Titration of Photophosphorylation in Spinach Chloroplasts with 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU)

The kinetics of the inhibition of photophosphorylation in chloroplasts from spinach (Spinacia oleracea) was investigated with 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) in small concentration intervals, starting at 10^-7M. Plots of the reciprocal of photophosphorylation against concentration of DCMU gave essentially the same straight line with 2 mM nicotinamide adenine dinucleotide phosphate (NADP) together with saturating amounts of ferredoxin or with 4 mM K₃Fe(CN)₆ as the final acceptors for electrons. Practically complete inhibition was obtained at 3 x 10^-6M DCMU. With 0.1 mM flavin mononucleotide (FMN) and ferredoxin, the inhibition between 10^-7M and 10^-6M DCMU was a little slower than in the other two cases. At 10^-6M DCMU a break occurred to a new straight line in the plots, indicating that another reaction was inhibited. Total photophosphorylation without DCMU was about 77 μmol ATP per mg chlorophyll and hour. At the breaking point 20% remained, and inhibition was not complete even at 8 x 10^-6M DCMU. The inhibitor constant for the high-DCMU reaction was in the order of 2 x 10^-5M; for the low-DCMU reaction some complication made the “constant” appear negative. With phenazine methosulfate (PMS) added, DCMU was without effect on photophosphorylation. – As earlier shown by us, titration curves for intact cells of the microalga Scenedesmus show the break at 10^-6M DCMU; and above 6 x 10^-6M photophosphorylation in the algae is not further decreased by DCMU. The data are compared and their possible significance is discussed.     

ATPase activity in plasmalemma-rich vesicles isolated by aqueous two-phase partitioning from Vicia faba mesophyll and epidermis: Characterization and influence of abscisic acid and fusicoccin

Plasmalemma-rich microsomal vesicles were prepared from whole leaf and acid-washed epidermal tissue of Vicia faba L. cv. Osnabrücker Markt by aqueous two-phase partitioning in dextran T-500 and polyethylenglycol 1350 aqueous phases. These vesicles were tightly sealed and predominantly right-side out, and contained a K⁺ -stimulated, mg²⁺-dependent and vanadate-sensitive ATPase. The enzyme from both tissues exhibited nearly identical properties: pH optimum 6.4, K_m for ATP 0.60 mM(whole leaf) and 0.67 mM (epidermis). V_max -480 nmol (mg protein)¹ min¹ (whole leaf) and 510 nmol (mg protein)¹ min¹ (epidermis), I₅₀ (Na₃,VO₄) 7.5 μM (whole leaf) and 15 μM (epidermis). The enzyme was not inhibited by NO₃(50 mM)or sodium azide (I mM). DCCD (20 μM) reduced enzyme activity to 50% (whole leaf) and 58% (epidermis), gramicidin S (20 μM) to 36% (whole leaf) and 41%(epidermis). Ca²⁺ inhibited the ATPase [I₅₀, C²⁺: 0.5 mM(whole leaf) and 0.8 mM(epidermis)]. Ca²⁺ inhibited the ATPase [I₅₀, C²⁺ 0.5 mM(whole leaf) und 0.8 (epidermis)]. The vanadate-sensitive ATPase from whole leaf and epidermal tissue was slightly but significantly stimulated by fusicoccin (FC) at a concentration (0.13 μM) promoting stomatal opening. The stimulation was not seen in the solubilized ATPase. Stomata of the cultivar used here were insensitive lo (±)ABA up to 2 μM level which is effective in most other cultivars and species. Likewise, at this concentration no effect of ABA on the activity of the epidermal ATPase was observed. The data are discussed with respect to the interaction of FC and ABA with the ATPase.     

In intact leaves, the maximum fluorescence level (FM) is independent of the redox state of the plastoquinone pool: A DCMU-inhibition study

The effects of DCMU (3-(3′,4′-dichlorophenyl)-1,1-dimethylurea) on the fluorescence induction transient (OJIP) in higher plants were re-investigated. We found that the initial (F₀) and maximum (F_M) fluorescence levels of DCMU-treated leaves do not change relative to controls when the treatment is done in complete darkness and DCMU is allowed to diffuse slowly into the leaves either by submersion or by application via the stem. Simultaneous 820 nm transmission measurements (a measure of electron flow through Photosystem I) showed that in the DCMU-treated samples, the plastoquinone pool remained oxidized during the light pulses whereas in uninhibited leaves, the F_M level coincided with a fully reduced electron transport chain. The identical F_M values with and without DCMU indicate that in intact leaves, the F_M value is independent of the redox state of the plastoquinone pool. We also show that (i) the generally observed F₀ increase is probably due to the presence of (even very weak) light during the DCMU treatment, (ii) vacuum infiltration of leaf discs leads to a drastic decrease of the fluorescence yield, and in DCMU-treated samples, the F_M decreases to the I-level of their control (leaves vacuum infiltrated with 1% ethanol), (iii) and in thylakoid membranes, the addition of DCMU lowers the F_M relative to that of a control sample.     

Indoleacetic Acid-Induced Decrease of the Ribomiclease Activity in vivo

A study has been made on the influence of indole-3-acetic acid (IAA) on the ribonuclease (RNase) activity in wheat coleoptile sections and green pea stem sections. The hormonal effects on the enzyme activity, ribonncleic acid (RNA) metabolism and growth have been compared. Addition of 10^?5M IAA to the plant sections causes their RNase activity to decrease and their elongation to increase. Removal of the added IAA results in increasing enzyme activity and decreasing growth. The altered enzyme activities are paralleled by opposite changes in the RNA net synthesis. Administration of crystalline RNase to the plant tissue depresses growth. There is thus evidence that the in vivo effect of IAA on the RNase activity is of importance for the hormonal regulation of RNA metabolism and growth. The IAA-induced reduction in the enzyme activity involves cellular metabolism. The effect can be suspended by means of p-chloromercuribenzoate. A possible mechanism for the reduction is discussed.     

The Effect of DCMU (3-(3,4-dichlorophenyl)-1,1-dimethylurea) on Photosynthesis, Glycolate Excretion ('Photorespiration') and Amino Acid Metabolism in the Blue-Green Alga Anacystis

When photosynthesis of the blue-green alga Anacystis nidulans was measured as ¹⁴CO₂-fixation, the inhibitory effect of DCMU at low concentrations was greatest when mainly Photosystem 1 (PS 1) (excitation at 446 or 687 nm) was operative. At concentrations above 10-⁶M the inhibition on ¹⁴CO₂-fixation was greatest when mainly Photosystem 2 (PS 2) was operative (excitation at 619). During excitation of PS 1, the excretion of glycolate was stimulated at low concentrations of DCMU (5 × 10^-8M and lower), while higher concentrations inhibited excretion. All concentrations of DCMU inhibited glycolate excretion when mainly PS 2 was excited. The curves showing the relative effect of DCMU on the two photosystems, measured as PS 1/PS 2, had opposite shapes for ¹⁴CO₂-fixation and glycolate excretion. An increase in ¹⁴CO₂-fixation coincided with a decrease in glycolate excretion and vice versa. It appears that the increased rate of photosynthesis when mainly PS 1 was operative relative to that when mainly PS 2 was excited, increases the consumption of glycolate in an oxidation process associated with the excitation of PS 1, resulting in less excretion of glycolate to the medium. The influence of DCMU inhibition on labelled amino acid pools connected to the glycolate pathway (glycine-serine) is quite similar to that for ¹⁴CO₂-fixation. At concentrations below 10^-6M DCMU, inhibition of ¹⁴CO₂- incorporation into the amino acids was greatest when PS 1 was excited, while at the higher concentrations tested, inhibition was greater when PS 2 was excited. We conclude that the metabolism of glycine and serine is closely connected to the rate of photosynthesis.     

Degradation of chlorinated and non-chlorinated aromatic solvents in soil suspensions by pure bacterial cultures

Summary Experiments were performed to test whether or not high concentrations of CaCl₂ (100 mM) are able to arrest and stabilize internal structures and associated functions in Euglena gracilis Z cells stored in darkness at 4° C. Storage of photoheterotrophically grown green cells in high Ca²⁺ media (2–100 mM) retards pheophytinization of the chlorophylls, preserves photosynthetic activities and stabilizes the structural organization of the associated light-harvesting complexes of the photosystem II units. Alterations of photosynthesis and respiration by chlorpromazine or by temperature are strongly reduced in cells stored under such conditions. More precisely, a chlorpromazine inhibition site is evidenced in the mitochondrial electron pathway and its location in the chloroplastic electron pathway is clarified. Adaptation of Euglena cells from 2 mM to 100 mM Ca²⁺ medium is accompanied by an increase both in the externally bound and total internal calcium concentration. A mechanism involving a Ca²⁺ deposit on internal membranes is proposed. Such interpretation is extended to the storage of cells immobilized in Ca²⁺-alginate gel.Nomenclature (Ca²⁺)ex external calcium concentration - Chl chlorophylls - (Cl^–)ex external chloride concentration - CPZ chlorpromazine or 2-chloro-10-(3-dimethylaminopropyl)-phenothiazine - DCMU diuron or (3,4-dichorophenyl)-1,1-dimethylurea - EGTA ethylene glycol-bis(-aminoethylether) N,N,N ,N-tetraacetic acid - Fc initial level of chlorophyll fluorescence with DCMU - Fmax maximal level of chlorophyll fluorescence with DCMU - Fo level of chlorophyll fluorescence after transients - Ft level of chlorophyll fluorescence with DCMU - Pheo pheophytins - PS I and PS II photosystems I and II - SMi storage medium Offprint requests to: C. Tamponnet     

Nitrite Uptake by Nitrogen-Depleted Wheat Seedlings

Intact, 14-day-old nitrogen-depleted wheat (Triticum vulgare cv. Blueboy) seedlings were exposed to solutions of 0.5 mM KNO₂, 0.05 mM CaSO₄ and 1 mM sodium 2-[N-morpholino]-ethanesulfonate, pH 6.1. Nitrite uptake was determined from depletion of the ambient solution or from incorporation of ¹⁵N in the tissue. An initial nitrite uptake shoulder was followed by a relatively slow uptake rate which subsequently increased to a substantially greater rate. This accelerated phase was maintained through 24 h. Nitrite accumulated to a slight extent in the root tissues during the first few hours but declined to low values when the accelerated rate was fully developed, indicating an increase in nitrite reductase activity paralleling the increase in nitrite uptake capacity. About 50% of the nitrogen absorbed as nitrite was translocated to the shoots by 9–12 h. Development of the accelerated nitrite uptake rate was restricted in excised roots, in intact plants kept in darkness, by 400 μg puromycin ml^?1 and by 1 mM L-ethionine. When puromycin and L-ethionine were added after the accelerated phase had been initiated, their effects were not as detrimental as when they were added at first exposure to KNO₂. The two inhibitors restricted translocation more than uptake. The data indicate an involvement of protein synthesis and a requirement for movement of a substance from shoots to roots for maximal development of the accelerated nitrite uptake phase. A requirement for protein synthesis in the transport of soluble organic nitrogen from roots to shoots is also suggested.     

Translocation of C Sucrose in Sugar Beet during Darkness

The time-course of arrival of ¹⁴C translocate in a sink leaf was studied in sugar beet (Beta vulgaris L. cultivar Klein Wanzleben) for up to 480 minutes of darkness. Following darkening of the source leaf, translocation rapidly declined, reaching a rate approximately 25% of the light period rate by 150 minutes. Comparison of data from plants that were girdled 1 cm below the crown with data from ungirdled plants indicates that after about 150 minutes darkness the beet root becomes a source of translocate to the sink leaf. After about 90 minutes darkness, starch-like reserve polysaccharide from the source leaf begins to contribute ¹⁴C to ethanol soluble pools in that leaf. Because of a 15% isotope mass effect, sucrose, at isotopic saturation, reaches a specific activity which is about 85% of the level of the supplied CO₂. The source leaf sucrose specific activity remains at the isotopic saturation level for about 150 minutes of darkness, after which time input from polysaccharide reserves causes the specific activity to drop to about 55% of that of the supplied CO₂. Sucrose specific activity determinations, polysaccharide dissolution measurements, and pulse labeling experiments indicate that following partial depletion of the sucrose pool, source leaf polysaccharide contributes to dark translocation. Respired CO₂ from the source leaf appears to be derived from a pool which, unlike sucrose, remains at a uniform specific activity.     

Incorporation of 3H-Gibberellin A20 in roots of G2 peas

Following application of ³H-Gibberellin A₂₀ (GA₂₀) to roots of G2 pea seedlings and homogenization of the roots, about 3% of the radioactivity in the tissue could be precipitated from a 30,000 × g supernatant with trichloroacetic acid (TCA) (soluble fraction) while about 5% of the radioactivity pelleted at 30,000 × g (particulate fraction). The radioactivity in the particulate fraction was soluble in sodium dodecyl sulfate (SDS), but was not dialyzable and was insoluble in ethanol. Electrophoresis of the soluble fraction gave only one band of radioactivity, while that of the particulate fraction gave multiple bands. Acid hydrolysis of the soluble fraction released radioactivity that ran coincident with acid-treated GA₂₀ on silicic-acid column chromatography. The particulate fraction gave numerous radioactive peaks following acid hydrolysis, two of which were coincident with GA₂₀ and GA₂₉ (hydroxylation product of GA₂₀) on silicic acid chromatography. Treatment of the particulate and soluble fractions with RNase, DNase, and proteases showed a significant solubilization of radioactivity only with the proteases, suggesting that the GA is bound to a proteinaceous macromolecule. Complete proteolytic hydrolyis followed by thin layer chromatography showed 65% of the radioactivity from the soluble fraction running separately from free GAs or the individual amino acids; the particulate fraction gave mainly (60%) free GAs on enzymatic hydrolysis and much smaller amounts (17%) in a position separate from that of the GAs or amino acids. Binding of ³H-GA to protease-sensitive material was obtained with biologically active ³H-GA₂₀ and ³H-GA₁.     

RNA polymerase activity in isolated nuclei ofNicotiana sanderae callus: Characteristics and modulation during differentiation

Isolated nuclei from differentiating cultures ofNicotiana sanderae showed increased levels of RNA polymerase activity as compared to the nuclei from callus cultures. The RNA synthetic activity was dependent on nucleotide triphosphates and Mg²⁺ and was destroyed by RNase. Maximum activity was obtained in the presence of 50 mM (NH₄)₂ SO₄ and α-amanitin inhibited 40% and 55% of the activity in the nuclei from callus and differentiating tissue respectively. The nuclei from differentiating tissue elicited a 3-fold increase in RNA polymerase I and a 4-fold augmentation in RNA polymerase II activities.     

Properties and subcellular distribution of ribonuclease activity in Chlorella

RNase activity from Chlorella was partially purified. Two RNase activities were demonstrated, one soluble and the other ribosomal. The effects on ribonuclease activity of variations in pH and temperature, and of Mg²⁺, Na⁺, and mononucleotides were examined. The RNase activities (phosphodiesterases EC 3.1.4.23) were both endonucleolytic, releasing oligonucleotides, and cyclic nucleotide intermediates, but exhibited different specificities in releasing mononucleotides from RNA. The ribosomal activity released 3′-GMP, and after prolonged incubation 3′-UMP, but the soluble activity released 3′-GMP, 3′-AMP and 3′-UMP. Neither ofthe RNase preparations hydrolysed DNA, nor released 5′-nucleotides from RNA. Increased ribosomal RNase activity was related to dissociation of ribosomes, and latency of ribosomal RNase activity was demonstrated. The possible in vivo distribution of RNases is discussed.     

Effects of Phosfon-S on Nucleic Acid Metabolism in Pisum sativum Alaska

Phosfon-S, a substance which inhibits stem elongation, alters nucleic acid metabolism in Pisum sativum Alaska. Methylated albumin kieselguhr (MAK) columns were used to fractionate ³²P-labeled nucleic acids. Phosfon-S treatment of the plants resulted in a decrease in soluble RNA and an increase in ribosomal RNA. Specific activities of the various nucleic acid fractions were lower as a result of treatment. The nucleic acids from treated tissues were more resistant to RNase degradation, and endogenous RNase activity was lower in treated tissues. When RNase treated nucleic acids were fractionated on MAK columns, the DNA-RNA fractions from treated plants had a higher specific activity than that of the control, which was not true before nuclease treatment. Spectrophotometric examination of this fraction revealed a difference in absorption spectra, possibly indicating a Phosfon-S nucleic acid complex. It is suggested that these alterations in nucleic acid metabolism could in turn alter a wide variety of metabolic processes, resulting in retarded growth.     

The control by RNA of pyruvate kinase activity in human skeletal muscle

Summary About 25% of total pyruvate kinase activity in human skeletal muscle is associated with the ribonucleoprotein complexes soluble in salt solutions of high ionic strength. These complexes, called form M_B, crystallize readily from 48% saturated ammonium sulfate at pH 5.6.Crystalline preparations represent a heterogenous population of ribonucleoprotein complexes displaying a graduated activity and a variable RNA content. Free protein was not detected in the preparations.Fractionation of crystalline complexes in salt solutions of varying ionic strength and pH, followed by gel filtration on Sephadex G-200 led to the separation of two nucleoprotein fractions with very high specific activity. Fractions containing 30% RNA and 85% RNA respectively revealed a specific activity of 660–670 U/mg protein at 25°C.Pyruvate kinase form M_A was extracted from muscle homogenate with distilled water, purified to homogeneity and crystallized. It contained less than 0.2% RNA and had a specific activity of 270 U/mg. Active ribonucleoprotein complexes gave in double immunodiffusion test the precipitation bands with the anti-M_A sera at the same protein concentration of both antigens, M_B and M_A.Pyruvate kinase M_B with high activity is sensitive to treatment with RNase. Digestion with RNase for 10 min at 25°C diminished the initial specific activity to about one third. Similar residual activity was found in crystalline ribonucleo protein complexes with low RNA content (3.5–20% RNA) which are resistant to further inactivation by RNase.These results implicate the enhancement and control of pyruvate kinase activity by RNA bound to the enzyme.This work was supported by a grant from the Biochemical and Biophysical Committee of Polish Academy of Sciences.     

Titration of Photophosphorylation, Oxidative Phosphorylation and Glycolysis in Scenedesmus with Desaspidin: Regulation between Pathways and Sites for Photophosphorylation

Narrow concentration intervals were used, covering 10^?6– 10^?4M desaspidin. The interaction with glycolysis involves three steps, the inhibitor constants (K_i:s) being in turn 2.7 × 10^?5M, 1.3 × 10^?4M, and high. About 18% of total glycolysis is inhibited in each of the two first steps, and 65% left for the third reaction. After compensation for glycolysis, oxidative phosphorylation may show a sudden jump to about 10% inhibition at 1.5 × 10^?5M desaspidin, the possible K_i of the reaction starting here being very high. Correcting for glycolysis, desaspidin affects total Photophosphorylation in two steps, with the K_i values of 7.8 × 10^?5M and 4.6 × 10^?4M respectively. Inhibition in the first step is about 27% of the total photophosphorylation. By applying 10^?6M DCMU[/3-(3, 4-dichlorophenyl)-l, l-dimethy lurea], one can abolish non-cyclic photophosphorylation. Desaspidin then reacts in a single step with a K_i of 1.4 × 10^?4M. At 5 × 10^?5M DCMU, also the pseudocyclic photophosphorylation is abolished. The remaining, true cyclic photophosphorylation has a single K_i of 2.3 × 10^?5M for desaspidin. Under non-cyclic conditions, the true cyclic process contributes about 25% to total Photophosphorylation. Under pseudocyclic conditions, no cyclic photophosphorylation occurs. Under true cyclic conditions, the non-cyclic and pseudocyclic processes are inoperative. This indicates a regulative system, so that either (1) the (non-cyclic + true cyclic), (2) only the pseudocyclic, or (3) only the true cyclic systems can be traced, dependent on the level of DCMU applied. There are two sites for non-cyclic Photophosphorylation, one of them common to the pseudocyclic pathway. Cyclic photophosphorylation has a third site, different from the other two.     

Membrane ATPase Activity of Barley Roots and Potassium Uptake by Isolated Stele and Cortex

Uptake of potassium ions by isolated stelar tissues of barley from 0.5 and 10 mM K⁺ was respectively 13 and 3.6% of that of the cortical tissues. 0.1 mM H₂PO₄, LO mM ATP and 10 mM Ca(NO₃)₂ did not increase the potassium uptake of either stele or cortex during 5 h of uptake period. A time-course incubation for histological demonstration of the ATPase activity of the plasmalemma and tonoplast of the matured sections of the roots demonstrated a greater activity for the cortical than the stelar tissue. In the stelar parenchyma cells, the plasma lemma showed a higher activity than the tonoplast. These results, which support the “leakiness hypothesis” of the stele, are discussed in relation to the proposed mechanisms of radial ion transport in roots.     

Manganese Absorption and Transport in Rice

The absorptive patterns of Mn²⁺ in excised rice roots, leaf tissues and intact plants, were studied. The rates of absorption of Mn²⁺ followed different patterns in the roots and the leaf tissues. The uptake from 0.1 and 5 mM MnSO₄ was found to be sensitive to metabolic inhibitors. The time course of uptake from 0.1 mM and 5 mM MnSO₄ followed a biphasic pattern which represented only the metabolic component of absorption. A secondary biphasic pattern of uptake at 5 mM MnSO₄ (one at 20 min and another at 80 min) appears quite anomalous and is probably related to structural virations or cellular compartments. When absorption and transport of Mn²⁺ were measured in intact rice and wheat plants, it was found that Mn²⁺ was easily translocated to shoot from roots and the transport of Mn²⁺ was comparable to that of K⁺.     

Stimulation of nitrogenase activity and photosynthesis in some cyanobacteria by glyoxylate

Abstract The effect og glyoxylate on nitrogenase activity (C₂H₂ reduction) and photosynthesis (H¹⁴CO₃ fixation and O₂ evolution) was in vestigated in the three heterocystous cyanobacteria Anabaena cylindrica, A. variabiltis and N. muscorum. Glyoxylate had virtually no effect on the rate of dark respiration and was unable to sustain photoheterotrophic growth, though some slight stimulation (= 30%) of photorophic growth was noted. A considerable stimulation of both nitrogenase and photosynthetic activities was observed in presence of glyoxylate. In the light the stimulation increased with time up to about 15-25 h after adding optimal concentrations of 4–6 mM glyoxylate. Placing glyoxylate treated samples in the dark or adding DCMU (30 μM) in the light, showed that glyoxylate initially supported significantly higher nitrogenase activity than did samples in absence of glyoxylate. However, after a prolonged incubation in the dark or in presence of DCMU glyoxylate is unable to relieve the adverse effects of such conditions. The stimulation of the nitrogenase activity was even more pronounced when the glyoxylate was added to cells preincubated in the dark (“carbon starved”) than for cells kept constantly in light. The results suggest that glyoxylate, or a metabolite, may act as an inhibitor of cyanobacterial photorespiration and this hypothesis is discussed.     

Preparation of Large Oligonucleotides by RNase T1 Partial Hydrolysis of MS2 Virus Treated with Succinic Nhydride

Intact MS2 virus was reacted with succinic anhydride to modify the protein coat and then treated with RNase Tl to obtain controlled hydrolysis of the viral RNA. Viral protein and enzyme were removed by phenol extraction. The RNA fraction was adsorbed on DEAE-cellulose and eluted stepwise with 0.1, 0.5, and 1.0 M NH₄HCO₃, pH 8.6. tRNA^arg, used as a marker, was eluted in the 1.0 M NH₄HCO₃ fraction. The oligomers in the 1.0 M fraction isolated from the viral derivative were further examined by paper chromatography, polyacrylamide gel electrophoresis, and sucrose gradient cestrifuga-tion. Yields of the large oligomer fraction as defined by elution with 1.0 MNH₄HCO₃ from DEAE-cellulose ranged from 51–86% of the amount applied.     

Expression of Three RNase Activities during Natural and Dark-Induced Senescence of Wheat Leaves

We have monitored the activities of RNases WL_A, WL_B, and WL_C (A Blank, TA McKeon [1991] Plant Physiol 97: 1402-1408) during leaf senescence in wheat (Triticum aestivum L. cv Chinese Spring). When seedlings were induced to senesce in darkness, protein loss from primary leaves began immediately. RNase WL_B activity was unchanged for 2 days and then rose linearly, reaching a sixfold elevation in 7 days. RNase WL_C activity declined for 2 days and then rose linearly, reaching a twofold elevation in 7 days. RNase WL_A activity declined in the first 2 days and was unchanged thereafter. Although differentially expressed, these RNase activities may respond to a common regulatory mechanism(s) which, at 2 days of darkness, signals progression into a more advanced stage of senescence. The RNase activities were also differentially expressed during light-induced recovery, returning to normal levels in dissimilar patterns. In flag leaves of greenhouse-grown wheat, the three RNase activities increased during the early postanthesis period when protein content was stable and underwent further, accelerated accumulation during senescence. RNase WL_B activity showed the largest overall senescence-associated elevation (sixfold), followed by RNase WL_C (fourfold) and RNase WL_A (threefold).     

Modulation in vivo by Nitrate Salts of the Activity and Properties of Phosphoenolpyruvate Carboxylase in Leaves of Alternanthera pungens (C4 plant) and A. sessilis (C3 species)

Feeding K⁺ or Na⁺ nitrate salts in vivo enhanced the activity of phosphoenolpyruvate carboxylase (PEPC) in the leaf extracts of Alternanthera pungens (C₄ plant) and A. sessilis (C₃ species). The increase was more pronounced in A. pungens than in A. sessilis. Chloride salts increased the PEPC activity only marginally. However, the sulfate salts were either not effective or inhibitory. Feeding nitrate modulated the regulatory properties of PEPC in A. pungens, resulting in increased K_I (malate) and decreased K_A (glucose-6-P). The sensitivity of PEPC to malate, which gives a measure of phosphorylation status of the enzyme, indicated that feeding leaves with NO₃ ^– enhanced the phosphorylation status of the enzyme. The reduction in PEPC activity due to cycloheximide treatment suggested that increased synthesis of PEPC protein kinase may be one of the reasons for the enhancement in PEPC activity, after the nitrate feeding. We suggest that nitrate salts could be used as a tool to modulate and analyze the properties of PEPC in C₃ and C₄ plants.