Repression of the K+ Uptake and Cation-Stimulated ATPase Activity Associated with the Plasma Membrane-Enriched Fraction of Cucumber Roots Due to Ca2+ Starvation

The uptake of K⁺ by cucumber plants decreased markedly duringCa²⁺ starvation. A plasma membrane-enriched fraction, judgedfrom the distribution of marker enzymes, was prepared from controland Ca²⁺-starved roots. The Mg²⁺- and K⁺-Mg²⁺-ATPase activitiesassociated with the plasma membrane-enriched fraction of controlroots were maxima at pH 6.5. Various monovalent cations andpotassium salts of monovalent anions stimulated Mg²⁺-ATPaseactivity. Vanadate, DES and DCCD inhibited K⁺- Mg²⁺-ATPase activity.Of the divalent cations and phosphate esters tested, Mg²⁺ andATP were most effective for the stimulation of ATPase by K⁺,whereas Ca²⁺ was ineffective in replacing Mg²⁺. Mg²⁺- and K⁺-Mg²⁺-ATPase activities associated with the plasmamembrane enriched fraction of Ca²⁺-starved roots were much lowerthan those of control roots. K_m values of K⁺-Mg²⁺-ATPase forATP were comparable for control and Ca²⁺-starved roots. The K⁺-stimulated activity of Mg²⁺-ATPase in Ca²⁺-starved rootswas approximately one fourth that of the control, whereas therate of stimulation was only slightly lower in Ca²⁺-starvedroots. (Received May 9, 1984; Accepted September 17, 1984)     

Characterization of ATPase Activity Associated with Plasma Membrane from Vigna Hypocotyls

A plasma membrane fraction was isolated from the hypocotylsof cowpea {Vigna unguiculata) by a combination of differentialcentrifugation and sucrose density gradient centrifugation.The ATPase activity of this fraction was dependent on divalentcations (Mn²⁺>Mg²⁺>Co²⁺>Ca²⁺>Fe²⁺>Zn²⁺>Ni²⁺)but was not further stimulated by monovalent cations (K⁺ and/orNa⁺). The pH optimum for the activation of ATPase by Mg²⁺ was7.0. This fraction hydrolyzed ATP or UTP as a substrate andthe ATPase activity obeyed a Michaelis-Menten type of kinetics.The K_m for MgATP ranged from 0.65 to 1.1 mM. The ATPase activitywas inhibited by inhibitors such as N, N'- dicyclohexylcarbodiimide,diethylstilbestrol and triphenyltin chloride, all of which arereported to block proton (H⁺) transport in plant cells, butwas insensitive to those of mitochondrial ATPase such as oligomycinand sodium azide. The ATPase activity was not stimulated bytreatment with ionophores (e.g., carbonyl cyanide p-trifluoromethoxyphenylhydrazone,3,5-di-ter-butyl-4-hydroxybenzilidenemalononitrile and valinomycin⁺KCl)which would be expected to dissipate the electrochemical potentialdifference of H⁺ or the membrane potential difference. The characteristics of the ATPase are compared with those ofplasma membrane ATPases of other plants and its possible rolein H⁺-transport is discussed. ¹ Present address: Institute of Applied Biochemistry, Yagi MemorialPark, Mitake, Gifu 505-01, Japan or Laboratory for Plant EcologicalStudies, Faculty of Science, Kyoto University, Kyoto 606, Japan. (Received April 20, 1984; Accepted August 14, 1984)     

Comparisons of Plasma Membranes from Shoots and Roots of Winter Rye (Secale cereale L. cv. Puma): Polypeptide Composition, ATPase Activity and Specific Naphthylphthalamic Acid Binding Capacity

