The function of tonoplast ATPase in intact vacuoles of red beet is governed by direct and indirect ion effects

The pH gradient and the electric potential across the tonoplast in mechanically isolated beetroot vacuoles has been studied by following the uptake of [¹⁴C]methylamine and [¹⁴C]triphenyl-methylphosphoniumchloride. In response to Mg-ATP, the vacuolar interior is acidified by 0.8 units. This strong acidification is accompanied by a slight hyperpolarization of the membrane potential, which is probably caused by a proton diffusion potential. In preparations where only a small acidification (0.4 units) occurred, the membrane potential was depolarized by the addition of Mg-ATP. Different monovalent cations and anions were tested concerning their effect on the pH gradient and ATPase activity in proton-conducting tonoplasts. Chloride stimulation and NO ₃ ^- inhibition were clearly present. The observed decline of the pH gradient upon the addition of Na⁺ salts is probably caused by an Na⁺/H⁺ antiport system.Abbreviations and symbol CCCP carbonylcyanide-m-chlorophenylhydrazone - Mes 2(N-morpholino)ethanesulfonic acid - TPMP⁺ triphenylmethylphosphoniumchloride - Tris 2-amino-2-(hydroxymethyl)-1,3-propanediol - membrane potential Dedicated to Professor A. Betz on the occasion of his 65th birthday     

Effects of abscisic acid on sequestration and exchange of Na+ by barley roots

Excised Na⁺-starved barley roots were suspended in solutions of Na⁺ in combination with NO ₃ ^- , Cl^-, and SO ₄ ^2- , and effects of the added phytohormone, abscisic acid (ABA), to the medium were determined. Abscisic acid increased the rate of Na⁺ (²²Na⁺) accumulation and the amount of Na⁺ deposited in the vacuoles. These stimulating effects of ABA were modified by anions following the sequence NO ₃ ^- >Cl^->SO ₄ ^2- . Testing whether the magnitude of the pH gradient across the plasmalemma of the cells of the root cortex affects rates of Na⁺ accumulation and their dependence upon ABA, we observed that, in the pH range from 4 to 8, the ABA-induced stimulation was strongest at pH 5.8, and least at pH 4. Changes in pH during the experiment caused changes in the rates of Na⁺ accumulation in agreement with experiments performed at constant pH values. Simultaneously with ABA-enhanced accumulation, loss of Na⁺ occurred. Loss of Na⁺ was strongest at pH 4 and was affected by anions, being greatest with SO ₄ ^2- and following the sequence SO ₄ ^2- >Cl^->NO ₃ ^- . On the basis of the finding that initial acceleration of uptake as well as loss of Na⁺ depended on the pH of the medium we suggest that, in barley roots, ABA stimulates an exchange of Na⁺ for H⁺ at the plasmalemma of the cortical cells. The results indicate that ABA-stimulated expulsion of Na⁺, in combination with ABA-stimulated sequestration in the vacuoles, constitutes one of the mechanisms which enable barley plants to tolerate higher than normal levels of Na⁺.Abbreviations ABA abscisic acid - FW fresh weight     

Characterization of esophageal desalination in the seawater eel,Anguilla japonica

To characterize mechanisms of esophageal desalination, osmotic water permeability and ion fluxes were measured in the isolated esophagus of the seawater eel. The osmotic permeability coefficient in the seawater eel esophagus was 2·10^-4 cm·s^-1. This value was much lower than those in tight epithelial, although the eel esophagus is a leaky epithelium with a tissue resistance of 77 ohm·cm^-2. When the esophagus was bathed in normal Ringer solutions on both sides no net ion and water fluxes were observed. However, when mucosal NaCl concentration was increased by a factor of 3, Na⁺ und Cl^- ions were transferred from mucosa to serosa (desalination). If only Na⁺ or Cl^- concentration in the mucosal fluid was increased by a factor of 3, net Na⁺ and Cl^- fluxes were reduced to 30–40%, indicating that 60–70% of the net Na⁺ and Cl^- fluxes are coupled mutually. The coupled NaCl transport seems to be effective in desalting the luminal high NaCl. The remaining 30–40% of the total Na⁺ and Cl^- fluxes seems to be due to a simple diffusion, because these components are independent of each other and follow their electrochemical gradients, and also because these fluxes remain even after treatment with NaCN or ouabain. A half of the coupled NaCl transport could be explained by a Na⁺/H⁺–Cl^-/HCO ₃ ^- double exchanger on the apical membrane of the esophageal epithelium, because mucosal amiloride and 4.4-diisothiocyanatostilbene-2,2-disulphonic acid inhibited the net Na⁺ and Cl^- fluxes by approximately 30%. The other half of the coupled NaCl transport, which follows their electrochemical gradients, still remains to be explained.Abbreviations DIDS 4,4-diisothiocyanatostilbene-2,2-disulphonic acid - NMDG N-methyl-d-glucosamine - P _Cl Cl^- permeability coefficient - PD transepithelial potential difference - P _Na Na⁺ permeability coefficient - P _osm osinotic permeability coefficient - TALH thick ascending limb of Henle's loop     

