition, greatly inhibited PEPC, particularly in extracts from young, developing leaves which were completely inhibited at an l-malate concentration of 1 millimolar. However, malate did not further inhibit PEPC activity in mature leaves when assayed at pH 7.1. The fairly constant day/night kinetic and regulatory properties of PEPC from P. camptotricha are unlike those of PEPC from CAM or C₄ species studied, and are consistent with the photosynthetic metabolism of this plant.     

The use and misuse of V c,max in Earth System Models

Earth System Models (ESMs) aim to project global change. Central to this aim is the need to accurately model global carbon fluxes. Photosynthetic carbon dioxide assimilation by the terrestrial biosphere is the largest of these fluxes, and in many ESMs is represented by the Farquhar, von Caemmerer and Berry (FvCB) model of photosynthesis. The maximum rate of carboxylation by the enzyme Rubisco, commonly termed V _c,max, is a key parameter in the FvCB model. This study investigated the derivation of the values of V _c,max used to represent different plant functional types (PFTs) in ESMs. Four methods for estimating V _c,max were identified; (1) an empirical or (2) mechanistic relationship was used to relate V _c,max to leaf N content, (3) V _c,max was estimated using an approach based on the optimization of photosynthesis and respiration or (4) calibration of a user-defined V _c,max to obtain a target model output. Despite representing the same PFTs, the land model components of ESMs were parameterized with a wide range of values for V _c,max (?46 to +77 % of the PFT mean). In many cases, parameterization was based on limited data sets and poorly defined coefficients that were used to adjust model parameters and set PFT-specific values for V _c,max. Examination of the models that linked leaf N mechanistically to V _c,max identified potential changes to fixed parameters that collectively would decrease V _c,max by 31 % in C₃ plants and 11 % in C₄ plants. Plant trait data bases are now available that offer an excellent opportunity for models to update PFT-specific parameters used to estimate V _c,max. However, data for parameterizing some PFTs, particularly those in the Tropics and the Arctic are either highly variable or largely absent.     

Developmental regulation of photosynthate distribution in leaves of rice

mRNA expression patterns of genes for metabolic key enzymes sucrose phosphate synthase (SPS), phosphoenolpyruvate carboxylase (PEPC), pyruvate kinase, ribulose 1,5-bisphosphate carboxylase/oxygenase, glutamine synthetase 1, and glutamine synthetase 2 were investigated in leaves of rice plants grown at two nitrogen (N) supplies (N_0.5, N_3.0). The relative gene expression patterns were similar in all leaves except for 9^th leaf, in which mRNA levels were generally depressed. Though increased N supply prolonged the expression period of each mRNA, it did not affect the relative expression intensity of any mRNA in a given leaf. SPS V_max, SPS limiting and PEPC activities, and carbon flow were examined. The ratio between PEPC activity and SPS V_max was higher in leaves developed at the vegetative growth stage (vegetative leaves: 5^th and 7^th leaves) than in leaves developed after the ear primordia formation stage (reproductive leaves: 9^th and flag leaves). PEPC activity and SPS V_max decreased with declining leaf N content. After using ¹⁴CO₂ the ¹⁴C photosynthate distribution in the amino acid fraction was higher in vegetative than in reproductive leaves when compared for the same leaf N status. Thus, at high PEPC/SPS activities ratio, more ¹⁴C photosynthate was distributed to the amino acid pool, whereas at higher SPS activity more ¹⁴C was channelled into the saccharide fraction. Thus, leaf ontogeny was an important factor controlling photosynthate distribution to the N- or C-pool, respectively, regardless of the leaf N status.     