Polypeptide compositions, ATPase characteristics, and the N-1-naphthylphthalamicacid binding capacity of plasma membranes prepared from winterrye (Secale cereale L. cv. Puma) shoots and roots were examinedand compared. Some unique polypeptides were revealed in each plasma membraneby one- and two-dimensional slab gel electrophoresis. A differencewas also detected in glycopeptide compositions. The plasma membranesfrom both organs contained Mg²⁺-stimulated ATPase exhibitingslightly different properties in the divalent cation specificityand the kinetic constants. The ATPase activities from both organsshowed a similar optimum pH around 6.5, simple Michaelis-Mentensaturation with increasing ATP-Mg concentrations, and littleK⁺-stimulation at the optimum pH. Both ATPases were inhibitedby orthovanadate, however, the degree of inhibition was a littledifferent in each membrane sample. The specific N-1-naphthylphthalamicacid binding capacity in the shoot plasma membrane was 2.6-foldhigher than that in the root plasma membrane. These results suggest that polypeptide compositions of plasmamembranes vary corresponding with a difference in the physiologicalfunctions of plasma membranes between shoots and roots of winterrye. ¹ Contribution No. 2670 from the Institute of Low TemperatureScience. (Received May 17, 1984; Accepted October 9, 1984)     

Presence of a Na+-activated ATPase in the Plasma Membrane of the Marine Raphidophycean Heterosigma akashiwo

Highly purified plasma membranes were isolated from Heterosigmaakashiwo cells, a marine raphidophycean unicellular biflagellate,by the silica microbead method, and the ATPase activity of themembranes was characterized. The ionic requirements and spectrumof effective inhibitors enable us to identify a novel Na⁺-activatedATPase in the plasma membrane of this organism. Furthermore,we detected two phosphorylated intermediate forms of ATPases,with molecular weights of 150 kDa and 95 kDa as judged by acidSDS-polyacrylamide gel electrophoresis of extracts of isolatedplasma membrane. The 150 kDa intermediate was phosphorylated in the presenceof both Mg²⁺ and Na⁺, while the 95 kDa intermediate was phosphorylatedin the presence of Mg²⁺ alone. Both were dephosphorylated inthe presence of monovalent cations. These results indicate thatthe former intermediate was a Na⁺-activated ATPase, similarto Na⁺,K⁺-ATPases from animals, and the latter was similar toH⁺,K⁺-ATPases from higher plants. The physiological significanceof the two kinds of ATPase in the plasma membrane of marinealgae. (Received March 15, 1989; Accepted June 23, 1989)     

Reconstitution and Characterization of H+-Translocating ATPase from the Plasma Membrane of Phaseolus mungo L. Roots

Plasma membrane H⁺-translocating ATPase was partially purifiedfrom mung bean (Phaseolus mungo L.) roots and reconstitutedinto soybean phospholipid (asolectin) liposomes by the n-octylglucosidedilution method. The resulting proteoliposomes were mainly unilamellarvesicles ranging in size from 0.05 to 0.2 µm. The existenceof ATP-drived H⁺-pumping across the proteoliposomes was demonstratedby the quenching of quinacrine fluorescence in the presenceof Mg²⁺. The quenching could be abolished by an uncoupler, FCCP,and an inhibitor of H⁺-translocating ATPase, vanadate. The reconstitutedATPase consisted of three major polypeptides of 105 KDa, 67KDa and 57 KDa. Its pH optimum, divalent cation stimulationand vanadate sensitivity were similar to those of partiallypurified ATPase. However, the specificity toward ATP was muchgreater following reconstitution. Also reconstitution reducedthe degree of inhibition by DCCD. Local anesthetics (e.g. dibucaine)had no effect on H⁺-pumping activity but increased the ATPaseactivity when proteoliposomes were reconstituted in their presence. (Received May 2, 1986; Accepted October 17, 1986)     