Properties of Plasmamembrane ATPase and Mitochondrial ATPase of Saccharomyces cerevisiae

The properties of mitochondrial ATPase and plasmamembrane ATPase of Saccharomyces cerevisiae are compared. The pH dependence differs considerably. At low pH plasmamembrane ATPase is inactivated. High salt concentrations protect the ATPase against acid inactivation. K⁺ is more effective than Na⁺. The sensitivity of mitochondrial ATPase towards azide, Dio 9 and oligomycin is far greater than found with plasmamembrane ATPase. There are no indications that the membrane ATPase is involved directly either in monovalent cation uptake or in divalent cation uptake. Sr²⁺ and Ca²⁺ do not activate plasmamembrane ATPase and inhibit Mg²⁺-activated ATPase. The substrate specificity of plasmamembrane ATPase is much greater than the substrate specificity of mitochondrial ATPase.     

Differential Ion Stimulation of Plasmalemma Adenosine Triphosphatase from Leaf Epidermis and Mesophyll of Nicotiana rustica L

An ATPase preparation, presumed to be associated with plasma membrane due to the coincidence on isopycnic gradients of cellulase and β-glucan synthetase at high substrate, has been isolated from the epidermal and mesophyll of tobacco leaf. The ATPase from both tissues was found to prefer ATP over other nucleotides. The pH optimum was 7.0 in the presence of 3 millimolar MgCl₂ and pH 6.5 in the presence of 3 millimolar MgCl₂ and 100 millimolar KCl. Monovalent ion stimulation patterns of the ATPases from these tissues were found to differ and ion accumulation patterns in these tissues reflect this difference: mesophyll accumulated roughly equal amounts of Na⁺ and Rb⁺ and its plasma membrane ATPase is also equally stimulated by these ions; on the other hand, epidermal ATPase preparations showed a stronger stimulation by Rb⁺ than Na⁺ and this tissue was found to accumulate Rb⁺ in preference to Na⁺. Abscisic acid and fusicoccin affected both ATPase activity and ion uptake, the former inhibiting and the latter stimulating these parameters. These data support the hypothesis that the epidermal plasmalemma ATPase is involved in stomatal opening.     

Is there a chloride ATPase in the gills of the shore crab Carcinus maenas?

Summary In gills of the shore crab Carcinus maenas an ATPase activity was found which was stimulated by bicarbonate and inhibited by low concentration of oligomycin and thiocyanate. This ATPase was activated by small hydrated alkali cations, i.e., activation was absent in the presence of Li⁺, small in the presence of Na⁺, and highest in the presence of K⁺ (K _m=4 mM). Inhibitor studies using ouabain, NEM, and vanadate suggest that this ATPase is different from (Na⁺+K⁺)-ATPase, the H⁺-ATPase of organelles, or an E ₁ E ₂-type ATPase represented by the H⁺/K⁺-ATPase in gastric mucosa. Results obtained by differential and density gradient centrifugation indicate that this ATPase is located in crab gill mitochondria, a location ruling out its direct participation in transepithelial ion transport. Since the ATPase lacked specific Cl^--activation it is not considered to be a Cl^- pump but a mitochondrial F ₁ F ₀-ATPase. Specific activities of mitochondrial ATPase and (Na⁺+K⁺)-ATPase were of comparable magnitude. Both ATPases were greatly increased in gills of crabs acclimated to brackish water (salinity 10) compared to crabs maintained in sea water (30). These results imply that low salinity-induced modifications in branchial tissues include mechanisms for active ion uptake as well as the elements for provision of cellular energy.Abbreviations ATPase adenosine triphosphatase - HEPES N-(2-hydroxyethyl)-1-piperazine-N(2-ethanesulfonic acid) - LDH lactate dehydrogenase - NADH reduced nicotinamide adenine dinucleotide - NEM Niethylmaleimide - PEP phosphoenolpyruvate - PK pyruvate kinase - TRIS TRIS (hydroxymethyl)aminomethane - S salinity     

Relationship between Sodium Influx and Salt Tolerance of Nitrogen-Fixing Cyanobacteria

The relationship between sodium uptake and cyanobacterial salt (NaCl) tolerance has been examined in two filamentous, heterocystous, nitrogen-fixing species of Anabaena. During diazotrophic growth at neutral pH of the growth medium, Anabaena sp. strain L-31, a freshwater strain, showed threefold higher uptake of Na⁺ than Anabaena torulosa, a brackish-water strain, and was considerably less salt tolerant (50% lethal dose of NaCl, 55 mM) than the latter (50% lethal dose of NaCl, 170 mM). Alkaline pH or excess K⁺ (>25 mM) in the medium causes membrane depolarization and inhibits Na⁺ influx in both cyanobacteria (S. K. Apte and J. Thomas, Eur. J. Biochem. 154:395-401, 1986). The presence of nitrate or ammonium in the medium caused inhibition of Na⁺ influx accompanied by membrane depolarization. These experimental manipulations affecting Na⁺ uptake demonstrated a good negative correlation between Na⁺ influx and salt tolerance. All treatments which inhibited Na⁺ influx (such as alkaline pH, K⁺ above 25 mM, NO₃⁻, and NH₄⁺), enhanced salt tolerance of not only the brackish-water but also the freshwater cyanobacterium. The results indicate that curtailment of Na⁺ influx, whether inherent or effected by certain environmental factors (e.g., combined nitrogen, alkaline pH), is a major mechanism of salt tolerance in cyanobacteria.     

黑河天涝池流域典型林分生态水文化学特征

采集了黑河天涝池流域典型林分林外雨、穿透雨、树干径流和枯透水,并检测水体pH值和12种离子(K~+、Ca~(2+)、Na~+、Mg~(2+)、NH_4~+、Cu~(2+)、Zn~(2+)、Pb~(2+)、Cd~(2+)、Cl~-、SO_4~(2-)、NO_3~3)的质量浓度。结果表明:天涝池流域大气降水pH均值为7.74,呈碱性,降水中离子绝对质量浓度较低,最高的是NO_3~-,质量浓度为1.1111 mg/L,最低的为Na~+,质量浓度为0.0108 mg/L;两种林分冠层有降低降雨pH值的作用,青海云杉林冠层对NH_4~+有升高作用,祁连圆柏林冠层对NH_4~+有降低作用,两种林冠层对NO_3~-和Cu~(2+)质量浓度有降低作用,对其它离子质量浓度均表现为升高作用;两种林分树干径流有提高穿透雨pH值的作用,与穿透雨相比,两种林分树干径流中阴离子均有升高,圆柏树干径流中所有阳离子质量浓度均有下降,云杉树干径流中Ca~(2+)、K~+、Mg~(2+)和Na~+减少,NH_4~+和Cu~(2+)增加;典型林分枯透水有提升穿透雨pH值的作用,与穿透雨相比,两种林分枯透水中阴离子质量浓度均有升高,云杉枯透水各阳离子均有降低,圆柏枯透水中Ca~(2+)、K~+和Mg~(2+)质量浓度升高,NH_4~+、Na~+和Cu~(2+)质量浓度下降;在采集的所有样本中,Pb~(2+)和Cd~(2+)均未检出,而Zn~(2+)仅在云杉树干径流中检出。     

Sodium gradient- and sodium plus potassium gradient-dependentl-glutamate uptake in renal basolateral membrane vesicles