Influence of Phosphorus Nutrition on Growth and Carbon Partitioning in Glycine max

Soybean plants (Glycine max [L.] Merr var Amsoy 71) were grown in growth chambers with high-phosphorus (high-P) and low-phosphorus (low-P) culture solutions. Low-P treatment reduced shoot growth significantly 7 days after treatment began. Root growth was much less affected by low-P, there being no significant reduction in root growth rate until 17 days had elapsed. The results suggest that low-P treatment decreased soybean growth primarily through an effect on the expansion of the leaf surface which was diminished by 85%, the main effect of low-P being on the rate of expansion of individual leaves. Low-P had a lesser effect on photosynthesis; light saturated photosynthetic rates at ambient and saturating CO₂ levels were lowered by 55 and 45%, respectively, after 19 days of low-P treatment. Low-P treatment increased starch concentrations in mature leaves, expanding leaves and fibrous roots; sucrose concentrations, however, were reduced by low-P in leaves and increased in roots. Foliar F-2,6-BP levels were not affected by P treatment in the light but in darkness they increased with high-P and decreased with low-P. The increase in the starch/sucrose ratio in low-P leaves was correlated primarily with changes in the total activities of enzymes of starch and sucrose metabolism.     

Nitrogenase activity, nodule respiration, and o(2) permeability following detopping of alfalfa and birdsfoot trefoil

Gas exchange measurements and noninvasive leghemoglobin (Lb) spectrophotometry (nodule oximetry) were used to monitor nodule responses to shoot removal in alfalfa (Medicago sativa L. cv Weevlchek) and birdsfoot trefoil (Lotus corniculatus L. cv Fergus). In each species, total nitrogenase activity, measured as H₂ evolution in Ar:O₂ (80:20), decreased to <50% of the initial rate within 1 hour after detopping, and net CO₂ production decreased to about 65% of the initial value. In a separate experiment in which nodule oximetry was used, nodule O₂ permeability decreased 50% within 5 hours in each species. A similar decrease in the O₂-saturated respiration rate (V_max) for the nodule central zone occurred within 5 hours in birdsfoot trefoil, but only after 24 hours in alfalfa. Lb concentration, also measured by oximetry, decreased after 48 to 72 hours. The decrease in permeability preceded the decrease in V_max in each species. V_max may depend mainly on carbohydrate availability in the nodule. If so, then the decrease in permeability could not have been triggered by decreasing carbohydrate availability. Both oximetry and gas exchange data were consistent with the hypothesis that, for the cultivars tested, carbohydrate availability decreased more rapidly in birdsfoot trefoil than in alfalfa nodules. Fractional Lb oxygenation (initially about 0.15) decreased during the first 24 hours after detopping but subsequently increased to >0.65 for a majority of nodules of each species. This increase could lead to O₂ inactivation of nitrogenase.     

Phosphate Deficiency in Maize. IV. Changes in Amounts of Sucrose Phosphate Synthase during the Course of Phosphate Deprivation

With low-P treatment of maize, the level of sucrose phosphatesynthase (SPS) protein decreased to 15% of the control, whilethe "V_max" activity stayed relatively high (100–80% ofthe control) and the substrate limiting activity increased about2 fold in the leaves. These results suggest that leaf phosphatestatus has dual effects on the amount of SPS protein and theactivation state of SPS. (Received January 20, 1992; Accepted April 14, 1993)     

Suppression of the High Affinity Phosphate Uptake System: A Mechanism of Arsenate Tolerance in Holcus lanatus L.

In Holcus lanatus L. phosphate and arsenate are taken up bythe same transport system. Short-term uptake kinetics of thehigh affinity arsenate transport system were determined in excisedroots of arsenate-tolerant and non-tolerant genotypes. In tolerantplants the V_max of ion uptake in plants grown in phosphate-freemedia was decreased compared to non-tolerant plants, and theaffinity of the uptake system was lower than in the non-tolerantplants. Both the reduction in V_max and the increase in K_m ledto reduced arsenate influx into tolerant roots. When the twogenotypes were grown in nutrient solution containing high levelsof phosphate, there was little change in the uptake kineticsin tolerant plants. In non-tolerant plants, however, there wasa marked decrease in the V_max to the level of the tolerant plantsbut with little change in the K_m. This suggests that the lowrate of arsenate uptake over a wide range of differing rootphosphate status is due to loss of induction of the synthesisof the arsenate (phosphate) carrier. Key words: Arsenate, Holcus lanatus L., phosphate uptake, tolerance mechanisms, uptake mechanisms     

Phosphate and Sulfate Activate the Phosphoenolpyruvate Carboxylase from the C4 Plant Cynodon dactylon L.

