Acetate oxidation by bloodstream forms of Trypanosoma cruzi.

Bloodstream forms of Trypanosoma cruzi had a substantial increase in respiration in the presence of acetate. Oxidation of acetate took place via the tricarboxylic acid cycle and involved an antimycin A-sensitive respiratory pathway. Oxygen uptake in the presence of acetate was a sensitive to antimycin A inhibition as was CO2 production. There was a 6--7% residual O2 uptake which was not inhibited by high antimycin concentrations. Human anti-T. cruzi sera had no effect on oxygen uptake.     

Role of <Emphasis Type="Italic">Trypanosoma cruzi</Emphasis> peroxiredoxins in mitochondrial bioenergetics

Trypanosoma cruzi cytosolic (TcCPx) and mitochondrial tryparedoxin peroxidase (TcMPx) play a fundamental role in H₂O₂ detoxification. Herein, mitochondrial bioenergetics was evaluated in cells that overexpressed TcCPx (CPx) and TcMPx (MPx) and in pTEX. In MPx, a higher expression was observed for TcCPx, and the same correlation was true for CPx. Differences in H₂O₂ release among the overexpressing cells were detected when the mitochondrial respiratory chain was inhibited using antimycin A or thenoyltrifluoroacetone. MPx had higher O₂ consumption rates than pTEX and CPx, especially in the presence of oligomycin. In all of the cells, the mitochondrial membrane potential and the ATP levels were similar. Because of the mild uncoupling that was observed in MPx, the presence or induction of a proton transporter in the mitochondrial membrane is suggested when TcMPx is expressed at higher levels. Our results show a possible interplay between the cytosolic and mitochondrial antioxidant systems in a trypanosomatid.     

The Mechanism of Acetate and Pyruvate Oxidation by Trypanosoma cruzi

The epimastigote or culture form of Trypanosoma cruzi oxidizes [3-¹⁴C] pyruvate and [2-¹⁴C] acetate to ¹⁴CO₂ without an apparent increase in overall respiration. This oxidation takes place through the tricarboxylic acid cycle as shown by (a) the incorporation of substrate ¹⁴C into cycle intermediates; (b) the earlier liberation of acetate carboxyl carbon as CO₂; and (c) the characteristic intramolecular distribution of pyruvate and acetate carbon atoms in the skeletal carbon of aspartic and glutamic acids. Upon oxidation of [3-¹⁴C] pyruvate and [2-¹⁴C] acetate, two of the products, alanine and glutamic acid, are found to account for more than 50% of incorporated ¹⁴C; labeling of alanine predominates with [3-¹⁴C] pyruvate while labeling of glutamic acid predominates with [2-¹⁴C] acetate. Using [1- or 6-¹⁴C] glucose as substrate, the pattern of ¹⁴C distribution in soluble metabolites closely resembles that obtained with [3-¹⁴C] pyruvate, in accordance with the joint operation of the Embden-Meyerhof pathway and Krebs cycle. The cycle operation depends on electron transport through the mitochondrial respiratory chain, since antimycin A, at a relatively low concentration, inhibits the oxidation of [2-¹⁴C] acetate to ¹⁴CO₂, to the same extent as the parasite respiration. Though functional in T. cruzi epimastigotes, the oxidative role of the Krebs’ cycle is apparently limited by the absence of an efficient oxidative apparatus. The cycle operation does, however, constitute an important source of skeletal carbon for the biosynthesis of amino acids and can contribute to the process of glycogenesis.     