Summary A membrane preparation enriched in the basolateral segment of the plasma membrane was isolated from the rat renal cortex by a procedure that included separation of particulates on a self-generating Percoll gradient. The uptake ofl-glutamate by the basolateral membrane vesicles was studied. A Na⁺ gradient ([Na⁺] _o >[Na⁺] _i ) stimulated the uptake ofl-glutamate and provided the driving force for the uphill transport of the acidic amino acid, suggesting a Na⁺-l-glutamate cotransport system in the basolateral membrane. A K⁺ gradient ([K⁺] _i >[K⁺] _o ) increased the uptake additionally. This effect was specific for K⁺ (Rb⁺). The action of the K⁺ gradient in enhancing the uptake ofl-glutamate had an absolute requirement for Na⁺. In the presence of Na⁺, but in the absence of a Na⁺ gradient. i.e., [Na⁺] _o =[Na⁺] _i , the K⁺ gradient also energized the concentrative uptake ofl-glutamate. This effect of the K⁺ gradient was not attributable to an alteration in membrane potential. The finding of a concentrative uptake system forl-glutamate energized by both Na⁺ ([Na⁺] _o >[Na⁺] _i and K⁺ ([K⁺] _i >[K⁺] _o ) gradients in the basolateral membrane, combined with previous reports of an ion gradient-dependent uphill transport system for this amino acid in the brush border membrane, suggests a mechanism by whichl-glutamate is accumulated intracellularly in the renal proximal tubule to extraordinarily high concentrations.     

Characterisation of a salt-stimulated ATPase activity associated with vacuoles isolated from storage roots of red beet (Beta vulgaris L.)

Ion stimulation and some other properties of an ATPase activity associated with vacuoles isolated from storage roots of red beet (Beta vulgaris L.) have been determined. The ATPase had a specific requirement for Mg²⁺ and in the presence of Mg²⁺ it was stimulated by salts of monovalent cations. The degree of stimulation by monovalent salts was influenced mainly by the anion and the order of effectiveness of the anions tested was Cl^->HCO ₃ ^- >Br^->malate>acetate>SO ₄ ^2- . For any given series of anions the magnitude of the stimulation obtained was influenced by the accompanying cation (NH ₄ ⁺ Na⁺>K⁺). This cation effect was abolished by 0.01% (v/v) Triton X-100 and it is suggested that it is the result of different permeabilities of membrane vesicles to the cations. There was no evidence of synergistic stimulation of the ATPase by mixtures of Na⁺ and K⁺. KCl- and NaCl-stimulation was maximal with salt concentrations in the range 60–150 mM. The true substrate of the enzyme was shown to be MgATP. It was shown that KCl stimulation was the result of an increase in V_max rather than a change in the affinity of the enzyme for MgATP. The ATPase was inhibited by N,N-dicyclohexylcarbodiimide, diethylstilbestrol, mersalyl and KNO₃ but other inhibitors tested (azide, oligomycin, orthovanadate, K₃[Cr(oxalate)₆] and ethyl-3-[3-dimethylaminopropyl]carbodiimide) were without effect or caused only partial inhibition at the highest concentration tested. The ATPase activity was equally distributed between pellet and supernatant fractions obtained after the subfractionation of vacuoles but the properties of the ATPase in each fraction were the same. It is suggested that beet vacuoles possess only one ATPase. The properties of the ATPase are compared with those of ATPases associated with other plant membranes and organelles and its possible role in transport at the tonoplast is discussed.Abbreviations ATP_F free ATP - ATP_T total ATP - BSA bovine serum albumen - DCCD N,N-dicyclohexylcarbodiimide - DES diethylstilbestrol - DNP 2,4-dinitrophenol - EDAC ethyl-3-(3-dimethylaminopropyl)carbodiimide - K_m apparent Michaelis constant - MgATP complex of Mg²⁺ and ATP - Mg _F ²⁺ free Mg²⁺ - Mg _T ² total Mg²⁺ - MES 2-(N-Morpholino)ethanesulphonic acid - Na₂EDTA disodium ethylenediaminetetraacetic acid - NEM N-ethylmaleimide - P_i inorganic phosphate - TCA trichloroacetic acid - Tris tris(hydroxymethyl)methylamine - V_max maximum velocity     

Growth,chemical composition,and nitrate reductase activity of Rumex species in relation to form and level of N supply

Growth, chemical composition, and nitrate reductase activity (NRA) of hydroponically cultured Rumex crispus, R. palustris, R. acetosa, and R. maritimus were studied in relation to form (NH₄ ⁺, NO₃ ^-, or both) and level of N supply (4 mM N, and zero-N following a period of 4mM N). A distinct preference for either NH₄ ⁺ or NO₃ ^- could not be established. All species were characterized by a very efficient uptake and utilization of N, irrespective of N source, as evident from high concentrations of organic N in the tissues and concurrent excessive accumulations of free NO₃ ^- and free NH₄ ⁺. Especially the accumulation of free NH₄ ⁺ was unusually large. Generally, relative growth rate (RGR) was highest with a combination of NH₄ ⁺ and NO₃ ^-. Compared to mixed N supply, RGR of NO₃ ^-- and NH₄ ⁺-grown plants declined on average 3% and 9%, respectively. Lowest RGR with NH₄ ⁺ supply probably resulted from direct or indirect toxicity effects associated with high NH₄ ⁺ and/or low Ca²⁺ contents of tissues. NRA in NO₃ ^- and NH₄NO₃ plants was very similar with maxima in the leaves of ca 40 μmol NO₂ ^- g^-1 DW h^-1. ‘Basal’ NRA levels in shoot tissues of NH₄ ⁺ plants appeared relatively high with maxima in the leaves of ca 20 μmol NO₂ ^- g^-1 DW h^-1. Carboxylate to organic N ratios, (C-A)/N_org, on a whole plant basis varied from 0.2 in NH₄ ⁺ plants to 0.9 in NO₃ ^- plants. After withdrawal of N, all accumulated NO₃ ^- and NH₄ ⁺ was assimilated into organic N and the organic N redistributed on a large scale. NRA rapidly declined to similar low levels, irrespective of previous N source. Shoot/root ratios of -N plants were 50–80% lower than those from +N plants. In comparison with +N, RGR of -N plants did not decline to a large extent, decreasing by only 15% in -NH₄ ⁺ plants due to very high initial organic-N contents. N-deprived plants all exhibited an excess cation over anion uptake (net proton efflux), and whole-plant (C-A)/N_org ratios increased to values around unity. Possible difficulties in interpreting the (C-A)/N_org ratio and NRA of plants in their natural habitats are briefly discussed.     

Effects of salinity on uptake and distribution of Na+, Cl- and K+ in two wheat cultivars

Plants of two wheat (Triticum aestivum L.) cultivars differing in salt tolerance were grown in sand with nutrient solutions. 35-d-old plants were subjected to 5 levels of salinity created by adding NaCl, CaCl₂ and Na₂SO₄. Growth reduction caused by salinity was accompanied by increased Na⁺ and Cl^- concentrations, Na⁺/K⁺ ratio, and decreased concentration of K⁺. The salt tolerant cv. Kharchia 65 showed better ionic regulation. Salinity up to 15.7 dS m^-1 induced increased uptake of Na⁺ and Cl^- but higher levels of salinity were not accompanied by further increase in uptake of these ions. Observed increases in Na⁺ and Cl^- concentrations at higher salinities seemed to be the consequence of reduction in growth. Uptake of K⁺ was decreased; more in salt sensitive cultivar. This was also accompanied by differences in its distribution.     

Bioenergetics of alkalophilic bacteria

Summary The central problem for organisms which grow optimally, and in some cases obligately, at pH values of 10 to 11, is the maintenance of a relatively acidified cytoplasm. A key component of the pH homeostatic mechanism is an electrogenic Na⁺/H⁺ antiporter which—by virtue of kinetic properties and/or its concentration in the membrane—catalyzes net proton uptake while the organisms extrude protons during respiration. The antiporter is also capable of maintaining a constant pH_in during profound elevations in pH_out as long as Na⁺ entry is facilitated by the presence of solutes which are taken up with Na⁺. Secondary to the problem of acidifying the interior is the adverse effect of the large pH gradient, acid in, on the total pmf of alkalophile cells. For the purposes of solute uptake and motility, the organisms appear to largely bypass the problem of a low pmf by utilizing a sodium motive force for energization. However, ATP synthesis appears not to resolve the energetics problem by using Na⁺ or by incorporating the proton-translocating ATPase into intracellular organelles. The current data suggest that effective proton pumping carried out by the alkalophile respiratory chain at high pH may deliver at least some portion of the protons to the proton-utilizing catalysts, i. e., theF ₁ F ₀-ATPase and the Na⁺/H⁺ antiporter, by some localized pathway.     

Sodium-potassium exchange in sea urchin egg. I. Kinetic and biochemical characterization at fertilization

Biochemical and kinetic characteristics of the Na⁺-K⁺ exchange were studied in Paracentrotus lividus eggs. Measurement of the ⁸⁶Rb uptake shows that ouabain-sensitive ⁸⁶Rb uptake is dramatically stimulated within the first minute following fertilization. The Na⁺-K⁺ pump-mediated K⁺ entry presents a maximal rate at 8 min postfertilization and then decreases to reach a plateau within 30 min. We assess that the steep rise in cell K⁺ occurring at fertilization (J.P. Girard, P. Payan, C. Sardet, Exp. Cell. Res. 142:215–221, 1982) does not originate from a net entry of external K⁺. Measured 30 min postfertilization, the half-maximal activation by K⁺ of the ouabain-sensitive Na⁺-K⁺ exchange is 5–6 mM and the ouabain lC₅₀ is 5.10^?5 M. Egg cortices from unfertilized and fertilized eggs show comparable Na⁺-K⁺ ATPase activity with a 50% ouabain-sensitive fraction. V_m and K_m for Na⁺ and K⁺ of the enzyme are of the same order of magnitude in cortices of unfertilized and fertilized eggs. Cortical Na⁺-K⁺ ATPase from unfertilized eggs shows a ten fold increase of activity between pH 6.7 and pH 7.7. The results strongly suggest that the plasma membrane of unfertilized eggs contains a preexisting Na⁺-K⁺ transporting system which is obligatorily stimulated at fertilization.     

Anion-sensitive ATPase activity and proton transport in isolated vacuoles of species of the CAM genus Kalanchoë

Vacuoles were isolated from leaves of Kalanchoë daigremontiana Hamet et Perrier de la Bathie, and the ionic sensitivity of the vacuolar ATPase was studied in vacuole homogenates desalted on Sephadex G-25. The ATPase activity was dependent on the presence of divalent cations (Mg²⁺≥ Mn²⁺≥ Ca²⁺, Co²⁺; Zn²⁺ had no effect). Mg²⁺-dependent ATPase activity was stimulated by anions (Cl^? > malate²⁺, HCO^?₃), with maximal stimulation at concentrations above 50 mM. Mg²⁺-Dependent activity was inhibited by NO^?₃ above 2 mM, but no saturation was observed up to 100 mM. No stimulation by K⁺ or Na⁺ was detected; stimulation by NH⁺₄ was abolished by 0.01% (w/v) Triton X-100, suggesting that the NH⁺₄ effect was due to the permeability of vacuolar membrane vesicles to NH₃. Trans-tonoplast electrical potentials (Δψ) and intra-vacuolar pH were measured with glass microelectrodes and antimony covered glass micro-pH-electrodes, respectively. Free vacuofes isolated from Kalanchoë tubiflora (Harv.) Hamet were slightly positive with respect to the suspension medium. This Δψ was insensitive to the protonophore FCCP and depolarized by about 4 mV on addition of 50 mM KCl, still remaining about +5 mV. Upon addition of 7 mM Mg-ATP, vacuoles showed an FCCP-sensitive increase of Δψ from +9.2 ± 2.8 (13) to +17.8 ± 3.7 (12) mV [given as x?± sd (n)] and an internal acidification from pH 5.4 ± 0.2 (11) to pH 4.3 ± 0.4 (12). Mg-ADP and ATP without Mg²⁺ had no effect on Δψ. It is concluded that the H4 pumping at the tonoplast is due to the functioning of the anion-sensitive vacuolar ATPase and that this is an essential part of the mechanism of nocturnal acid accumulation in CAM.     

Proton-pumping adenosine triphosphatase in membrane vesicles of tobacco callus: Sensitivity to vanadate and K+

H⁺-pumping adenosinetriphosphatases (ATPases, EC 3.6.1.3) were demonstrated in sealed microsomal vesicles of tobacco callus. Quinacrine fluorescence quenching was induced specifically by MgATP and stimulated by EGTA and Cl^?. Fluorescence quenching reflected a relative measure of pH gradient formation (inside acid), as it could be reversed by gramicidin (an H⁺/cation conductor) or 10 mM NH₄Cl (an uncoupler). H⁺ pumping was inhibited by tributyltin (an ATPase inhibitor) and sodium vanadate, but it was insensitive to oligomycin or fusicoccin. The vanadate concentration required to inhibit pH gradient formation was similar to that needed to inhibit KCl-stimulated Mg²⁺-ATPase activity and generation of a membrane potential (measured by ATP-dependent ³⁵SCN^? uptake). About 45% of all three activities (ATPase, pH gradient, membrane potential generation) were vanadate-insensitive, supporting the idea that non-mitochondrial membranes of plants have at least two types of electrogenic H⁺ pump.A vanadate-insensitive, H⁺-pumping ATPase previously shown by methylamine accumulation was characterized to be anion-sensitive and possibly enriched in vacuolar membranes (Churchill, K.A. and Sze, H. (1983) Plant Physiol. 71, 610–617). Yet, pH gradient formation determined by quinacrine fluorescence quenching was decreased by monovalent cations with a sequence K⁺, Rb⁺, Na⁺ > Cs⁺,Li⁺> choline, bisTris-propane. Since K⁺ stimulated ATPase activity more than Bistris-propane, K⁺ appeared to collapse formation of the pH gradient by an H⁺/K⁺ countertransport. The sensitivity to vanadate and K⁺ provides evidence that the plasma-membrane ATPase is an electrogenic H⁺ pump.     