The effect of phosphate, sulfate and other inorganic ions on the activity of phosphoenolpyruvate carboxylase (PEPC) from the C₄ plant Cynodon dactylon were investigated for the first time, as well as their interaction with Clc-6-P, AMP and ma-late. Activation of PEPC by phosphate and sulfate ions was demonstrated and it was not dependent on the accompanying cations, something that was not clarified for PEPCs from other plant sources. No activation of this enzyme was observed by nitrate. PEPC activation was found to be competitive with glucoses-phosphate (Clc-6-P) and AMP stimulation and less sensitive to malate inhibition. This work showed that PEPC from C₄plants could exhibit similar activation properties with the enzyme from CAM plants and different activation properties in plants of the same type, rendering the study of this enzyme from different plant sources necessary.     

Leaf phosphate status, photosynthesis, and carbon partitioning in sugar beet: I. Changes in growth, gas exchange, and calvin cycle enzymes

Sugar beets (Beta vulgaris L. cv F58-554H1) were cultured hydroponically for 2 weeks in growth chambers with two levels of orthophosphate (Pi) supplied in half strength Hoagland solution. Low-P plants were supplied with 1/20th of the Pi supplied to control plants. With low-P treatment, the acid soluble leaf phosphate and total leaf P decreased by about 88%. Low-P treatment had a much greater effect on leaf area than on photosynthesis. Low-P decreased total leaf area by 76%, dry weight per plant by 60%, and the rate of photosynthesis per area at light saturation by 35%. Low-P treatment significantly decreased the total extractable activity of phosphoglycerate kinase (by 18%) and NADP-glyceraldehyde-3-phosphate dehydrogenase (by 16%), but did not decrease the total activities of ribulose-1,5-bisphosphate (RuBP) carboxylase (RuBPCase) and ribulose-5-phosphate kinase. Low-P treatment decreased the initial activities of three rate-limiting Calvin cycle enzymes, but had no effect on the initial activity of RuBPCase. Furthermore, low-P treatment significantly increased the total extractable activities of fructose-1,6-bisphosphatase (by 61%), fructose-1,6-bisphosphate aldolase (by 53%), and transketolase (by 46%). The results suggest that low-P treatment affected photosynthetic rate through an effect on RuBP regeneration rather than through RuBPCase activity and that the changes in Calvin cycle enzymes with low-P resulted in an increased flow of carbon to starch.     

Sucrose translocation and storage in the sugar beet

Several physiological processes were studied during sugar beet root development to determine the cellular events that are temporally correlated with sucrose storage. The prestorage stage was characterized by a marked increase in root fresh weight and a low sucrose to glucose ratio. Carbon derived from ¹⁴C-sucrose accumulation was partitioned into protein and structural carbohydrate fractions and their amino acid, organic acid, and hexose precursors. The immature root contained high soluble acid invertase activity (V_max 20 micromoles per hour per milligram protein; K_m 2 to 3 millimolar) which disappeared prior to sucrose storage. Sucrose storage was characterized by carbon derived from ¹⁴C-sucrose uptake being partitioned into the sucrose fraction with little evidence of further metabolism. The onset of storage was accompanied by the appearance of sucrose synthetase activity (V_max 12 micromoles per hour per milligram protein; K_m 7 millimolar). Neither sucrose phosphate synthetase nor alkaline invertase activities were detected during beet development. Intact sugar beet plants (containing a 100-gram beet) exported 70% of the translocate to the beet, greater than 90% of which was retained as sucrose with little subsequent conversions.     

Leaf Phosphate Status, Photosynthesis, and Carbon Partitioning in Sugar Beet: III. Diurnal Changes in Carbon Partitioning and Carbon Export