Titles of related papers in other sections

Culture of Trypanosoma cruzi (Tulahuen strain) in the presence of ethidium bromide (1–20 μg/ml) resulted in dyskinetoplasty and inhibition of growth, to an extent depending on the dye concentration and the medium composition. The ethidium bromide-induced dyskinetoplasty caused a decrease of (a) the cytochrome content of epimastigotes (a,a₃ and b species); (b) the rate of respiration (endogenous or supported by D-glucose); and (c) the rate of production of ¹⁴CO₂ from [2-¹⁴C]acetate and [1-¹⁴C]glucose. [2-¹⁴C]Acetate oxidation to ¹⁴CO₂ was affected by dyskinetoplasty more than [1-¹⁴C]glucose oxidation, particularly at the exponential growth phase. With dyskinetoplastic epimastigotes, diminution of ¹⁴CO₂ production from [2-¹⁴C]acetate largely exceeded that of oxygen uptake, while with [1-¹⁴C]glucose, ¹⁴CO₂production and respiration were affected to about the same extent. Dyskinetoplasty also decreased the incorporation of [2-¹⁴C]acetate carbon into intermediates of the tricarboxylic acid cycle and related amino acids, and modified the distribution pattern of ¹⁴C in accordance with the decrease of respiration. Reduction of cytochrome content of epimastigotes by restriction of heme compounds during growth decreased ¹⁴CO₂ production from [2-¹⁴C]acetate, like the ethidium-induced dyskinetoplasty. The same occurred after inhibition of electron transfer by antimycin and cyanide, though to a much more significant extent, thus confirming the functional association of electron transport at the mitochondrial cytochrome system of T. cruzi and the enzymatic reactions of the tricarboxylic acid cycle.     

Evidence for Cyclic Photophosphorylation during CO(2) Fixation in Intact Chloroplasts: Studies with Antimycin A, Nitrite, and Oxaloacetate

This study examines the effect of antimycin A and nitrite on ¹⁴CO₂ fixation in intact chloroplasts isolated from spinach (Spinacia oleracea L.) leaves. Antimycin A (2 micromolar) strongly inhibited CO₂ fixation but did not appear to inhibit or uncouple linear electron transport in intact chloroplasts. The addition of small quantities (40-100 micromolar) of nitrite or oxaloacetate, but not NH₄Cl, in the presence of antimycin A restored photosynthesis. Antimycin A inhibition, and the subsequent restoration of photosynthetic activities by nitrite or oxaloacetate, was observed over a wide range of CO₂ concentration, light intensity, and temperature. High O₂ concentration (up to 240 micromolar) did not appear to influence the extent of the inhibition by antimycin A, nor the subsequent restoration of photosynthetic activity by nitrite or oxaloacetate. Studies of O₂ exchanges during photosynthesis in cells and chloroplasts indicated that 2 micromolar antimycin A stimulated O₂ uptake by about 25% while net O₂ evolution was inhibited by 76%. O₂ uptake in chloroplasts in the presence of 2 micromolar antimycin A was 67% of total O₂ evolution. These results suggest that only a small proportion of the O₂ uptake measured was directly linked to ATP generation. The above evidence indicates that cyclic photophosphorylation is the predominant energy-balancing reaction during photosynthesis in intact chloroplasts. On the other hand, pseudocyclic O₂ uptake appears to play only a minimal role.     

Trypanosoma cruzi dihydrolipoamide dehydrogenase as target of reactive metabolites generated by cytochrome c/hydrogen peroxide (or linoleic acid hydroperoxide)/phenol systems

Abstract

This study determines that cytochrome c (cyt c) catalyses the oxidation of phenol compounds (Phen) in the presence of H₂O₂ or linoleic acid hydroperoxide (LOOH), generating Phen-derived free radicals or other reactive metabolites. These products irreversibly inactivated the dihydrolipoamide dehydrogenase from Trypanosoma cruzi (T cruzi LADH), depending on: the Phen structure, peroxide type, activated cyt c, incubation time and presence of an antioxidant. Nordihydroguaiaretic acid (NDGA) and caffeic acid (CAFF) with cyt c/H₂O₂ or cyt c/LOOH were the most effective inhibitors of T cruzi LADH. The comparison of inactivation values for T cruzi and mammalian heart enzymes demonstrated a greater sensitivity of T cruzi LADH to Phen. GSH, N-acetylcysteine, NAD(P)H, ascorbate and trolox, prevented T cruzi LADH inactivation by acetaminophen. The role of the Phen as potential trypanocidal systems is discussed.     

Fermentation parameters of Peptostreptococcus productus on gaseous substrates (CO,H2/CO2)

Intrinsic growth and substrate uptake parameters were obtained for Peptostreptococcus productus, strain U-1, using carbon monoxide as the limiting substrate. A modified Monod model with substrate inhibition was used for modeling. In addition, a product yield of 0.25 mol acetate/mol CO and a cell yield of 0.034 g cells/g CO were obtained. While CO was found to be the primary substrate, P. productus is able to produce acetate from CO₂ and H₂, although this substrate could not sustain growth. Yeast extract was found to also be a growth substrate. A yield of 0.017 g cell/g yeast extract and a product yield of 0.14 g acetate/g yeast extract were obtained. In the presence of acetate, the maximum specific CO uptake rate was increased by 40% compared to the maximum without acetate present. Cell replication was inhibited at acetate concentrations of 30 g/l. Methionine was found to be an essential nutrient for growth and CO uptake by P. productus. A minimum amount of a complex medium such as yeast extract (0.01%) is, however, required.     

Regulation of nonphosphorylating electron transport pathways in soybean cotyledon mitochondria and its implications for fat metabolism

The respiration of mitochondria isolated from germinating soybean cotyledons was strongly resistant to antimycin and KCN. This oxygen uptake was not related to lipoxygenase which was not detectable in purified mitochondria. The antimycin-resistant rate of O₂ uptake was greatest with succinate as substrate and least with exogenous NADH. Succinate was the only single substrate whose oxidation was inhibited by salicyl hydroxamic acid alone, indicating engagement of the alternative oxidase. Concurrent oxidation of two or three substrates led to greater involvement of the alternative oxidase. Despite substantial rotenone-resistant O₂ uptake with NAD-linked substrates, respiratory control was observed in the presence of antimycin, indicating restriction of electron flow through complex I. Addition of succinate to mitochondria oxidizing NAD-linked substrates in state four stimulated O₂ uptake substantially, largely by engaging the alternative oxidase. We suggest that these properties of soybean cotyledon mitochondria would enable succinate received from the glyoxysome during lipid metabolism to be rapidly oxidized, even under a high cytosolic energy charge.     

Factors affecting glucose uptake by the ruminal bacterium Bacteroides ruminicola

Summary Glucose uptake by whole cells of Bacteroides ruminicola B₁4 is constitutive. Potassium concentrations between 10 and 150 mm stimulated uptake over fourfold, while sodium had little effect on uptake. The involvement of potassium in glucose uptake by B. ruminicola was supported by strong inhibition of uptake by the ionophores valinomycin, lasalocid, and monensin. The electron transport inhibitor antimycin A had little effect on uptake, but menadione and acriflavine inhibited uptake by 30 and 48%, respectively. Potent inhibitors of uptake included oxygen, p-chloromercuribenzoate, HgCl₂, and o-phenanthroline. Sodium arsenate decreased uptake by 40%, suggesting that a high-energy phosphate compound and possibly a binding protein may be involved in glucose uptake. The protonophores carbonyl cyanide m-chlorophenylhydrazone and 2,4-dinitrophenol inhibited glucose uptake by 37 and 22%, respectively. Little change in uptake activity was observed at extracellular pH values between 4.0 and 8.0. Excess (10 mm) cellobiose, maltose, and sucrose inhibited glucose uptake less than 15%. High levels (0.15% w/v) of p-coumaric acid and vanillin decreased uptake by 32 and 37%, respectively, while 0.15% ferulic acid decreased uptake by 15%.     

Effect of ubiquinone extraction on the reaction of the mitochondrialbc 1 complex with ferricyanide

Depletion of endogenous ubiquinone by pentane extraction of mitochondrial membranes lowered succinate-ferricyanide reductase activity, whereas quinone reincorporation restored the enzymatic activity as well as antimycin sensitivity. The oxidant-induced cytochromeb extrareduction, normally found upon ferricyanide pulse in intact mitochondria in the presence of antimycin, was lost in ubiquinone-depleted membranes, even if cytochromec was added. Readdition of ubiquinone-2 restored the oxidant-induced extrareduction with an apparent half saturation at 1 mol/molbc ₁ complex saturating at about 5 mol/mol. These findings demonstrate a requirement for the ubiquinone pool of the cytochromeb extrareduction. Since the initial rates of cytochromeb reoxidation upon ferricyanide addition, in the presence of antimycin, did not saturate by any ferricyanide concentration in ubiquinone-depleted mitochondria, a direct chemical reaction between ferricyanide and reduced cytochromeb was postulated. The fact that such direct reaction is much faster in ubiquinone-depleted mitochondria may explain the lower antimycin sensitivity of the succinate ferricyanide reductase activity after removal of endogenous ubiquinone.     

Light-driven reduction of oxygen as a method for studying electron transport in the green photosynthetic bacterium Chlorobium limicola

The use of O₂ uptake as a valid assay for non-cyclic photosynthetic electron flow in membranes from Chlorobium limicola is discussed. It is recommended that methyl viologen, catalase and superoxide dismutase should be added to the experimental medium. The addition of methyl viologen more than doubled the rate of O₂ uptake observed on illumination with 1 mM sulphide as donor. Superoxide dismutation was shown to be efficient under the experimental conditions by means of standard additions of potassium superoxide dissolved in dimethylsulphoxide. The highest rates of light stimulated O₂ uptake were obtained with sulphide as electron donor, and approached 50 mol O₂ · h^-1 · mg bacteriochlorophyll c ^-1 with 0.2 mM sulphide. The presence of 5 mM 2-mercaptoethanol or 3 mM sulphite as electron donor led to lower light stimulated rates of O₂ uptake, while 5 mM thiosulphate had little effect. The rates were insensitive to uncoupler. The light stimulated O₂ uptake with 0.2 mM sulphide as donor was 20–30% inhibited by 10 M antimycin A and 50 M cyanide.Abbreviations APS Adenosine 5-phosphosulphate - FCCP carbonylcyanide-p-trifluoromethoxyphenylhydrazone - HEPES 4-(2-hydroxyethyl)-1-piperazineethanesulphonic acid - MeV methyl viologen - P-840 the photoreactive bacteriochlorophyll     

Absence of Transitory [Ca2+]I Flux During Early In Vitro Metacyclogenesis of Trypanosoma cruzi1

ABSTRACT. The phorbol ester TPA (phorbol 12-myristate 13-acetate) substitutes for CO₂ as an agonist for transforming Trypanosoma cruzi epimastigotes to the metacyclic trypomastigote stage in a starvation medium consisting of phosphate buffered saline + 10 mM proline, 10 mM sodium acetate and 0.035% NaHCO₃. Since TPA is thought to stimulate protein kinase C by mimicking the activity of the secondary messenger diacylglycerol, the above result suggested that T. cruzi metacyclogenesis could be activated by a Ca^2+-dependent protein kinase C signal induction pathway. Accordingly, cytosolic calcium flux ([Ca²⁺]_i) in epimastigotes, activated with 5% CO₂ or TPA (10^-7 M), was measured with the Ca²⁺ molecular probe, fluo-3AM. In addition, [Ca²⁺]_i was measured in cells incubated with putative metacyclogenic agonists (e.g. proline, glutamate, bioamines, ionophores and catecholamines). None of the compounds studied, except for EGTA, affected cytosolic Ca²⁺ levels. Control assays with 11 μM thapsigargin, which mobilizes noncytoplasmic Ca²⁺ stores by inhibiting endoplasmic reticulum Ca²⁺-ATPase. validated our fluorometric assay procedure. Although thapsigargin significantly increases cytoplasmic Ca²⁺ fluorescence, it has no effect on transformation. The protein kinase C inhibitors staurosporine, H-7 and HA 1004 were tested for their effect on T. cruzi metacyclogenesis. Low concentrations of staurosporine and HA 1004 significantly elevated Pent strain transformation while H-7 had no effect on Peru strain metacyclogenesis. Inhibitor H-7 did significantly depress CL transformation. the results indicate that induction of T. cruzi metacyclic trypomastigote formation by CO₂ and TPA is not accompanied by changes in cytosolic Ca²⁺ and do not provide supporting evidence for participation of a protein kinase C-mediated phosphoinositide cascade in metacyclogenesis.     

Proton nuclear magnetic resonance analysis of metabolic end products of the Bolivia strain of Trypanosoma cruzi and three of its clones

Proton nuclear magnetic resonance (¹H NMR) was used to study the in vivo metabolism of Trypanosoma cruzi, the pathogen causing American trypanosomiasis (Chagas' disease). Three clones were isolated from a strain of T. cruzi (Bolivia strain), The clones I, II and III and the original strain were characterized according to the spectra of their metabolic pathways to test the hypothesis that clonal evolution of T. cruzi has a major impact on biologically relevant properties of this parasite. T. cruzi (Bolivia strain) excreted acetate, alanine, glycerol, and succinate as major end products, in the proportion 6:4:2:2. Comparing the spectra of T. cruzi clones with the original Bolivia strain revealed both quantitative, as well as qualitative differences in the metabolites excreted: the clones I and II, as opposed to the Bolivia strain and clone III, excreted significant quantities of ethanol.     

Competition between Fe(III)-Reducing and Methanogenic Bacteria for Acetate in Iron-Rich Freshwater Sediments

The kinetics of acetate uptake and the depth distribution of [2-¹⁴C]acetate metabolism were examined in iron-rich sediments from a beaver impoundment in northcentral Alabama. The half-saturation constant (K_m) determined for acetate uptake in slurries of Fe(III)-reducing sediment (0.8 µM) was more than 10-fold lower than that measured in methanogenic slurries (12 µM) which supported comparable rates of bulk organic carbon metabolism and V_max values for acetate uptake. The endogenous acetate concentration (S _n) was also substantially lower (1.7 µM) in Fe(III)-reducing vs methanogenic (9.0 µM) slurries. The proportion of [2-¹⁴C]acetate converted to ¹⁴CH₄ increased with depth from ca 0.1 in the upper 0.5 cm to ca 0.8 below 2 cm and was inversely correlated (r² = 0.99) to a decline in amorphous Fe(III) oxide concentration. The results of the acetate uptake kinetics experiments suggest that differences in the affinity of Fe(III)-reducing bacteria vs methanogens for acetate can account for the preferential conversion of [2-¹⁴C]acetate to ¹⁴CO₂ in Fe(III) oxide-rich surface sediments, and that the downcore increase in conversion of [2-¹⁴C]acetate to ¹⁴CH₄ can be attributed to progressive liberation of methanogens from competition with Fe(III) reducers as Fe(III) oxides are depleted with depth.     

Fermentative Metabolism of Chlamydomonas reinhardii: III. Photoassimilation of Acetate

The anaerobic photodissimilation of acetate by Chlamydomonas reinhardii F-60 adapted to a hydrogen metabolism was studied utilizing manometric and isotopic techniques. The rate of photoanaerobic (N₂) acetate uptake was approximately 20 μmoles per milligram chlorophyll per hour or one-half that of the photoaerobic (air) rate. Under N₂, cells produced 1.7 moles H₂ and 0.8 mole CO₂ per mole of acetate consumed. Gas production and acetate uptake were inhibited by monofluoroacetic acid (MFA), 3-(3′,4′-dichlorophenyl)-1,1-dimethylurea (DCMU) and by H₂. Acetate uptake was inhibited about 50% by 5% H₂ (95% N₂). H₂ in the presence of MFA or DCMU stimulated acetate uptake and the result was interpreted to indicate a transition from oxidative to reductive metabolism. Carbon-14 from both [1-¹⁴C]- and [2-¹⁴C]acetate was incorporated under N₂ or H₂ into CO₂, lipids, and carbohydrates. The methyl carbon of acetate accumulated principally (75-80%) in the lipid and carbohydrate fractions, whereas the carboxyl carbon contributed isotope primarily to CO₂ (56%) in N₂. The presence of H₂ caused a decrease in carbon lost from the cell as CO₂ and a greater proportion of the acetate was incorporated into lipid. The results support the occurrence of anaerobic and light-dependent citric acid and glyoxylate cycles which affect the conversion of acetate to CO₂ and H₂ prior to its conversion to cellular material.     

All-atom molecular dynamics simulations reveal significant differences in interaction between antimycin and conserved amino acid residues in bovine and bacterial bc1 complexes

Antimycin A is the most frequently used specific and powerful inhibitor of the mitochondrial respiratory chain. We used all-atom molecular dynamics (MD) simulations to study the dynamic aspects of the interaction of antimycin A with the Q_i site of the bacterial and bovine bc₁ complexes embedded in a membrane. The MD simulations revealed considerable conformational flexibility of antimycin and significant mobility of antimycin, as a whole, inside the Q_i pocket. We conclude that many of the differences in antimycin binding observed in high-resolution x-ray structures may have a dynamic origin and result from fluctuations of protein and antimycin between multiple conformational states of similar energy separated by low activation barriers, as well as from the mobility of antimycin within the Q_i pocket. The MD simulations also revealed a significant difference in interaction between antimycin and conserved amino acid residues in bovine and bacterial bc₁ complexes. The strong hydrogen bond between antimycin and conserved Asp-228 (bovine numeration) was observed to be frequently broken in the bacterial bc₁ complex and only rarely in the bovine bc₁ complex. In addition, the distances between antimycin and conserved His-201 and Lys-227 were consistently larger in the bacterial bc₁ complex. The observed differences could be responsible for a weaker interaction of antimycin with the bacterial bc₁ complex.     

The response of intact Strongyloides ratti infective (L3) larvae to substrates and inhibitors of respiratory electron transport

Live, intact third-stage larvae (L3s) of Strongyloides ratti in the absence of exogenous substrates consumed oxygen at a rate (E-QO₂) of 181.8 ± 12.4 ng atoms min⁻¹ mg dry weight⁻¹ at 35°C. Respiratory electron transport (RET) Complex I inhibitor rotenone (2 μm) produced 33 ± 6.5% inhibition of the E-QO₂. Unusually the rotenone-induced inhibition was not relieved by 5 μm-succinate. The E-QO₂ of intact L3s was refractory to RET Complex III inhibitor antimycin A at 2 μm; 4 μm-antimycin inhibited ≤ 10% of the E-QO₂. The electron donor couple ascorbate/TMPD augmented the E-QO₂ in the presence of rotenone (2 μm) and antimycin A (4 μm) by 110%. Azide (1 mm) stimulated the antimycin A refractory QO₂ by 36.6 ± 7.2% which was only partially inhibited by 1.0 mm-KCN (IC₅₀ = 0.8 mm). The data suggest the presence of classical (CPW) and alternate (APW) electron transport pathways in S. ratti L3s.     

