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Nitrogen Utilization in Lemna1: II. Studies of Nitrate Uptake Using 13NO3−

Ingemarsson, Björn; Oscarson, Petter; af Ugglas, Magnus; Larsson, Carl-Magnus
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em /11/1987 EN
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465.65848%
13N-labeled nitrate was used to trace short-term nitrate influx into Lemna gibba L. G3 in experiments where disappearance of both radioactivity and total nitrate from the incubation medium was measured continuously and simultaneously. In plants performing net nitrate uptake from an initial nitrate concentration of 40 to 60 micromolar, there was no discrepancy between net uptake and influx, irrespective of the N status of the plants, indicating that concomitant nitrate efflux was low or nil. Plants treated with tungstate to inactivate nitrate reductase were able to take up nitrate following induction of the uptake system by exposure to a low amount of nitrate. Also, in this case, net uptake was equivalent to influx. In tungstate-treated plants preloaded with nitrate, both net uptake and influx were nil. In contrast to these observations, a clear discrepancy between net uptake and influx was observed when the plants were incubated at an initial nitrate concentration of approximately 5 micromolar, where net uptake is low and eventually ceases. It is concluded that plasmalemma nitrate transport is essentially unidirectional in plants performing net uptake at a concentration of 40 to 60 micromolar, and that transport is nil when internal nitrate sinks (vacuole...

Calcium Transport in Sealed Vesicles from Red Beet (Beta vulgaris L.) Storage Tissue 1: II. Characterization of 45Ca2+ Uptake into Plasma Membrane Vesicles

Giannini, John L.; Ruiz-Cristin, Jose; Briskin, Donald P.
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em /12/1987 EN
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465.65848%
Calcium uptake was examined in sealed plasma membrane vesicles isolated from red beet (Beta vulgaris L.) storage tissue using 45Ca2+. Uptake of 45Ca2+ by the vesicles was ATP-dependent and radiotracer accumulated by the vesicles could be released by the addition of the calcium ionophore A23187. The uptake was stimulated by gramicidin D but slightly inhibited by carbonylcyanide m-chlorophenylhydrazone. Although the latter result might suggest some degree of indirect coupling of 45Ca2+ uptake to ATP utilization via δμH+, no evidence for a secondary H+/Ca2+ antiport in this vesicle system could be found. Following the imposition of an acid-interior pH gradient, proton efflux from the vesicle was not enhanced by the addition of Ca2+ and an imposed pH gradient could not drive 45Ca2+ uptake. Optimal uptake of 45Ca2+ occurred broadly between pH 7.0 and 7.5 and the transport was inhibited by orthovanadate, N,N′-dicyclohexylcarbodiimide, and diethylstilbestrol but insensitive to nitrate and azide. The dependence of 45Ca2+ uptake on both calcium and Mg:ATP concentration demonstrated saturation kinetics with Km values of 6 micromolar and 0.37 millimolar, respectively. While ATP was the preferred substrate for driving 45Ca2+ uptake, GTP could drive transport at about 50% of the level observed for ATP. The results of this study demonstrate the presence of a unique primary calcium transport system associated with the plasma membrane which could drive calcium efflux from the plant cell.

Induced K+ Efflux from Cultured Rose Cells 1: Effects of Protein-Synthesis Inhibitors

Murphy, Terence M.
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em /03/1988 EN
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Inhibitors of translation, cycloheximide, emetine, and puromycin, and inhibitors of transcription, actinomycin D and cordycepin, stimulate a net efflux of K+ from cultured cells of Rosa damascena. In the case of cycloheximide and emetine, this efflux bears many similarities to the efflux induced by ultraviolet radiation, including a lag period of 0.25 to 2.5 hours and a limited total loss of K+. The efflux is transient, and continued incubation of cells with cycloheximide and emetine allows the cells to recover the K+; after this, the cells no longer release K+ in response to UV or to cycloheximide treatment. This suggests that the efflux process depends on continued protein synthesis. Other treatments such as cerulenin, low temperature (0°C), and high temperature (40°C) also inhibit the UV- and cycloheximide-induced K+ efflux, suggesting that the induction of efflux may involve the synthesis of new plasma membrane.

l-Glutamate-Dependent Medium Alkalinization by Asparagus Mesophyll Cells 1: Cotransport or Metabolism?