The effect of low phosphate supply (low P) was determined on the diurnal changes in the rate of carbon export, and on the contents of starch, sucrose, glucose, and fructose 2,6-bisphosphate (F2,6BP) in leaves. Low-P effects on the activities of a number of enzymes involved in starch and sucrose metabolism were also measured. Sugar beets (Beta vulgaris L. cv. F58-554H1) were cultured hydroponically in growth chambers and the low-P treatment induced nutritionally. Low-P treatment decreased carbon export from the leaf much more than it decreased photosynthesis. At growth chamber photon flux density, low P decreased carbon export by 34% in light; in darkness, export rates fell but more so in the control so that the average rate in darkness was higher in low-P leaves. Low P increased starch, sucrose, and glucose contents per leaf area, and decreased F2, 6BP. The total extractable activities of enzymes involved in starch and sucrose synthesis were increased markedly by low P, e.g. adenosine 5-diphosphoglucose pyrophosphorylase, cytoplasmic fructose-1,6-bisphosphatase, uridine 5-diphosphoglucose pyrophosphorylase, and sucrose-phosphate synthase. The activities of some enzymes involved in starch and sucrose breakdown were also increased by low P. We propose that plants adapt to low-P environments by increasing the total activities of several phosphatases and by increasing the concentrations of phosphate-free carbon compounds at the expense of sugar phosphates, thereby conserving Pi. The partitioning of carbon among the various carbon pools in low-P adapted leaves appears to be determined in part by the relative capacities of the enzymes for starch and sucrose metabolism.     

Mechanism of photosynthetic response in Microcystis aeruginosa PCC7806 to low inorganic phosphorus

Photosynthetic response of Microcystis aeruginosa PCC7806 to different concentrations of phosphorus supply was studied so as to elucidate if the declining process of Microcystis bloom under freshwater ecosystem is related to soluble reactive phosphorus (SRP) decrease in water volume. Growth rate of M. aeruginosa PCC7806 was significantly reduced under P-deficient conditions, and its photosynthetic activity in terms of rETR_max (maximum electron transport rate) decreased significantly after 48 h growth, while it kept elevating and reached to a relative stable value when supplied with rich phosphorus of 0.6 mg/L. With the increasing actinic irradiance along the rapid light curves of M. aeruginosa PCC7806 cultured under low-phosphorus level, qP (photochemical quenching) and rETR (relative electron transport rate) decreased greatly, and the increase in qN (non-photochemical quenching) and Φ_PS (actual photochemical efficiency of PSII) was obviously inhibited. The affinity of M. aeruginosa PCC7806 to inorganic carbon was reduced evidently in 0.02 mg/L P compared with in 0.6 mg/L P. When P was reduced from 0.6 to 0.02 mg/L, the decreasing rate of rETR_max (77%) was significantly greater than that of photosynthetic carbon assimilation (22%), which indicated that down-regulation of CO₂ affinity caused by P-deficiency was, but not the only reason that resulted in the decline of photosynthetic efficiency. Instantaneous low-temperature significantly limited rETR_max under rich-P condition but had no effect on it when P was insufficient, and 1% ethanol could enhance rETR_max at low-P level but did not influence it at rich-P level. These two results proved that the decrease in thylakoid membrane fluidity caused by P-deficiency was another important reason that results in the decline of photosynthetic efficiency of M. aruginosa PCC7806.     

The relationship of leaf photosynthetic traits – Vcmax and Jmax – to leaf nitrogen,leaf phosphorus,and specific leaf area: a meta‐analysis and modeling study

Great uncertainty exists in the global exchange of carbon between the atmosphere and the terrestrial biosphere. An important source of this uncertainty lies in the dependency of photosynthesis on the maximum rate of carboxylation (V_cmax) and the maximum rate of electron transport (J_max). Understanding and making accurate prediction of C fluxes thus requires accurate characterization of these rates and their relationship with plant nutrient status over large geographic scales. Plant nutrient status is indicated by the traits: leaf nitrogen (N), leaf phosphorus (P), and specific leaf area (SLA). Correlations between V_cmax and J_max and leaf nitrogen (N) are typically derived from local to global scales, while correlations with leaf phosphorus (P) and specific leaf area (SLA) have typically been derived at a local scale. Thus, there is no global-scale relationship between V_cmax and J_max and P or SLA limiting the ability of global-scale carbon flux models do not account for P or SLA. We gathered published data from 24 studies to reveal global relationships of V_cmax and J_max with leaf N, P, and SLA. V_cmax was strongly related to leaf N, and increasing leaf P substantially increased the sensitivity of V_cmax to leaf N. J_max was strongly related to V_cmax, and neither leaf N, P, or SLA had a substantial impact on the relationship. Although more data are needed to expand the applicability of the relationship, we show leaf P is a globally important determinant of photosynthetic rates. In a model of photosynthesis, we showed that at high leaf N (3 gm⁻²), increasing leaf P from 0.05 to 0.22 gm⁻² nearly doubled assimilation rates. Finally, we show that plants may employ a conservative strategy of J_max to V_cmax coordination that restricts photoinhibition when carboxylation is limiting at the expense of maximizing photosynthetic rates when light is limiting.     