Responses of the macroalga Gracilaria tenuistipitata var. liui (Rhodophyta) to iron stress

Chlorophyll (Chl), phycoerythrin (PE), total nitrogen (TN% dw) and Fein tissues were measured in Fe-deficient cultures of Gracilariatenuistipitata var. liui over a period of 60 days. ⁵⁵Fe uptakeand photosynthetic carbon fixation (NaH¹⁴CO³) werecompared in Fe-rich and Fe-deficient cultures and analyzed the effects ofFe-deficiency on the ultrastructure. The maximum carbon fixationdecreased significantly (p < 0.01) under Fe-deficiency. Thechlorophyll and phycoerythrin contents also declined with decreasing tissueiron content, falling, respectively, to 7.9 and 33.8% of their originallevel. Photosynthesis in Fe-deficient cells became light-saturated at lowerirradiance than the control. Total N in tissue decreased from 3.65 to2.49%. ⁵⁵Fe uptake rate for cultures grown on NO₃ ^-was measured following resuspension in either NH₄ ⁺ orNO₃ ^- as N source. Enhanced Fe uptake developedunder Fe stress, especially with cells resuspended in NH⁺ ₄-N medium. The Vmaxfor Fe uptake was higher with NH₄ ⁺ thanNO₃ ^- (62.8 versus 12.1 pmol mg dw^-1 h^-1). The requirement for N accelerates further Fe uptake. Ultrastructuralobservations of Fe-deficient cells showed reductions in chloroplast number,degeneration of lamellar organization, decrease in mitochondrial matrixdensity and variation in accumulation body number and morphology. During Fe-deficiency, the growth rate continued to decline and after 40days of iron deficiency, no further growth was detectable, and eventuallyiron deficiency resulted in chlorosis. The results suggest that the lowergrowth rate of Gracilaria tenuistipitata var. liui underFe-deficiency may result from largely from inhibition of photosynthesis andnitrogen utilization.     

Na<Superscript>+</Superscript>-mediated piezoprotection in<Emphasis Type="Italic"> Rhodotorula rubra</Emphasis>

Sodium concentrations as low as 2 mM exerted a significant protective effect on the high-pressure inactivation (160–210 MPa) of Rhodotorula rubra at pH 6.5, but not on two other yeasts tested (Shizosaccharomyces pombe and Saccharomyces cerevisiae). A piezoprotective effect of similar magnitude was observed with Li⁺ (2 and 10 mM), and at elevated pH (8.0–9.0), but no effect was seen with K⁺, Ca²⁺, Mg²⁺, Mn²⁺, or NH₄ ⁺. Intracellular Na⁺ levels in cells exposed to low concentrations of Na⁺ or to pH 8.0–9.0 provided evidence for the involvement of a plasma membrane Na⁺/H⁺ antiporter and a correlation between intracellular Na⁺ levels and pressure resistance. The results support the hypothesis that moderate high pressure causes indirect cell death in R. rubra by inducing cytosolic acidification.Communicated by K. Horikoshi     

Sodium-dependent succinate uptake in purple bacterium Ectothiorhodospira shaposhnikovii

Succinate, malate and fumarate uptake in purple sulfur bacterium Ectothiorhodospira shaposhnikovii, strain 1 K MSU, obligatorily depends on the presence of Na⁺. Other monovalent cations such as K⁺, Li⁺, NH₄⁺ could not replace Na⁺. Experiments with energy-depleted cells have shown that succinate uptake against its concentration gradient can be energized by artificially imposed sodium gradients (ΔpNa).An artificial membrane potential (inside negative) inhibited ΔpNa-driven succinate uptake at pH 7.0 but stimulated it at pH 9.0.The results confirm the suggestion that succinate uptake in E. shaposhnikovii is carried out in symport with Na⁺.