Effects of cations on the sugar receptor-site of the blowfly, Phormia

1) Ca^{+ +} (1 to 10 mM) lowered the binding affinity of sugarreceptor-site for sucrose in the labellar sugar receptor ofthe blowfly, Phormia regina, without changing the maximum-responseamplitude. It also elevated the values of the Hill coefficient(n_H) in some degrees. 2) Other divalent cations such as Mg^{+ +}, Ba^{+ +} or Cd^{+ +} alsoshowed almost the same property as above. The sequence of theeffect is as follows: Ba^{+ +}, Mg^{+ +} x Ca^{+ +} x Cd^{+ +}. Trivalentcation, La^{+ + +} (1 mM), changed the value of n_H from 1 (La^{++ +}-free) to 2. 3) On the contrary, the action of monovalent cations such asK⁺ or Na⁺, of which ionic strength was made the same as thatof the divalents hardly suppressed the response. 4) The results obtained do not support the hypothesis, at leaston the sugar receptor of the fly, that the receptor potentialis attributable to a change of the surface potential (zeta potential)as is proposed for the frog sugar receptor.     

Effects of the Herbicide MCPA, and the Heavy Metals, Cadmium and Nickel on the Lipid Composition, Mg2+-ATPase Activity and Fluidity of Plasma Membranes from Rice, Oryza sativa (cv. Bahia) Shoots

Application of the herbicide 4-chloro-2-methylphenoxyaceticacid (MCPA), cadmium or nickel to the roots of rice, Oryza sativa(cv. Bahia) grown in solution culture resulted in changes tothe lipid composition and increases in the Mg²⁺-ATPase activityof the shoot plasma membranes. These increases in Mg²⁺-ATPaseactivity were associated with reductions in the campesterol:sitosterol ratio. There was no significant effect on membranefluidity (measured by steady-state fluorescence polarization)as a result of the treatments. The results suggested that therewas no consistent relationship between fluidity and Mg²⁺-ATPaseactivity. A possible involvement of sterols in the regulationof plasma membrane-bound ATPase activity is proposed. Key words: MCPA, heavy-metals, sterols, ATPase activity, plasma membranes     

Cation and Chloride Partitioning through Xylem and Phloem within the Whole Plant of Ricinus communis L. under Conditions of Salt Stress

Uptake and partitioning through the xylem and phloem of K⁺,Na⁺, Mg²⁺ , Ca²⁺ and Cl^– were studied over a 9 d intervalduring late vegetative growth of castor bean (Ricinus communisL.) plants exposed to a mean salinity stress of 128 mol m^–3NaCl. Empirically based models of flow and utilization of eachion within the whole plant were constructed using informationon ion increments of plant parts, molar ratios of ions to carbonin phloem sap sampled from petioles and stem internodes andpreviously derived information on carbon flow between plantsparts in xylem and phloem in identical plant material. Salientfeatures of the plant budget for K⁺ were prominent depositionin leaves, high mobility of K⁺ in phloem, high rates of cyclingthrough leaves and downward translocation of K⁺ providing theroot with a large excess of K⁺ . Corresponding data for Na⁺showed marked retention in the root, lateral uptake from xylemby hypocotyl, stem internodes and petioles leading to low intakeby young leaf laminae and substantial cycling from older leavesback to the root. The partitioning of the anionic componentof NaCl salinity, Cl^–, contrasted to that of Na⁺ in thatit was not substantially retained in the root, but depositedmore or less uniformly in stem, petiole and leaf lamina tissues.The flow pattern for Mg²⁺ showed relatively even depositionthrough the plant but some preferential uptake by young leaves,generally lesser export than import by leaf laminae, and a returnflow of Mg²⁺ from shoot to root considerably less than the recordedincrement of the root. Ca²⁺ partitioning contrasted with thatof the other ions in showing extremely poor phloem mobility,leading to progressive preferential accumulation in leaf laminaeand negligible cycling of the element through leaves or root.Features of the response of Ricinus to salinity shown in thepresent study were discussed with data from similar modellingstudies on white lupin (Lupinus albus L.) and barley (Hordeumvulgare L.) Key words: Ricinus communis L, potassium, sodium, chloride, calcium, magnesium, phloem, xylem, transport, partitioning, salinity     

Monovalent ion stimulated adenosine triphosphatase from oat roots

Monovalent ion stimulated ATPase activity from oat (Avena sativa) roots has been found to be associated with various membrane fractions (cell wall, mitochondrial and microsomal) of oat roots. The ATPase requires Mg²⁺ (or Mn⁺²) but is further stimulated by K⁺ and other monovalent ions. The monovalent ions are ineffective in the absence of the divalent activating cation. The ATPase has been described with respect to monovalent ion specificity, temperature, pH, substrate specificity, and Mg²⁺ and K⁺ concentrations. It was further shown that oligomycin inhibits a part of the total ATPase activity and on the basis of the oligomycin sensitivity it appears that at least 2 membrane associated ATPases are being measured. The mitochondrial fraction is most sensitive to oligomycin and the microsomal fraction is least sensitive to oligomycin. The oligomycin insensitive ATPase appears to be stimulated more by K⁺ than the oligomycin sensitive ATPase.     

Characterization of Plasma Membrane-associated Adenosine Triphosphase Activity of Oat Roots

ATPase activity of plasma membranes isolated from oat (Avena sativa L. cv. Goodfield) roots was activated by divalent cations (Mg²⁺ = Mn²⁺ > Zn²⁺ > Fe²⁺ > Ca²⁺) and further stimulated by KCl and a variety of monovalent salts, both inorganic and organic. The enzyme exhibited greater specificity for cations than anions. The presence of Mg²⁺ was necessary for KCl stimulation. Ca²⁺ was ineffective in replacing Mg²⁺ for activation of plasma membrane ATPase, but it did activate other membrane-bound ATPases. The pH optima for Mg²⁺ activation and KCl stimulation of the plasma membrane ATPase were 7.5 and 6.5, respectively.     

Isolation and characterization of adenosine triphosphatase from Salicornia pacifica var. utahensis

Membrane-bound ATPase associated with plasma membrane and solubleATPase associated with the cytoplasm were prepared from shootsof Salicornia pacifica var. utahensis by sucrose density gradientcentrifugation. The isolated ATPases were tolerant to high concentrationsof NaCl. The Km for membrane-bound ATPase was 1.75 mM and forsoluble ATPase, it was 1.4 mM. The relative effectiveness ofdivalent cations for stimulation of membrane-bound ATPase wasMg²⁺>Fe²⁺>Mn²⁺>Co²⁺>Cu²⁺. Soluble ATPase activitywas stimulated by Ba²⁺>Ca²⁺>Mg²⁺ and was inhibited byCu²⁺, Zn²⁺, Co²⁺ and Fe²⁺. The compounds N,N¹-dicyclohexylcarbodiimide,NaF and ADP, did inhibit the ATPases but ouabain, triphenyltinhydroxide, sodium azide, indoleacetic acid and abscisic aciddid not inhibit the ATPases from s. pacifica var. utahensis. ¹ Present address: Department of Biology, Kong-Ju National College,Kong-Ju, Korea. (Received April 1, 1980; )     

Modulation of Ca2+-gated cardiac muscle Ca2+-release channel (ryanodine receptor) by mono- and divalent ions

The effects of mono- and divalent ions onCa²⁺-gated cardiac muscleCa²⁺-release channel (ryanodinereceptor) activity were examined in [³H]ryanodine-bindingmeasurements. Ca²⁺ bound with thehighest apparent affinity to Ca²⁺activation sites in choline chloride medium, followed by KCl, CsCl,NaCl, and LiCl media. The apparentCa²⁺ binding affinities ofCa²⁺ inactivation sites were lowerin choline chloride and CsCl media than in LiCl, NaCl, and KCl media.Sr²⁺ activated the ryanodinereceptor with a lower efficacy thanCa²⁺. Competition studiesindicated that Li⁺,K⁺,Mg²⁺, andBa²⁺ compete withCa²⁺ forCa²⁺ activation sites. In 0.125 MKCl medium, the Ca²⁺ dependence of[³H]ryanodine bindingwas modified by 5 mM Mg²⁺ and 5 mM,-methyleneadenosine 5'-triphosphate (a nonhydrolyzable ATPanalog). The addition of 5 mM glutathione was without appreciable effect. Substitution of Clby 2-(N-morpholino)ethanesulfonic acid ion caused anincrease in the apparent Ca²⁺affinity of the Ca²⁺ inactivationsites, whereas an increase in KCl concentration had the oppositeeffect. These results suggest that cardiac muscle ryanodine receptoractivity may be regulated by 1)competitive binding of mono- and divalent cations toCa²⁺ activation sites,2) binding of monovalent cations toCa²⁺ inactivation sites, and3) binding of anions to anionregulatory sites.

     

Effect of Sudden Salt Stress on Ion Fluxes in Intact Wheat Suspension Cells

Although salinity is one of the major problems limiting agriculturalproduction around the world, the underlying mechanisms of highNaCl perception and tolerance are still poorly understood. Theeffects of different bathing solutions and fusicoccin (FC),a known activator of plasma membrane ATPase, on plasma membranepotential (E_m) and net fluxes of Na⁺, K⁺and H⁺were studied inwheat suspension cells (Triticum aestivum) in response to differentNaCl treatments. E_mof cells in Murashige and Skoog (MS) mediumwas less negative than in cells exposed to a medium containing10 mM KCl + 0.1 m M CaCl₂(KSM) and to a basic salt medium (BSM),containing 1 m M KCl and 0.1 m M CaCl₂. Multiphasic Na⁺accumulationin cells was observed, peaking at 13 min after addition of 120m M NaCl to MS medium. This time scale was in good agreementwith net Na⁺flux changes measured non-invasively by moving ion-selectivemicroelectrodes (the MIFE system). When 120 m M NaCl was addedto all media studied, a quick rise of Na⁺influx was reversedwithin the first 20 min. In both 120 and 20 m M NaCl treatmentsin MS medium, net Na⁺efflux was observed, indicating that activeNa⁺transporters function in the plant cell response to saltstress. Lower external K⁺concentrations (KSM and BSM) and FCpre-treatment caused shifts in Na⁺fluxes towards net influxat 120 m M NaCl stress. Copyright 2000 Annals of Botany Company Sodium, potassium, proton, membrane potential, fusicoccin, salt stress, wheat, Triticum aestivum     

Effects of Sodium Chloride Stress on Callus Cultures of Cicer arietinum L. cv. BG-203: Growth and Ion Accumulation

Growth and ion accumulation were measured in callus culturesof Cicer arietinum L. cv. BG-203, grown on media supplementedwith 0–200 mol m^–3 NaCl. Fresh and dry weights decreasedat concentrations ranging from 100–200 mol m^–3,the reduction being greater during the third and fourth weeksof culture. Slight stimulation of growth was observed at 25and 50 mol m^–3 NaCl. There was also a decrease in tissuewater content (fresh weight: dry weight) at 100–200 molm^–3 NaCl. The concentration of Na⁺ and Cl^– in thetissue increased with increasing salinity of the medium. Mostof the accumulation of these ions occurred by the first weekwhile significant growth inhibition became apparent by onlythe third week of culture. Tissue K⁺ and Mg²⁺ decreased withincreasing salinization, the decrease being greater in K⁺ levels.Levels of Ca²⁺, however, were maintained throughout the experimentalrange. Key words: Cicer arietinum, NaCl stress, Callus cultures, Ion accumulation     

NaCl Effects on Root Plasma Membrane ATPase of Salt Tolerant Wheat

Wheat seedlings of a salt tolerant cultivar were grown hydroponically in presence and absence of 100 mM NaCl. Roots were harvested, and the plasma membrane (PM) fraction was purified. PM ATPase required a divalent cations for activity (Mg > Mn > Ca > Co > Zn > Ni > Cu), and it was further stimulated by monovalent cations (K > Rb > NH₄ > Li > choline > Cs). The pH optima were 6.0 and 5.6 in absence and presence of 25 mM KCl, respectively. The enzyme was sensitive to vanadate and DCCD but insensitive to azide, oligomycine and nitrate. The enzyme displayed a high preference for ATP but was also able to hydrolyze other nucleotide tri- and diphosphates. The enzyme activity showed a simple Michaelis-Menten kinetics for the substrate Mg²⁺-ATP in both control and salt exposed roots. The polypeptide patterns of control and salt stressed PM fractions, detected by SDS-PAGE, were very similar. NaCl substantially reduced the PM ATPase specific activity, whereas it had little effect on the apparent K_m for Mg²⁺-ATP. Since the root PM ATPase of salt sensitive and resistant genotypes responded similarly to salinity stress, it seems unlikely that the mechanism of salt tolerance in wheat is primarily based on differences in PM ATPase characteristics.     

Effect of Divalent Cations on Na+ Permeability of Chara corallina and Freshwater Grown Chara buckellii

The effect of elevated Na⁺ concentration on Na⁺ permeability(P_Na) and Na⁺ influx in the presence of two levels of externaldivalent cations was determined in Chara corallina and freshwater-culturedChara buckellii. When Na⁺ in the medium was increased from 1.0to 70 mol m^–3, Na⁺ influx increased in both species ifCa²⁺ was low (0.1 mol m–3). If Ca²⁺ was increased to 7.0mol m^–3 when Na⁺ was increased, Na⁺ influx remained atthe low control level in C. corallina, and showed only a temporaryincrease in C. buckellii. Mg²⁺ was a better substitute for Ca²⁺in C. buckellii than in C. corallina. Na⁺ permeability data suggest that when the external Ca²⁺ concentrationis low, P_Na does not increase in the presence of elevated NaCl;the increase in Na⁺ influx appears to be due to the increasein external Na⁺ concentration alone. Ca^{2 +} supplementation appearsto decrease P_Na whereas supplemental Mg²⁺ has no effect. Na⁺ effluxes were computed from previously determined net fluxesand the influxes. It was found that for both species, fluxesin both directions were stimulated in response to all experimentaltreatments, but Na⁺ influx always exceeded efflux. This resultedin net Na⁺ accumulation in the vacuoles of both species. The results are discussed with reference to net flux and electrophysiologicaldata obtained previously under identical conditions, as wellas the comparative salinity tolerance of both species and theNa⁺/divalent cation ratio. Key words: Na⁺ influx, Na⁺ tolerance, membrane potential, permeability, Chara     

Divalent cation and anion currents are activated during the dark-induced transient hyperpolarization of the plasma membrane of the green alga Eremosphaera viridis

Darkening after illumination induces a transient hyperpolarizationof the plasma membrane of the unicellular green alga Eremosphaeraviridis de Bary. With electro-physiological methods, in particularthe two electrode voltage-clamp technique, we investigated theion fluxes involved in this transient potential change (TP).The question was whether other ion currents besides those carriedby the known Ca²⁺-dependent K⁺ channel take part in this actionpotential-like, but hyperpolarizing, response. At maximum hyperpolarizationvoltage-clamp measurements resulted in ‘N-shaped’I/V curves, known from other botanical systems. The differentinstantaneous current components of the N-shaped I/V curvesoccurred at different times during a single transient potentialchange (TP). Substitution of alkali metal cations in the bathingsolution by NMG/NO₃ showed that the inward currents in the I/Vcurves were not carried by an influx of K⁺ into the cytoplasm.The voltage amplitude of the TP not only depended on the externalK⁺ concentration, but also on the Mg²⁺ concentration in thebathing solution. Increasing Mg²⁺ concentrations shifted themembrane potential in the top of the TP in the direction ofthe Nernst potential of Mg²⁺ and resulted in an increased inwardcurrent component of the N-shaped I/V curves. Another currentcomponent was found to be carried not by cations but by an effluxof anions. It was a voltage-dependent component with a maximumcurrent amplitude at voltages of about –220 to –240mV, and was blocked by the anion channel inhibitors anthracen-9-carboxylicacid (A9C), (5-nitro-2-3-phenylpropylamino) benzoic acid (NPPB)and ZnCI2. Based on these data a model is proposed which explainsthe N-shape of the I/V curves observed during the transientpotential change of the alga E. viridis by the combination ofan inward cation current with an inward anion current and theoutward cation current carried by the Ca²⁺-dependent K⁺ channels. Key words: Anion current, cation current, Eremosphaera viridis, potassium channel, voltage-clamp     

The Role of the Stem in Phloem Loading of Minerals in Lupinus albus L. cv. Ultra

Collections of phloem sap made over a 40-day period from a varietyof locations on nodulated white lupin plants (Lupinus albusL. cv. ultra) showed considerable enrichment with K⁺ and Mg²⁺in the phloem streams destined for the shoot apices or fruitsrelative to the streams arising from the leaflets (up to 5.5times). Sodium showed enrichment in the streams destined forthe roots (up to 2.5 times) but only when present in the watersupply at a high level (3 mM). The stem, in view of its centrallocation in the transport pathway, is seen as an organ capableof redistributing minerals in the phloem independently of photosynthate. Lupinus albus L., lupin, phloem loading, magnesium, potassium, sodium, mineral elements     

The concentration of selected cations in normal and crown-gall tumors obtained from potato discs

Crown-gall tumor tissue obtained from potato discs inoculatedwith virulent strains of Agrobacterium tumefadens containedhigher concentrations of K⁺, Mg²⁺ and Ca²⁺ than the correspondingnormal tissue. These tumors also contained higher concentrationsof these cations than normal tissue inoculated with an avirulentstrain of Agrobacterium tumefadens, or than normal tissue adjacentto the crown-gall tumors on the same potato disc. The concentrationof these cations remained significantly higher than controltissue regardless of the tumor age. 2,4-Dinitrophenol and myo-inositol,while affecting the concentration of Ca²⁺ in these tissues,had no effect on the Mg²⁺ and K⁺ concentrations. These resultssuggest increased concentrations of certain cations may be aspecific property of crown-gall tumors. (Received August 16, 1978; )     

Ion Effects on the H+-Translocating Adenosine Triphosphatase and Pyrophosphatase Associated with the Tonoplast of Chora corallina

H⁺-translocating ATPase and pyrophosphatase (PPase) associatedwith the tonoplast of Chara corallina were isolated with theaid of a perfusion technique, and the effects of ions on theiractivities were studied. All the alkali metal cations testedstimulated the ATPase and ATPdependent H⁺ pumping activitiesonly by 10 to 40%. Anions, on the other hand, strongly affectedthe activities. Potassium salts of Cl^- and Br^- stimulated them,while F^- and NO₃^- inhibited them. By contrast, the H⁺-translocatingPPase was insensitive to anions but sensitive to cations. Theorder of cation stimulation was Rb⁺=K⁺>Cs⁺>Na⁺=Li⁺>choline⁺.NO₃^- (50 mil), thought to be a specific inhibitor of the tonoplast-typeH⁺-ATPase, inhibited the ATPdependent H⁺ pumping almost completelybut the ATPase activity by only about 50%. Na⁺ inhibited thePP₁-dependent H⁺ pumping (I_5O=5OmM) in the presence of 50 mMKCl but not the ATP-dependent one. The PPase was more sensitiveto F^- (I₅₀=400µM) than the ATPase. Both the H⁺-ATPaseand the H⁺-PPase required Mg²⁺ for their activities, althoughan excess was inhibitory to both. The different sensitivitiesof the PP₁-dependent and the ATP-dependent H⁺- pumping enzymesto ions correspond to the tonoplast enzymes of higher plantsand may be used as "markers" to distinguish between these enzymesin characean cells (Received October 2, 1987; Accepted May 18, 1988)