Elicitor-Inducible Caffeoyl-Coenzyme A 3-O-Methyltransferase from Petroselinum crispum Cell Suspensions : Purification, Partial Sequence, and Antigenicity

Physiological regulation of nodule gas permeability has a central role in the response of legumes to such diverse factors as drought, defoliation, and soil nitrate. A new method for quantifying nodule respiration and O₂ permeability, based on noninvasive spectrophotometry of leghemoglobin, was evaluated using intact, attached nodules of Lotus corniculatus. First, the relationship between nodule respiration (O₂ consumption) rate and internal O₂ concentration was determined from the rate of decrease in fractional oxygenation of leghemoglobin (FOL) under N₂. The rate of increase of FOL under 100% O₂ was then used to calculate nodule O₂ permeability, after correcting for respiration. Inactivation of nitrogenase by exposure to 100% O₂ for 15 minutes led to decreases in both permeability and O₂-saturated respiration (V_max), but the brief (<15 seconds) exposures to 100% O₂ required by the assay itself had little effect on either parameter. A gradual increase in external O₂ concentration from 20 to 40% resulted in a reversible decrease in permeability, but no change in V_max. The new method is likely to be useful for research on nodule physiology and might also be applicable to agronomic research and crop improvement programs.     

Characterization of phosphate absorption in maize root cortex segments

Uptake of phosphate ions by 1 mm segments of isolated maize root cortex layers was studied. Cortex segments (from roots of 8 days old maize plants) absorb phosphate ions from 1 mM KH₂PO₄ in 0.2 mM CaSCO₄ at the average rate of 34.3 ±3.2 μg P_i g^?1 (fr. m.) h^?1,i.e. 0.35± 0.02 μmol P_i g^?1 (fr. m.) h^?1. Phosphate uptake considerably increases after a certain period of “augmentation”,i.e. washing in aerated 0.2 mM CaSO₄. This increase is completely blocked by the presence of 10 μg ml^?1 cycloheximide. The relation of uptake rate to phosphate concentration in the medium was shown to have 3 phases in the concentration range of 0.02 - 40 mM. Transition points were found between 0.8–1 mM and 10–20 mM. Following K_m and V_max values were found: K_m[mM] : 0.37 - 3.82 - 27.67 V_max[μg P_i g^?1 (fr. m.) h^?1] : 3.33 - 39.40 - 66.67 We have found no sharp pH optimum for phosphate uptake. It proceeds at almost constant rate till pH 6.0 and then the uptake rate drops with increasing pH. At low phosphate concentrations (1 mM) the lowest uptake rate was found at 5 and 13 °C, while the uptake is higher at 5 °C than at 13 °C at phosphate concentrations higher than 1 mM. At these concentrations uptake rate at 35 °C is lower than at 25 °C. Phosphate uptake considerably decreased in anaerobic conditions. DNP and iodoacetate (0.1 mM) completely blocked phosphate uptake from 1 mM KH₂PO₄, while uptake from 5 and 10 mM KH₂PO₄ was left unaffected by these substances. The inhibitors of active - SH groups NEM and PCMB inhibited phosphate uptake: 10^?3 M NEM by 81.6%, 10⁴ M NEM by 42% and 10^?4 M PCMB by 42%.     

Regulation of Ribulose-1,5-Bisphosphate Carboxylase Activity in Response to Changing Partial Pressure of O(2) and Light in Phaseolus vulgaris

The regulation of ribulose-1,5-bisphosphate (RuBP) carboxylase (rubisco) activity in Phaseolus vulgaris was studied under moderate CO₂ and high light, conditions in which photosynthesis in C₃ plants can be insensitive to changes in O₂ partial pressure. Steady state RuBP concentrations were higher, the calculated rate of RuBP use was lower and the activation state of rubisco was lower in low O₂ relative to values observed in normal O₂. It is suggested that the reduced activity of rubisco observed here is related to feedback effects which occur when the rate of net CO₂ assimilation approaches the maximum capacity for starch and sucrose synthesis (triose phosphate utilization). The activation state of rubisco was independent of O₂ partial pressure when light or CO₂ was limiting for photosynthesis. Reduced activity of rubisco was also observed at limiting light. However, in this species light dependent changes in the concentration of an inhibitor of rubisco controlled the apparent V_max of rubisco in low light while changes in the CO₂-Mg²⁺ dependent activation of rubisco controlled the apparent V_max in high light.     