McCutcheon, Steve L.; Ciccarelli, Bruce W.; Chung, Induk; Shelp, Barry; Bown, Alan W.
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em /12/1988 EN
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Mechanically isolated Asparagus sprengeri Regel mesophyll cells cause alkalinization of the suspension medium on the addition of l-glutamate or its analog l-methionine-d,l-sulfoximine. Using a radiolabeled pH probe, it was found that both compounds caused internal acidification whereas l-aspartate did not. Fusicoccin stimulated H+ efflux from the cells by 111% and the uptake of l-[U-14C]glutamate by 55%. Manometric experiments demonstrated that, unlike l-methionine-d,l-sulfoximine, l-glutamate stimulated CO2 evolution from nonilluminated cells. Simultaneous measurements of medium alkalinization and 14CO2 evolution upon the addition of labeled l-glutamate showed that alkalinization was immediate and reached a maximum value after 45 minutes whereas 14CO2 evolution exhibited a lag before its appearance and continued in a linear manner for at least 100 minutes. Rates of alkalinization and uptake of l-[U-14C]glutamate were higher in the light while rates of 14CO2 evolution were higher in the dark. The major labeled product of glutamate decarboxylation, γ-aminobutyric acid, was found in the cells and the suspension medium. Its addition to the cell suspension did not result in medium alkalinization and evidence indicates that it is lost from the cell to the medium. The data suggest that the origin of medium alkalinization is co-transport not metabolism...

Characterization of the Effects of Divalent Cations on the Coupled Activities of the H+-ATPase in Tonoplast Vesicles

Tu, Shu-I; Nungesser, Edwin; Brauer, David
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em /08/1989 EN
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465.71402%
The substrate requirement of the H+-ATPase in purified corn root tonoplast vesicles was investigated. The coupled activities, ATP hydrolysis and proton pumping, were simultaneously supported only by Mg2+ or Mn2+. The presence of Ca2+ or Ba2+ did not significantly affect the coupled activities. The addition of Cd2+, Co2+, Cu2+, and Zn2+ inhibited both the hydrolysis of Mg-ATP and the proton transport. However, the inhibition of proton pumping was more pronounced. Based on equilibrium analysis, both ATP-complexed and free forms of these cations were inhibitory. Inhibition of the hydrolysis of Mg-ATP could be correlated to the concentrations of the ATP-complex of Zn. On the other hand, the free Cu2+ and Co2+ were effective in inhibiting hydrolysis. For proton pumping, the ATP complexes of Co2+, Cu2+, and Zn2+ were effective inhibitors. However, this inhibition could be further modulated by free Co2+, Cu2+, and Zn2+. While the equilibrium concentrations of Cd-ATP and free Cd2+ were not estimated, the total concentration of this cation needed to inhibit the coupled activities of the H+-ATPase was found to be in the range of 10 to 100 micromolars. The presence of free divalent cations also affected the structure of the lipid phase in tonoplast membrane as demonstrated by the changes of emission intensity and polarization of incorporated 1...

Subunit Composition of Cytochrome c Oxidase in Mitochondria of Zea mays1

Hawkesford, Malcolm J.; Liddell, Andrew D.; Leaver, Christopher J.
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em /12/1989 EN
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465.71402%
Cytochrome c oxidase has been purified from Zea mays mitochondria by a solubilization with dodecyl maltoside followed by a simple and rapid two step fast protein liquid chromatographic method involving anion exchange on Mono Q and size exclusion chromatography on Superose 12. The preparation obtained was resolved by urea sodium dodecyl sulfate-polyacrylamide gel electrophoresis and had a subunit composition comprising polypeptides of apparent molecular masses of 48, 31, and 25 kilodaltons at least one at 16 and 11 kilodaltons and three subunits below 10 kilodaltons. Comparison with a purified yeast cytochrome c oxidase revealed that the four largest subunits showed similar electrophoretic mobilities. Subunits I and II cross-reacted with antibodies raised against the yeast homologous polypeptides. Polypeptides of the plant ubiquinone:cytochrome c reductase complex have also been identified by cross-reaction with antibodies raised against yeast cytochrome b and c1 subunits and by inference from comigration.

ΔpH-Dependent Amino Acid Transport into Plasma Membrane Vesicles Isolated from Sugar Beet Leaves: I. Evidence for Carrier-Mediated, Electrogenic Flux through Multiple Transport Systems

Li, Zhen-Chang; Bush, Daniel R.
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em /09/1990 EN
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Amino acid transport into plasma membrane vesicles isolated from mature sugar beet (Beta vulgaris L. cv Great Western) leaves was investigated. The transport of alanine, leucine, glutamine, glutamate, isoleucine, and arginine was driven by a trans-membrane proton concentration difference. ΔpH-Dependent alanine, leucine, glutamine, and glutamate transport exhibited simple Michaelis-Menten kinetics, and double-reciprocal plots of the data were linear with apparent Km values of 272, 346, 258, and 1981 micromolar, respectively. These results are consistent with carrier mediated transport. ΔpH-Dependent isoleucine and arginine transport exhibited biphasic kinetics, suggesting these amino acids may be transported by at least two transport systems. Symport mediated alanine transport was electrogenic as demonstrated by the effect of membrane potential (ΔΨ) on ΔpH-dependent flux. In the absence of significant charge compensation, a low rate of alanine transport was observed. When ΔΨ was held at 0 millivolt with symmetric potassium concentrations and valinomycin, the rate of flux was stimulated fourfold. In the presence of a negative ΔΨ, alanine transport increased sixfold. These results are consistent with an electrogenic transport process which results in a net flux of positive charge into the vesicles. The effect of changing ΔΨ on the kinetics of alanine transport altered Vmax with no apparent change in Km. Amino acid transport was inhibited by the protein modifier diethyl pyrocarbonate...

Mutants of Synechocystis PCC6803 Defective in Inorganic Carbon Transport 1

Ogawa, Teruo
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em /10/1990 EN
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465.76043%
Eighty mutants of Synechocystis PCC6803 that require high CO2 for growth were examined with a mass spectrometer for their ability to take up CO2 in the light. Two of these mutants (type A) did not show any CO2 uptake while the rest of the mutants (type B) took up CO2 actively. Type A mutants (RKa and RKb) and one type B mutant (RK11) were partially characterized. At 3% CO2, growth rates of the mutants and the wild type (WT) were similar. Under air levels of CO2, growth of RKa and RKb was very slow, and RK11 did not grow at all. The photosynthetic affinities for inorganic carbon (Ci) in these three mutants were about 100 times lower than the affinity in WT. The following characteristics of type A mutants indicated that the mutants have a defect in their CO2-transport system: (a) the activity of 13C18O2 uptake in RKa and RKb in the light was less than 5% the activity in WT, and (b) each mutant had only a low level of activity of 14CO2 uptake as measured by the method of silicone oil-filtering centrifugation. The HCO3−-transport system was also impaired in these mutants. The activity of H14CO3− uptake was negligibly low in RKb and was one-third the activity of WT in RKa. On the other hand, the type B mutant, RK11, transported CO2 and HCO3− into the intracellular Ci pool as actively as WT but was unable to utilize it for photosynthesis. Complementation analysis of type A mutants indicated that RKa and RKb have mutations in different regions of the genome. These results suggested that at least two kinds of proteins are involved in the Ci-transport system.

Molecular Cloning of Tomato Plasma Membrane H+-ATPase 1

Ewing, Nicholas N.; Wimmers, Larry E.; Meyer, David J.; Chetelat, Roger T.; Bennett, Alan B.
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em /12/1990 EN
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465.71402%
Two cDNA clones (LHA1 and LHA2) from tomato (Lycopersicon esculentum) which likely encode isoforms of the plasma membrane H+-ATPase were isolated. The longest cDNA (3229 base pairs), LHA1, comprises an open reading frame that encodes a 956 amino acid, 105 kilodalton polypeptide with several potential transmembrane domains. In vitro transcription and translation of LHA1 yields a major translation product of approximately 100 kilodaltons that is immunoprecipitable with antiserum to the corn root plasma membrane H+-ATPase. LHA2 encodes a portion of a coding sequence that is 96% identical to LHA1, suggesting that LHA2 encodes an isoform of the H+-ATPase. Genomic DNA gel blot analysis indicates that both LHA1 and LHA2 hybridize to a common set of six to eight restriction fragments at moderate stringency and to single distinct fragments at high stringency. LHA1 and LHA2 map to distinct sites on chromosomes three and six, respectively. RNA gel blot analysis indicates that both LHA1 and LHA2 hybridize to 3.4 kilobase pair transcripts present in both leaves and roots, although the LHA2 transcript is relatively more abundant in leaves than in roots. These results indicate that in tomato as many as six to eight genes may encode the plasma membrane H+-ATPase...

Substrate Specificity of the H+-Sucrose Symporter on the Plasma Membrane of Sugar Beets (Beta vulgaris L.) 1: Transport of Phenylglucopyranosides

Hecht, Roland; Slone, J. Henry; Buckhout, Thomas J.; Hitz, William D.; VanDerWoude, William J.
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em /06/1992 EN
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Previous results (TJ Buckhout, Planta [1989] 178: 393-399) indicated that the structural specificity of the H+-sucrose symporter on the plasma membrane from sugar beet leaves (Beta vulgaris L.) was specific for the sucrose molecule. To better understand the structural features of the sucrose molecule involved in its recognition by the symport carrier, the inhibitory activity of a variety of phenylhexopyranosides on sucrose uptake was tested. Three competitive inhibitors of sucrose uptake were found, phenyl-α-d-glucopyranoside, phenyl-α-d-thioglucopyranoside, and phenyl-α-d-4-deoxythioglucopyranoside (PDTGP; Ki = 67, 180, and 327 micromolar, respectively). The Km for sucrose uptake was approximately 500 micromolar. Like sucrose, phenyl-α-d-thioglucopyranoside and to a lesser extent, PDTGP induced alkalization of the external medium, which indicated that these derivatives bound to and were transported by the sucrose symporter. Phenyl-α-d-3-deoxy-3-fluorothioglucopyranoside, phenyl-α-d-4-deoxy-4-fluorothioglucopyranoside, and phenyl-α-d-thioallopyranoside only weakly but competively inhibited sucrose uptake with Ki values ranging from 600 to 800 micromolar, and phenyl-α-d-thiomannopyranoside, phenyl-β-d-glucopyranoside, and phenylethyl-β-d-thiogalactopyranoside did not inhibit sucrose uptake. Thus...

Proton Transport Activity of the Purified Vacuolar H+-ATPase from Oats 1: Direct Stimulation by Cl−

Ward, John M.; Sze, Heven
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em /07/1992 EN
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465.76043%
To determine whether the detergent-solubilized and purified vacuolar H+-ATPase from plants was active in H+ transport, we reconstituted the purified vacuolar ATPase from oat roots (Avena sativa var Lang). Triton-solubilized ATPase activity was purified by gel filtration and ion exchange chromatography. Incorporation of the vacuolar ATPase into liposomes formed from Escherichia coli phospholipids was accomplished by removing Triton X-100 with SM-2 Bio-beads. ATP hydrolysis activity of the reconstituted ATPase was stimulated twofold by gramicidin, suggesting that the enzyme was incorporated into sealed proteoliposomes. Acidification of K+-loaded proteoliposomes, monitored by the quenching of acridine orange fluorescence, was stimulated by valinomycin. Because the presence of K+ and valinomycin dissipates a transmembrane electrical potential, the results indicate that ATP-dependent H+ pumping was electrogenic. Both H+ pumping and ATP hydrolysis activity of reconstituted preparations were completely inhibited by <50 nanomolar bafilomycin A1, a specific vacuolar type ATPase inhibitor. The reconstituted H+ pump was also inhibited by N,N′-dicyclohexylcarbodiimide or NO3− but not by azide or vanadate. Chloride stimulated both ATP hydrolysis by the purified ATPase and H+ pumping by the reconstituted ATPase in the presence of K+ and valinomycin. Hence...

In Vitro Analysis of the H+-Hexose Symporter on the Plasma Membrane of Sugarbeets (Beta vulgaris L.) 1

Tubbe, Axel; Buckhout, Thomas J.
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em /07/1992 EN
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465.76043%
The mechanism of hexose transport into plasma membrane vesicles isolated from mature sugarbeet leaves (Beta vulgaris L.) was investigated. The initial rate of glucose uptake into the vesicles was stimulated approximately fivefold by imposing a transmembrane pH gradient (ΔpH), alkaline inside, and approximately fourfold by a negative membrane potential (ΔΨ), generated as a K+-diffusion potential, negative inside. The -fold stimulation was directly related to the relative ΔpH or ΔΨ gradient imposed, which were determined by the uptake of acetate or tetraphenylphosphonium, respectively. ΔΨ- and ΔpH-dependent glucose uptake showed saturation kinetics with a Km of 286 micromolar for glucose. Other hexose molecules (e.g. 2-deoxy-d-glucose, 3-O-methyl-d-glucose, and d-mannose) were also accumulated into plasma membrane vesicles in a ΔpH-dependent manner. Inhibition constants of a number of compounds for glucose uptake were determined. Effective inhibitors of glucose uptake included: 3-O-methyl-d-glucose, 5-thio-d-glucose, d-fructose, d-galactose, and d-mannose, but not 1-O-methyl-d-glucose, d- and l-xylose, l-glucose, d-ribose, and l-sorbose. Under all conditions of proton motive force magnitude and glucose and sucrose concentration tested...

Effect of Elicitors on the Plasmamembrane of Petunia hybrida Cell Suspensions 1: Role of ΔpH in Signal Transduction

Hagendoorn, Marc J. M.; Poortinga, Arne M.; Sang, Harro W. Wong Fong; van der Plas, Linus H. W.; van Walraven, Hendrika S.
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em /08/1991 EN
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465.71402%
Primary processes during elicitation of the phenylpropanoid pathway (PPP) were studied in Petunia hybrida cell suspensions. We tested the hypothesis that decrease of the proton gradient across the plasma membrane activates the PPP. Induction of the PPP was determined by measuring phenylalanine ammonia lyase activity. A variety of ATPase inhibitors and ionophores were tested for the ability to elicit the PPP. The ATPase inhibitors orthovanadate and N,N′-dicyclohexylcarbodiimide and the ionophores carbonyl cyanide-4-trifluoromethoxyphenylhydrazone and nigericin were all effective elicitors. Carbonyl cyanide-4-trifluoromethoxyphenylhydrazone and nigericin elicit also when used in combination with N,N′-dicyclohexylcarbodiimide. Valinomycin had little effect on phenylalanine ammonia lyase activity. Treatment with orthovanadate or nigericin led to the formation of lignin. Alkalinization of the external medium by N,N′-dicyclohexylcarbodiimide, carbonyl cyanide-4-trifluoromethoxyphenylhydrazone, and nigericin was observed directly with the use of a sensitive pH electrode and internal acidification was deduced from the changes in emission intensity of the fluorescent probe bis[3-propyl-5-oxoisoxazol-4-yl] pentamethineoxonol. These data indicate that changes in the activity of the plasmamembrane H+-ATPase...

ΔpH-Dependent Amino Acid Transport into Plasma Membrane Vesicles Isolated from Sugar Beet (Beta vulgaris L.) Leaves: II. Evidence for Multiple Aliphatic, Neutral Amino Acid Symports

Li, Zhen-Chang; Bush, Daniel R.
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em /08/1991 EN
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Proton-coupled aliphatic, neutral amino acid transport was investigated in plasma membrane vesicles isolated from sugar beet (Beta vulgaris L., cv Great Western) leaves. Two neutral amino acid symport systems were resolved based on inter-amino acid transport competition and on large variations in the specific activity of each porter in different species. Competitive inhibition was observed for transport competition between alanine, methionine, glutamine, and leucine (the alanine group) and between isoleucine, valine, and threonine (the isoleucine group). The apparent Km and Ki values were similar for transport competition among amino acids within the alanine group. In contrast, the kinetics of transport competition between these two groups of amino acids did not fit a simple competitive model. Furthermore, members of the isoleucine group were weak transport antagonists of the alanine group. These results are consistent with two independent neutral amino acid porters. In support of that conclusion, the ratio of the specific activity of alanine transport versus isoleucine transport varied from two- to 13-fold in plasma membrane vesicles isolated from different plant species. This ratio would be expected to remain relatively stable if these amino acids were moving through a single transport system and...

Evidence for a Highly Specific K+/H+ Antiporter in Membrane Vesicles from Oil-Seed Rape Hypocotyls

Cooper, Sagi; Lerner, Henri R.; Reinhold, Leonora
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em /11/1991 EN
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465.76043%
We present evidence strongly suggesting that a proton gradient (acid inside) is used to drive an electroneutral, substrate-specific, K+/H+ antiport in both tonoplast and plasma membrane-enriched vesicles obtained from oilseed rape (Brassica napus) hypocotyls. Proton fluxes into and out of the vesicles were monitored both by following the quenching and restoration of quinacrine fluorescence (indicating a transmembrane pH gradient) and of oxonol V fluorescence (indicating membrane potential.) Supply of K+ (with Cl− or SCN−) after a pH gradient had been established across the vesicle membrane by provision of ATP to the H+-ATPase dissipated the transmembrane pH gradient but did not depolarize the positive membrane potential. Evidence that the K+/H+ exchange thus indicated could not be accounted for by mere electric coupling included the findings that, first, no positive potential was generated when KSCN or KCl was supplied, even in the absence of 100 millimolar Cl− and, second, efflux of K+ from K+-loaded vesicles drives intravesicular accumulation of H+ against the electrochemical potential gradient. Neither was the exchange due to competition between K+ and quinacrine for membrane sites, nor to inhibition of the H+-ATPase. Thus...

Evidence for the Existence of One Antenna-Associated, Lipid-Dissolved and Two Protein-Bound Pools of Diadinoxanthin Cycle Pigments in Diatoms[C][W]

Lepetit, Bernard; Volke, Daniela; Gilbert, Matthias; Wilhelm, Christian; Goss, Reimund
Fonte: American Society of Plant Biologists Publicador: American Society of Plant Biologists
Tipo: Artigo de Revista Científica
EN
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We studied the localization of diadinoxanthin cycle pigments in the diatoms Cyclotella meneghiniana and Phaeodactylum tricornutum. Isolation of pigment protein complexes revealed that the majority of high-light-synthesized diadinoxanthin and diatoxanthin is associated with the fucoxanthin chlorophyll protein (FCP) complexes. The characterization of intact cells, thylakoid membranes, and pigment protein complexes by absorption and low-temperature fluorescence spectroscopy showed that the FCPs contain certain amounts of protein-bound diadinoxanthin cycle pigments, which are not significantly different in high-light and low-light cultures. The largest part of high-light-formed diadinoxanthin cycle pigments, however, is not bound to antenna apoproteins but located in a lipid shield around the FCPs, which is copurified with the complexes. This lipid shield is primarily composed of the thylakoid membrane lipid monogalactosyldiacylglycerol. We also show that the photosystem I (PSI) fraction contains a tightly connected FCP complex that is enriched in protein-bound diadinoxanthin cycle pigments. The peripheral FCP and the FCP associated with PSI are composed of different apoproteins. Tandem mass spectrometry analysis revealed that the peripheral FCP is composed mainly of the light-harvesting complex protein Lhcf and also significant amounts of Lhcr. The PSI fraction...

A Protein Kinase, Calcineurin B-Like Protein-Interacting Protein Kinase9, Interacts with Calcium Sensor Calcineurin B-Like Protein3 and Regulates Potassium Homeostasis under Low-Potassium Stress in Arabidopsis1[W][OA]

Liu, Li-Li; Ren, Hui-Min; Chen, Li-Qing; Wang, Yi; Wu, Wei-Hua
Fonte: American Society of Plant Biologists Publicador: American Society of Plant Biologists
Tipo: Artigo de Revista Científica
EN
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Potassium (K+) is an essential macronutrient for plant growth and development. Previous studies have demonstrated that Calcineurin B-Like Protein1 (CBL1) or CBL9 and CBL-Interacting Protein Kinase23 (CIPK23) regulate K+ uptake in Arabidopsis (Arabidopsis thaliana) roots by modulating K+ channel Arabidopsis K+ Transporter1. In this study, we show that the protein kinase CIPK9 interacts with the calcium sensor CBL3 and plays crucial roles in K+ homeostasis under low-K+ stress in Arabidopsis. Arabidopsis wild-type plants showed leaf chlorotic symptoms when grown for 10 d on low-K+ (100 μm) medium. Here, we show that plants lacking CIPK9 displayed a tolerant phenotype to low-K+ stress, which still maintained green leaves when the wild-type plants showed typical K+-deficient symptoms. Overexpressing lines of CIPK9 resulted in a low-K+-sensitive phenotype compared with wild-type plants. Furthermore, CBL2 and CBL3 were identified as upstream regulators of CIPK9. Both CBL2- and CBL3-overexpressing lines displayed similar low-K+-sensitive phenotypes and K+ contents to CIPK9-overexpressing lines. However, only cbl3 mutant plants, but not cbl2 mutant plants, showed the low-K+-tolerant phenotype similar to cipk9 mutants. Taken together, these results demonstrate that CIPK9 and CBL3 work together and function in K+ homeostasis under low-K+ stress in Arabidopsis.

Cation transport mechanisms in Mycoplasma mycoides var. Capri cells. The nature of the link between K+ and Na+ transport

Benyoucef, Mohammed; Rigaud, Jean-Louis; Leblanc, Gérard
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em 15/12/1982 EN
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We have studied the links between the mechanisms of Na+, K+ and H+ movements in glycolysing Mycoplasma mycoides var. Capri cells. In the light of the results reported in the preceding paper [Benyoucef, Rigaud & Leblanc (1982) Biochem. J. 208, 529–538], we investigated certain properties of the membrane-bound ATPase of Mycoplasma cells, with special reference to its ionic requirements and sensitivity to specific inhibitors. Our findings show, first, that, although Na+ stimulated ATPase activity, K+ did not affect it, and, secondly, that NN′-dicyclocarboidi-imide and 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole (NBD) were potent inhibitors of the basal ATPase activity, which was unaffected by vanadate and ouabain. We also investigated the movements of Na+ and H+ under the experimental conditions applied to the study of the K+ uptake reported in the preceding paper, and found that when `Na+-loaded cells' previously equilibrated with 22Na+ were diluted in a sodium-free medium, addition of glucose induced a rapid efflux of 22Na+. This energy-dependent efflux was independent of the presence of KCl in the medium. Studies of the changes in internal pH by 9-aminoacridine fluorescence or [14C]methylamine distribution indicated that the movement of Na+ was coupled to that of protons moving in the opposite direction...

Cytoplasmic Membrane Changes during Adaptation of the Fresh Water Cyanobacterium Synechococcus 6311 to Salinity 1

Lefort-Tran, Marcelle; Pouphile, Monique; Spath, Susan; Packer, Lester
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em /07/1988 EN
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In this investigation, changes were characterized in cell structure and cytoplasmic membrane organization that occur when the freshwater cyanobacterium Synechococcus 6311 is transferred from `low salt' (0.03 molar NaCl) to `high salt' (0.5 molar NaCl) media (i.e. sea water concentration). Cells were examined at several time points after the imposition of the salt stress and compared to control cells, in thin sections and freeze fracture electron microscopy, and by flow cytometry. One minute after exposure to high salt, i.e. `salt shock,' virtually all intracellular granules disappeared, the density of the cytoplasm decreased, and the appearance of DNA material was changed. Glycogen and other granules, however, reappeared by 4 hours after salt exposure. The organization of the cytoplasmic membrane undergoes major reorganization following salt shock. Freeze-fracture electron microscopy showed that small intramembrane particles (diameters 7.5 and 8.5 nanometers) are reduced in number by two- to fivefold, whereas large particles, (diameters 14.5 and 17.5 nanometers) increase two- to fourfold in frequency, compared to control cells grown in low salt medium. The changes in particle size distribution suggest synthesis of new membrane proteins...

Immunocytolocalization of Plasma Membrane H+-ATPase

Parets-Soler, Antonio; Pardo, Jose M.; Serrano, Ramon
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em /08/1990 EN
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465.76043%
The localization of plasma membrane H+-ATPase has been studied at the optical microscope level utilizing frozen and paraffin sections of Avena sativa and Pisum sativum, specific anti-ATPase polyclonal antibody, and second antibody coupled to alkaline phosphatase. In leaves and stems the ATPase is concentrated at the phloem, supporting the notion that it generates the driving force for phloem loading. In roots the ATPase is concentrated at both the periphery (rootcap and epidermis) and at the central cylinder, including endodermis and vascular cells. This supports a `two-pump' mechanism for ion absorption, involving active uptake at the epidermis, symplast transport across the cortex, and active efflux at the xylem. The low ATPase content of root meristem and elongation zone may explain the observed transorgan H+ currents, which leave nongrowing parts and enter growing tips.