Mass spectrometric determination of O2 gas exchange during a dark-to-light transition in higher-plant cells

The exchange of O₂ and CO₂ by photoautotrophic cells of Euphorbia characias L. was measured using a mass-spectrometry technique. During a dark-tolight transition the O₂ uptake rate was little affected whereas CO₂ efflux was decreased by 40%. In order to differentiate eventual superimposed O₂-uptake processes, the kinetics of O₂ exchange resulting from brief illuminations were measured with a highly sensitive device. When the cells were exposed to a saturating light for short periods, the rate of O₂ uptake passed through a series of transients: there was first a stimulation occurring 2–3 s after the appearance of O₂ from water-splitting, followed 30 s later by an inhibition. These two transients were reduced 80% by 3-(3,4-dichlorophenyl)1, 1-dimethylurea (DCMU), indicating that they relied on the linear transport of electrons in the chloroplasts. The first transient (stimulation of an O₂ uptake) was little affected by mitochondrial inhibitors such as antimycin A and oligomycin or the uncoupler carbonylcyanide m-chlorophenylhydrazone (CCCP) but was increased in presence of KCN. When spaced flashes (2 us duration; 100-ms intervals) were used instead of continuous light, this transient was almost suppressed indicating that it was dependent on the saturation of some component of the chloroplastic chain. The second transient (inhibition of O₂ uptake) was present when spaced flashes were used instead of continuous light. It was markedly decreased by addition of CCCP and mitochondrial inhibitors (antimycin A, oligomycin, KCN) which strongly indicates that it relied on mitochondrial respiration. It is concluded from these experiments that illumination of the cells resulted in an inhibition of mitochondrial respiration, but the resulting inhibition of O₂ uptake was hidden by the appearance of an O₂-uptake process of extramitochondrial origin, presumably located in the chloroplast.Abbreviations CCCP carbonylcyanide mchlorophenylhydrazone - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - Rubisco ri-bulose-1,5-bisphosphate carboxylase/oxygenase The authors thank Drs A. Vermeglio, P. Thibault and P. Gans for helpful discussions.     

Carrier-mediated acetate uptake in Corynebacterium glutamicum

Acetate is effectively taken up by whole cells of Corynebacterium glutamicum via a specific carrier with a pH optimum of 8. The K _m of acetate uptake was 50 μM and the V _max 25–35 nmol/mg dw min. The activation energy was determined to be 70 kJ/mol. Acetate uptake was competitively inhibited by propionate with a K _i of about 30 μM and blocked by addition of sulfhydryl reagents. The transport activity was clearly dependent on the membrane potential, but independent of the presence of Na⁺-ions. It is concluded that uptake of acetate proceeds by a secondary, proton coupled mechanism.     

PHOTOASSIMILATION OF 14C-ACETATE BY ULVA LACTUCA1

The effect of light on the uptake of ¹⁴C-labeled acetate, glucose, α-ketoglutarate, mannitol, and glycine was investigated in Ulva lactuca var. rigida. Uptake in the light over that in the dark of ¹⁴C-acetale (8-fold) was far higher than that of the other compounds tested. Further study of the phenomenon showed that (1) an increase in light intensity from 60 through 1000 ft-c results in increased ¹⁴C-acetate uptake, the kinetics of which differ from that of ¹⁴CO₂ uptake versus light intensity over the same range: (2) the action spectrum of acetate uptake, between 450 and 730 nm conforms to the action spectrum for photosynthesis in Ulva; (3) the acetate uptake process is sensitive to photosynthetic poisons including N'- (3-4, dichlorophenyl)-N, N-dimethyl urea (DCMU), and phenazine methosulfate (PMS), and inhibitors of the Krebs cycle including sodium monofluoroacetate (MFA) and malonate; (4) uptake of acetate is favored by low CO₂ concentrations; (5) uptake of acetate is not sensitive to 10^?4 M uranyl nitrate; (6) continuous white light, with and without far-red radiation, indicates no significant phytochrome involvement in the process. These results point to an intracellular dependence of the assimilation of acetate on the Krebs cycle in some manner (possibly in cooperation with the glyoxylate cycle) other than simply releasing CO₂ from the acetate moiety with subsequent fixation via the Calvin cycle, and on some product(s) of photo-system 2, eg, NADPH⁺, reduced ferredoxin, or O₂.