Regulation of Phosphoenolpyruvate Carboxylase from the Green Alga Selenastrum minutum: Properties Associated with Replenishment of Tricarboxylic Acid Cycle Intermediates during Ammonium Assimilation

Two isoforms of phosphoenolpyruvate carboxylase (PEPC) with very different regulatory properties were partially purified from the green alga Selenastrum minutum. They were designated PEPC₁ and PEPC₂. PEPC₁ showed sigmoidal kinetics with respect to phosphoenolpyruvate (PEP) whereas PEPC₂ exhibited a typical Michaelis-Menten response. The S_0.5(PEP) of PEPC₁ was 2.23 millimolar. This was fourfold greater than the S_0.5(PEP) of PEPC₂, which was 0.57 millimolar. PEPC₁ was activated more than fourfold by 2.0 millimolar glutamine and sixfold by 2.0 millimolar dihydroxyacetone phosphate (DHAP) at a subsaturating PEP concentration of 0.625 millimolar. In contrast, PEPC₂ showed only 8% and 52% activation by glutamine and DHAP, respectively. The effects of glutamine and DHAP were additive. PEPC₁ was more sensitive to inhibition by glutamate, 2-oxoglutarate, and aspartate than PEPC₂. Both isoforms were equally inhibited by malate. All of these metabolites affected only the S_0.5(PEP) not the V_max. The regulatory properties of S. minutum PEPC in vitro are discussed in terms of (a) increased rates of dark carbon fixation (shown to be catalyzed predominantly by PEPC) and (b) changes in metabolite levels in vivo during enhanced NH₄₊ assimilation. Finally, a model is proposed for the regulation of PEPC in vivo in relation to its role in replenishing tricarboxylic acid cycle intermediates consumed in NH₄₊ assimilation.     

Overexpression of phosphoenolpyruvate carboxylase from Corynebacterium glutamicum lowers the CO2 compensation point (*) and enhances dark and light respiration in transgenic potato

The effect of decreased or increased phosphoenolpyruvate carboxylase (PEPC) activity on the CO2 compensation point, respiration in the light or dark as well as the partitioning of carbon into starch, soluble sugars, organic acids, and amino acids was investigated using transgenic potato plants. Engineered PEPC activity ranged form 0.5-fold wild-type level in antisense plants to 5-fold wild-type levels in lines overexpressing the ccpc gene of Corynebacterium glutamicum encoding for a PEPC not modulated by protein phosphorylation. The CO2 compensation point determined according to Brooks and Farquhar (1985) was lower in PEPC overexpressors (32 l l^-1 CO2) compared to control potato lines (38 l l^-1 CO2), but was increased in antisense PEPC plants (42 l^-1 CO2). 3-fold overexpression of PEPC gave a minimum CO2 compensation point of 32 l l^-1 CO2. Increased PEPC activity resulted in enhanced respiration in the light and dark. Altered PEPC activity had no effect on the pattern of ¹⁴CO2 incorporation into leaf discs in the light. ¹⁴C pulse-chase experiments in the dark, demonstrated that substantially more total label was lost in the leaf discs from PEPC overexpressors. Metabolite levels were determined in 21 PEPC overexpressing lines after 8 h in the light. A 5-fold increase in PEPC over the wild-type increased malate (61%), starch (75%) and significantly increased sucrose contents 9150%). Total amino acid contents were only marginally increased. From gas exchange characteristics and labeling experiments it was concluded that PEP carboxylation, followed by an increased rate of respiratory CO2 release, might work as a HCO3^-/CO2 pump. This might result in elevated CO2/O2 ratios in the mesophyll, concomitant with a more favoured carboxylation/oxygenation ratio of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco).