Fluxes of Na+, Rb+ and Cl- Ions in Excised Roots of Sugar Beet Seedlings

Uptake and fluxes of sodium, rubidium (instead of potassium), and chloride ions in segments of 3-week-old sugar beet roots were studied. Radioactive ²²Na, ⁸⁶Rb and ³⁶Cl were used for labelling of the ions. Compartmental analysis was used to obtain the fluxes and concentrations in the cell compartments. The passive or active character of the movements was examined by the Ussing-Teorell equation and compared with electropotential measurements. In the case of sodium, net flux was in the outward direction over both tonoplast and plasmalemma, but the active components were directed away from the cytoplasm. Potassium was close to equilibrium. Chloride was actively transported from the medium to the cytoplasm, and — contrary to observations in other systems — from the vacuole to the cytoplasm. This unusual situation may be caused by a loss of sugar, both by lowering the energy supply and by formation of organic acids.     

Electrochemical gradients and k and cl fluxes in excised corn roots

The compartmental analysis method was used to estimate the K⁺ and Cl⁻ fluxes for cells of excised roots of Zea mays L. cv. Golden Bantam. When the measured fluxes are compared to those calculated with the Ussing-Teorell flux-ratio equation, an active inward transport of Cl⁻ across the plasmalemma is indicated; the plasmalemma K⁺ fluxes are not far different from those predicted for passive diffusion, although an active inward transport cannot be precluded. Whether fluxes across the tonoplast are active or passive depends upon the vacuolar potential which is unknown. Assuming no electropotential gradient, the tracer flux ratios are fairly close to those predicted for passive movement. However, if the vacuole is positive by about 10 millivolts relative to the cytoplasm, the data suggest active inward transport for K⁺ and outward transport for Cl⁻.     

Control of Ion Fluxes in Stomatal Guard Cells

Opening and closing of the stomatal pore is associated with very large changes in K-salt accumulation in stomatal guard cells. This review discusses the ionic relations of guard cells in relation to the general pattern of transport processes in plant cells, in plasmalemma and tonoplast, involving primary active transport of protons, proton-linked secondary active transport, and a number of gated ion channels. The evidence available suggests that the initiation of stomatal opening is regulated through the uptake mechanisms, whereas initiation of stomatal closing is regulated by control of ion efflux at the plasmalemma, and of fluxes to and from the vacuole. In response to a closing signal there are large transient increases in efflux of both Cl^? (or Br^?) and Rb⁺ (K⁺) at the plasmalemma, with also a probable increase in anion flux from vacuole to cytoplasm and decrease in anion flux from cytoplasm to vacuole. A speculative hypothetical sequence of events is discussed, by which the primary response to a closing signal is an increase in Ca²⁺ influx at the plasmalemma, producing depolarisation and increase in cytoplasmic Ca²⁺. The consequent opening of Ca²⁺-sensitive Cl^? channels, and voltage-sensitive K⁺ channels (also Ca²⁺-sensitive?) in the plasmalemma, and of a Ca²⁺-sensitive nonspecific channel in the tonoplast, could produce the flux effects identified by the tracer work; this speculation is also consistent with the Ca²⁺-sensitivity of the response to closing signals and with evidence from patch clamping that such channels exist in at least some plant cells, though not yet all shown in guard cells.     

Compartments and Fluxes of K, NA, and CL in Avena Coleoptile Cells

By the compartmental analysis method of MacRobbie and Dainty, and Pitman, estimates of K⁺, Na⁺, and Cl⁻ concentrations and fluxes were obtained for the cytoplasm and vacuole of coleoptile cells of oat, Avena sativa L. cv. Victory. Double labeling was used in experiments with ⁴²K plus ²²Na and with ⁴²K plus ³⁶Cl in a complete nutrient solution. At the plasmalemma, according to the Ussing-Teorell flux ratio equation, Na⁺ is pumped out and Cl⁻ is actively transported inward. The results with K⁺ are less conclusive, but it is probably pumped in. At the tonoplast there is an active inward transport of Na⁺ and probably of K⁺, but the status of Cl⁻ is uncertain, depending upon whether there is an electrical potential difference between the cytoplasm and vacuole. The results suggest that ion selectivity resides mostly in the plasmalemma. Possible errors in the estimates and interpretations are discussed.     

Studies on the tonoplast action potential ofNitella flexilis

Summary The origins of the two peaks of the action potential inNitella flexilis were analyzed by inserting two microelectrodes. one into the vacuole and the other into the cytoplasm. It was unequivocally demonstrated that the rapid first peak was generated at the plasmalemma and the slow second peak at the tonoplast. MnCl₂ applied in the external medium abolished the second, tonoplast, peak but not the first, plasmalemma, peak, MnCl₂ also inhibited the cessation of the cytoplasmic streaming accompanying the action potential. CaCl₂ added in MnCl₂-containing medium recovered generation of the tonoplast action potential and the streaming cessation. Since it has been established that the cessation of cytoplasmic streaming on membrane excitation is caused by an increase in cytoplasmic free Ca^2– (Williamson, R.E., Ashley, C.C., 1982.Nature (London) 296:647–651: Tominaga, Y., Shimmen, T., Tazawa, M., 1983,Protoplasma 116:75–77), it is suggested that the tonoplast action potential is also induced by an increase in cytoplasmic Ca²⁺ resulting from the plasmalemma excitation. When vacuolar Cl was replaced with SO ₄ ² by vacuolar perfusion, the polarity of the second, slow peak was reversed from vacuolar positive to vacuolar negative with respect to the cytoplasm, supporting the previous report that the tonoplast action potential is caused by increase in Cl permeability (Kikuyama, M., Tazawa, M., 1976.J. Membrane Biol.29:95–110).     

Cation fluxes in excised and intact roots in relation to specific and varietal differences

Summary In this short survey differences between species and varieties in the four major mechanisms that affect selective uptake of potassium and sodium to the plant within the root are considered. These include influx selectivity, K⁺/Na⁺ exchange at the plasmalemma, and selectivity at the tonoplast as well as at the symplasm-xylem boundary. The affinity of various plants for potassium influx in system 1 is rather uniform although varietal differences in barley have been observed. Differences are much more pronounced for sodium influx, for which Helianthus showed rather high and Fagopyrum rather low affinity. There is substantial variation between species in the efficiency of K⁺/Na⁺ exchange at the plasmalemma of cortical root cells; the three cereals Hordeum, Triticum, and Secale were highly efficient while K⁺/Na⁺ exchange in Atriplex, Helianthus and Allium was poor, even if the cytoplasmic sodium content was accounted for. Apparently there was no direct relation between salt tolerance and K⁺/Na⁺ exchange. The observed differences in the efficiency of K⁺-dependent sodium extrusion or K⁺/Na⁺ exchange were not due to the use of excised roots, they were observed also when roots of whole seedlings were investigated. At the tonoplast a 11 exchange of vacuolar potassium for sodium has been observed in roots of Hordeum. By this exchange sodium ions are removed from the symplasm and potassium ions are recovered from vacuoles and thus made available for transport to the shoot. Indications for specific differences in this exchange have been observed; the exchange appears to be more efficient in Helianthus than in Hordeum roots. More comparative studies are needed here. At the boundary between symplasm and xylem vessels selectivity can be set up during xylem release of cations and there are reports that suggest a preference for sodium (Lycopersicum cheesemanii, Solanum pennellii, and Suaeda) and for varietal differences amongst tomatoes. Selectivity at this boundary, the plasmalemma of the xylem parenchyma cells was described in this paper by the selectivity ratio of transport that relates the rates of xylem transport to the cytoplasmic sodium and potassium concentrations. Based on this ratioAtriplex hortensis was shown to discriminate for sodium during xylem release while there was little selectivity in Hordeum and possibly some discrimination in favour of K⁺ in Allium roots. The data are shortly discussed in relation to salt tolerance and to the breeding of salt-tolerant crop varieties.     

Tonoplast action potential inNitella in relation to vacuolar chloride concentration

Summary The action potential ofNitella internode was studied in relation to K⁺ and Cl^– concentrations in the vacuole. When the vacuole ofNitella pulchella was filled with an artificial solution with extremely low Cl^– concentration, a diphasic action potential (DAP) was observed. The first phase consists of a rapid depolarization followed by a relatively rapid repolarization, and the second one consists of a strong hyperpolarization followed by a gradual return to the resting potential.When the cell was stimulated immediately after the generation ofDAP, a monophasic action potential which resembles an action potential of the natural cell was observed, indicating that theDAP consists of two components with different refractory periods. The refractory period of the component responsible for the depolarizing phase is shorter than that of a component responsible for the hyperpolarizing phase. Measuring the plasmalemma potential and vacuolar potential separately, it was demonstrated that the hyperpolarizing component ofDAP originates from the tonoplast.The action potential of the tonoplast, in contrast with that of the plasmalemma, could be generated independently of concentration of K⁺ in the vacuole. Since the maximum amplitude of hyperpolarization decreased significantly by increasing Cl^– concentration of the vacuole, it is concluded that the tonoplast is very sensitive to Cl^– during excitation.     

Movement of Abscisic Acid Across the Plasmalemma and the Tonoplast of Guard Cells of Valerianella locusta

Uptake experiments and efflux compartmental analyses of abscisic acid (ABA) with acid treated epidermal peels of Valerianella locusta were performed to elucidate the mechanisms of transport of ABA across the plasmalemma and tonoplast of guard cells. ABA uptake across the plasmalemma is linearly correlated with external ABA concentration in the incubation medium. Under alkaline conditions ABA-uptake was not significantly above background, indicating that ABA uptake occurs mainly by diffusion of undissociated ABAH as the most permeable species, which is trapped afterwards in the alkaline cytosol as impermeable ABA^?. Efflux analysis of ABA revealed a saturable component of ABA transfer across the tonoplast. A Woolf-Augustinsson-Hofstee analysis suggested the existence of two transport systems for ABA at the tonoplast. The high affinity transport system had a K_M of 0.21 mol m^?3 and a V_max 85.8 amol ABA cell^?1 h^?1. Using the data of the uptake and efflux experiments we calculated the permeability coefficients of ABA for the plasmalemma and the tonoplast of guard cells, which are 2.46 10^?7 m s–¹ and 1.26 10^?8m s^?1, respectively. The distribution of the pH-probe (¹⁴C)-DMO between medium, cytosol and vacuole was investigated and used to calculate cytosolic and vacuolar pH. The vacuolar pH is too low to explain the high vacuolar ABA concentration by trapping of ABA^?, whereas the cytosol is sufficiently alkaline to act as an efficient anion trap. Therefore we conclude that ABA transport across the guard cell tonoplast is catalyzed by a saturable uptake component.     

Electrical Potentials and Na, K, and Cl Concentrations in the Vacuole and Cytoplasm of Nitella translucens

The potential differences across the tonoplast and plasmalemmamembranes have been measured in the single cells of Nitellatranslucens, the cells being immersed in an artificial pondwater (composition: NaCl _1.0 mM., KC1 _0.1 mM., CaCl₂, _0.1 mM.).The potential of the cytoplasm is –138 m V with respectto the bathing medium and –18 mV with respect to the vacuole.The concentrations of Na, K, and Cl have been measured in thetwo cell fractions. The concentrations in the flowing cytoplasmare: Na 14 mM., K 119 mM., and Cl 65 mM.; the vacuolar concentrationsare: Na 65 mM., K 75 mM.,and Cl 160 mM. The observed potential differences across the two membranesare compared with the Nernst potentials for all three ions.This analysis shows that all three ions are actively transportedat the plasmalemma: Na is pumped outwards while K and Cl arepumped inwards. At the tonoplast Na is pumped into the vacuolewhile K and Cl are close to electrochemical equilibrium. The inhibitor, ouabain, has no effect on the cell resting potential.     

Abscisic-acid-induced turion formation in Spirodela polyrrhiza L. IV. Comparative ion flux characteristics of the turion and the vegetative frond and the effect of ABA during early turion development

Abstract Using the method of compartmental analysis, the ion fluxes and compartment concentrations of Ca²⁺, K⁺ and Cl^- have been compared in the untreated vegetative frond and the abscisic acid (ABA) induced turion of Spirodela polyrrhiza. The ABA-induced turion is characterized by reduced Ca²⁺ exchange across the tonoplast and low vacuolar Ca²⁺ concentration relative to the vegetative frond. In addition the turion exhibits a higher plasmalemma flux with a correspondingly high Ca²⁺ concentration in the cytoplasm. The concentration of K⁺ and Cl^- is much lower in the cytoplasm of the ABA-induced turion than in the vegetative frond with the influx/efflux ratio at both the plasmalemma and the tonoplast being less than 1, a finding exhibited also in dormant storage tissue. Treatment of vegetative fronds with ABA for 18 h resulted in a reduced K⁺ plasmalemma efflux relative to untreated vegetative fronds and a concomitant increase in the cytoplasmic concentration. There was no rapid effect of ABA on Ca²⁺, K⁺ or Cl^- fluxes through either membrane. These results are consistent with the notion that drastic changes in ion fluxes and concentrations in the turion are a secondary consequence of ABA-induced development, possibly due to prior regulation by ABA of enzymes inherent to processes involved in membrane transport.     

Two components of auxin transport

The transport of indoleacetic acid-1-¹⁴C out of sunflower stem sections has been analyzed by a compartmental analysis procedure in which the radioactivity moving out of the tissue (log per cent) is plotted against time. The analysis indicates that indoleacetic acid is transported via a fast transport system (t_½ of about 30 minutes) and a slow transport system (t_½ about 10 hours). While we do not know the sources of these two pools, by analogy with ion transport studies, the fast efflux is characteristic of transport from the cytoplasm across the plasmalemma and the slow efflux is characteristic of transport across the tonoplast and thus out of the vacuole. Both components of transport are inhibited by 2,3,5-triiodobenzoic acid.     

Activity of Ion Transporters and Salt Tolerance in Barley

The authors attempted to relate the cultivar-specific salt tolerance in barley (Hordeum distichum L.) to the efficiency of ion transporters in the plasmalemma and tonoplast. The study involved plasmalemma and tonoplast membrane vesicles isolated from roots and leaves of the 7-day-old barley seedlings exposed to elevated NaCl concentrations. Two barley cultivars were employed: salt-tolerant cv. Elo and salt-susceptible cv. Belogorskii. The vesicles were used to measure the transport activity of plasmalemma and tonoplast proton pumps and the cation/anion exchange. The data obtained in the experiments demonstrated that the changes in the activity of ion transporters under salt stress conditions correlated with the barley cultivar-specific tolerance to elevated NaCl concentrations.     

The Membrane Potential of Nitella translucens

The effects of changing the external concentrations of Na, K,Ca, and Cl on the potentials of the cytoplasm and the vacuolewith respect to the bathing medium of the internodal cells ofNitella translucens have been investigated. The potential differencebetween the vacuole and the cytoplasm is practically unaffectedby the concentration changes. The observed changes of potentialdifference are therefore attributed to the boundary separatingthe cytoplasm from the medium; this boundary is possibly a plasmalemma–cellwall complex. The difference of potential between the cell walland the medium has also been measured and, in the presence ofCa, shown to be markedly sensitive only to the external Ca concentration.The results are divided into two sections: (a) for cells pretreatedin 5 mM NaCl, the subsequent experiments being carried out inCa-free media, and (b) for cells initially immersed in a standardartificial pond water containing the chlorides of Na, K, Ca.With the pretreated cells the external Na/K ratio was variedwith the total NaCl+KCl concentration kept constant at 1.1 mM.The results suggest that over a limited range of concentrationsthe cytoplasm-medium potential difference can be described byan equation similar in form to a Goldman equation but containingonly terms for Na and K, the average value of the permeabilityratio (= P_Na/P_K) being 0.27. In the presence of Ca the effectsof Na and K on the cytoplasm-medium potential difference aregreatly reduced, while the effect of Ca is relatively large.The results cannot be fitted to any form of Goldman equationcontaining terms for the major ions. The possibility of a contributionto the plasmalemma potential from electrogenic pumps is brieflydiscussed. Measurements of the Na and K content of the cytoplasmand the vacuole have been made for the pretreated cells. TheNa concentration in the cytoplasm is 37 mM and in the vacuole73 mM; the K concentration is 93 mM in the cytoplasm and 67mM in the vacuole. The Nernst potentials for both ions are comparedwith the cytoplasm-medium and cytoplasm-vacuole potential differences.This analysis shows that Na is actively transported from thecytoplasm into the medium as well as into the Vacuole; K ispumped into the cytoplasm from the medium but appears to beclose to electrochemical equilibrium across the tonoplast. ThisConfirms previously published work.     

Pinocytosis in the root cells of a salt-accumulating halophyte <Emphasis Type="Italic">Suaeda altissima</Emphasis> and its possible involvement in chloride transport

Ultrastructure of root cells in salt-accumulating halophyte Suaeda altissima (L.) Pall. was examined with transmission electron microscopy. Plants were grown hydroponically on nutrient media containing 3, 50, 250, and 500 mM NaCl. Some plants were exposed to hypersomotic salt shock by an abrupt increase in NaCl concentration from 50 to 400 mM. Growing S. altissima plants at high NaCl concentrations induced the formation of type 1 pinocytotic structures in root cells. Type 1 structures appeared as pinocytotic invaginations of two membranes, the plasmalemma and tonoplast. These invaginations into vacuoles gave rise to freely ‘floating’ multivesicular bodies (MVB) enclosed by a double membrane layer. The pinocytotic invaginations and MVB contained the plasmalemma-derived vesicles and membranes of endosome origin. The hyperosmotic salt shock led to formation of type 2 and type 3 pinocytotic structures. The type 2 structures were formed as pinocytotic invaginations of the tonoplast and gave rise to MVB in vacuoles. Unlike type 1 MVB, the type 2 MVB had only one enclosing membrane, the tonoplast. The type 3 structures appeared as the plasmalemma-derived vesicles located in the periplasmic space. The cytochemical electron-microscopy method was applied to determine the intracellular Cl^? localization. This method, based on sedimentation of electron-dense AgCl granules in tissues treated with silver nitrate, showed that the pinocytotic structures of all types contain Cl^? ions. The presence of Cl^? in pinocytotic structures implies the involvement of these structures in Cl^? transport between the apoplast, cytoplasm, and the vacuole.     

Impact of hypoosmotic challenges on spongy architecture of the cytoplasm of the giant marine alga Valonia utricularis

Summary. The ultrastructure of the several micrometers thick cytoplasmic layer of the giant marine alga Valonia utricularis displays characteristics which are apparently linked with the capability of this alga to regulate turgor pressure. Transmission and scanning electron microscopy of cells prefixed in different ways, including a protocol that allows prefixation of the alga in a turgescent state, revealed a highly dendritic network of cytoplasmic strands connecting and enveloping the chloroplasts and the nuclei. Innumerable vacuolar entities are embedded in the network, giving the cytoplasm a spongy appearance. Vacuolar perfusion of turgor-pressure-clamped cells with prefixation solution containing tannic acid presented evidence that these vacuolar entities together with the huge central vacuole form a large unstirred continuum. In contrast to the tonoplast, the plasmalemma followed smoothly the lining of the cell wall, even at the numerous cell wall ingrowths. Sucrose, but not polyethylene glycol 6000, induced chloroplast clustering. Acute hypoosmotic treatment (established by reduction of external NaCl or by replacement of part of the external NaCl by equivalent osmotic concentrations of sucrose or polyethylene glycol 6000) resulted in a local relocation of the chloroplasts and cytoplasm towards the central vacuole. This effect did not occur when the relatively low reflection coefficients of these two osmolytes were taken into account. The increase in spacing between the spongy cytoplasm and the plasmalemma by chloroplast relocation (viewed by confocal laser scanning microscopy) was associated with a speckled appearance of the affected surface area under the light microscope. As indicated by electron microscopy, hypoosmotically induced chloroplast relocation resulted from disproportionate swelling of the vacuolar entities located close to the plasmalemma. The cytoskeleton in the cytoplasm and the mucopolysaccharide network in the central vacuole apparently resisted swelling of these compartments. This finding has the important consequence that relevant hydrostatic pressure gradients can be built up throughout the entire multifolded vacuolar space. This gradient could represent the trigger for turgor pressure regulation which is manifested electrically first in the tonoplast.Correspondence and reprints: Lehrstuhl für Biotechnologie, Biozentrum, Am Hubland, 97074 Würzburg, Federal Republic of Germany.     

Protein kinase activities in tonoplast and plasmalemma membranes from corn roots

Protein kinase and phosphatase activities were studied in plasmalemma and tonoplast membrane fractions from corn (Zea mays L.) roots in order to test the hypothesis that the tonoplast H⁺-ATPase is regulated by intrinsic protein phosphorylation (G Zocchi, SA Rogers, JB Hanson 1983 Plant Sci Lett 31: 215-221), and to facilitate future purification of kinase activities from these membranes. Kinase activity in the plasmalemma was about three-fold higher than in the tonoplast, and displayed Michaelis Menten-type behavior with a K_m value for MgATP²⁻ of about 50 micromolar. Both activities were optimal at 3 millimolar free Mg²⁺ and had pH optima at 6.6 and 7.0 for the plasmalemma and tonoplast, respectively. Kinase activities in both fractions were stimulated by 1 micromolar free Ca²⁺, but calmodulin had no stimulatory effect, and chlorpromazine was inhibitory only at high concentrations. The pattern of phosphopeptides on SDS polyacrylamide gel electrophoresis was similar in both fractions except for one band of 50 kilodaltons that was present only in the tonoplast. A partially purified H⁺-ATPase fraction was prepared from tonoplast membranes, incubated under conditions optimal for protein phosphorylation. The three polypeptides (of 67, 57, and 36 kilodaltons), enriched in this fraction, did not become phosphorylated, suggesting that this protein is not regulated by endogenous protein phosphorylation. Protein phosphatase activity was detected only in the plasmalemma fraction. These results indicate that a regulatory cycle of protein phosphorylation and dephosphorylation may operate in the plasmalemma. The activity in the tonoplast appears to originate from plasmalemma contamination.     

Electrophysiological Techniques and the Magnitudes of the Membrane Potentials and Resistances of Nitella translucens

The membrane resistance of internodal cells of Nitella translucensincreased by 50 per cent during the first 5 h after insertionof two microelectrodes into the vacuole even when precautionswere taken to eliminate external disturbances. The insertionof a third microelectrode into the cytoplasm did not affectthe resistance. In artificial pond water the final value forthe plasmalemma resistance was 112 k cm² and that for the tonoplastwas 6 k cm². The increase in the membrane potential after thefirst hour was less than 10 per cent. A recent suggestion that accurate measurements of the plasmalemmaresistance can be made with a microelectrode outside the plasmalemma,but in close contact with it, is criticized. Tests were made of the claim that leakage of current at thepoint where microelectrodes enter the cytoplasm gives rise toa local increase in current density at the tonoplast and henceleads to an overestimate of the tonoplast resistance. Valuesfor the tonoplast resistance obtained when the cytoplasmic microelectrodewas inserted through the plasmalemma were similar to those observedwhen it was pushed across the cell and inserted through thetonoplast at a point remote from the postulated current leakage.Furthermore, the tonoplast resistance stayed remarkably constantwhen the plasmalemma resistance varied in a way which wouldcause different proportions of the applied current to pass throughthe leak resistance and produce variations in the apparent tonoplastresistance. It is concluded that published values of the tonoplastresistance are not grossly inaccurate.     

灵武长枣果实质膜和液泡膜H~+-ATPase和H~+-PPase活性与果实糖分积累

以不同发育时期灵武长枣(Ziziphus jujuba cv.Lingwuchangzao)的果实为材料,通过测定与分析果肉组织中细胞质膜、液泡膜H+-ATPase和H+-PPase活性、果实糖分含量变化,研究了灵武长枣果实质膜、液泡膜H+-ATPase和H+-PPase活性与糖积累特性的关系。结果表明:(1)果实第二次快速生长期之前主要积累葡萄糖和果糖,之后果实迅速积累蔗糖,葡萄糖和果糖含量则逐渐下降,成熟期果实主要积累蔗糖。(2)在果实发育的缓慢生长期S1,质膜H+-ATPase活性最低;第一次快速生长期,质膜H+-ATPase活性最高;缓慢生长期S2,其活性降低;第二次快速生长期,质膜H+-ATPase活性升至次高;完熟期,质膜H+-ATPase活性下降幅度较大。(3)在果实发育过程中,液泡膜H+-ATPase和H+-PPase活性的变化趋势相似。缓慢生长期S1,液泡膜H+-ATPase和H+-PPase活性较低;从缓慢生长期S1至第一次快速生长期缓慢下降至最低;从第一次快速生长期开始,液泡膜H+-ATPase和H+-PPase活性呈现为逐渐增高的变化趋势;除第二次快速生长期以外,液泡膜H+-PPase活性始终高于H+-ATPase。由此推测,质膜H+-ATPase和液泡膜H+-ATPase、H+-PPase对灵武长枣果实糖分的跨膜次级转运起到重要的调控作用。     

Reactions of yeast cells to glycerol treatment alterations to membrane structure and glycerol uptake

Summary Ultrastructural alterations to the plasmalemma and tonoplast ofSaccharomyces cerevisiae were studied after incubation in hypertonic solutions of glycerol and sorbitol. After 20 to 30 minutes incubation in glycerol, the cells had shrunk to about 40% of their original volume. Large depressions of the plasmalemma were then always found associated with the typical plasmalemma invaginations. The vacuoles of treated cells changed to an irregular form, the tonoplast intramembranous particles were clustered, and large smooth areas appeared. After 6 to 12 hours incubation, cell and vacuole volume, as well as plasmalemma and tonoplast ultrastructure, had reverted to normal. The rate of recovery was strongly temperature dependent.Protoplasts could be similarly shrunk, but no alterations to the plasmalemma ultrastructure were then observed; however, the tonoplast revealed particle clustering as observed in whole cells. Protoplasts also reverted to normal volume and ultrastructure after prolonged incubation. Cells and protoplasts treated with sorbitol showed similar phenomena, but remained shrunken.By the use of radioactive tracers, glycerol was shown to penetrate cells, protoplasts and isolated vacuoles, but no uptake of sorbitol could be demonstrated.During the glycerol permeation period (0.5 to 6 hours), numerous vesicles were found in the cytoplasm and these were possibly engulfed by the vacuole. Associated with the engulfment, patches of tonoplast intramembranous particles were found in a semicrystalline array. Osmotic stress induced alterations to membrane ultrastructure, due to the use of cryoprotective agents, are discussed.A preliminary note of the paper was given at the Sixth European Congress on Electron Microscopy, Jerusalem, 1976.     

Interpretation of the dual isotherm for ion absorption in beet tissue

Beet discs aged in 0.5 mM CaSO₄ develop a capacity to absorb K⁺ and Cl⁻ from solutions of low concentration. The initial influx of these ions is described by a hyperbolic relationship with concentration in the range 0.01 to 0.5 mM KCl, which is identical with the system 1 absorption isotherm found in other tissues. A second hyperbolic isotherm, attributable to system 2, is found at higher concentrations (1-50 mM KCl).

When the transport of labeled ion to the vacuole is studied by wash-exchanging the bulk of the cytoplasmic label following the absorption period, it is noted that in the range of system 1, isotope influx to the vacuole increases with time as the concentration of labeled ions in the cytoplasm increases, while in the range of system 2, influx to the vacuole is constant from the beginning. Diminution of the cytoplasmic specific activity during radio-isotope absorption by prefilling the cytoplasm with the analogous unlabeled salt, markedly reduces subsequent radioisotope uptake to the vacuole only in the range of system 1. These experiments suggest that the cytoplasm serves as a mixing chamber, and that the plasma membrane controls ion uptake to the tissue at low concentrations, indicating that the system 1 isotherm reflects ion movement into the cytoplasm through the plasma membrane. Flux experiments support this conclusion, showing that development with age of the system 1 isotherm corresponds to a quantitatively similar increase in plasma membrane influx in 0.2 mM KCl.

At higher concentrations the outer membrane no longer rate-limits entry of ions to the vacuole. Isotope influx under these conditions, described by the system 2 isotherm, presumably reflects movement across the tonoplast.