Metabolism-linked Binding of Rubidium in the Free Space of Wheat Roots and its Relation to Active Uptake

Components of the initial uptake and active transport of rubidium were studied in potassium-starved and potassium-rich wheat plants. A metabolism-linked component of the initial uptake in the root was identified by means of DNP-treatment. It was shown that there is a direct proportionality between the magnitude of the metabolism-linked component of the initial uptake and the rate of active uptake. It is suggested that, in the free space, this metabolism-linked ion binding represents an initial step of the active ion transport preceding the subsequent carrier-mediated transport across the plasmalemma.     

Apparent Bicarbonate Uptake and Possible Plasmalemma Proton Efflux in Chara corallina

It is shown that the apparent uptake of bicarbonate by cells of Chara corallina could be the result of a proton efflux coupled to extracellular production of CO₂ from bicarbonate, with CO₂ being taken up by the cell. The theoretical results presented here show that the influx of CO₂ across the plasmalemma can be much greater than previously thought, if there is a large efflux of protons across the plasmalemma, and that, if this occurs, there would be a much steeper gradient of pH near the cell surface than previously thought possible.     

Mechanism of sugar retention by roots of intact maize and field bean plants

The re-uptake of sugars driven by the proton gradient was studied in sugar net-release and net-uptake experiments using roots of intact maize (Zea mays cv. Blizzard) and field bean (Vicia faba L. cv. Alfred) plants. The net release of sugars into the root medium (0.1 mM CaSO₄) was stimulated by: the protonophore CCCP (10 M); the sulfhydryl reagent NEM (300 M); the specific inhibitor of plasmalemma ATPase vanadate (0.5 mM); and the inhibitor of the glucose carrier phlorizin (2 mM). Net uptake of glucose, fructose and arabinose from 10 M external concentrations was also inhibited by these substances. Surprisingly fusicoccin, a stimulator of net proton release did not effect net sugar uptake. Medium pH values only influenced sugar net uptake if the pH was above 7. It is concluded that a degradation of the proton gradient across the plasmalemma stimulates net sugar release because of disturbed re-uptake of sugars (in particular glucose) via a proton/sugar cotransport system. Thus, the retention of sugars by root cells not only depends on the plasmalemma permeability but also on the electro-chemical proton gradient. If an electro-chemical proton gradient is established by plasmalemma ATPase activity the re-uptake of sugars by proton/sugar cotransport minimizes the release of sugars into the rhizosphere.     

Bioregulator enhancement of sink activity in sugar beet

The sink mobilizing abillity is partially determined by sugar uptake rates of storage cells. Two synthetic growth regulators (Pix and BAS 106W) were tested for their effect on sucrose uptake in root tissue discs or glucose uptake in cell cultures of sugar beet. In tissue discs, uptake at the plasmalemma was determined by incubating the discs for 1 h in the presence of 5 mM sucrose and at the tonoplast for 4 h in the presence of 40 mM sucrose. Cell cultures were incubated for 1 h in the presence of 1 mM glucose. Pix (10 mg l^–1) caused a 20% stimulation of active sucrose uptake at the plasmalemma. Active sucrose uptake at the tonoplast was increased 67% by 100 mg l^–1 Pix. No effect of BAS 106W was observed on sucrose uptake in tissue discs. In cell cultures, a 65% enhancement of active glucose uptake was observed with both Pix and BAs 106W. When the bioregulators were applied to the root medium of seedlings, Pix but not BAS 106W resulted in increased root/shoot ratio, translocation of ¹⁴C-assimilates, and allocation of more biomass to the root sink. The data suggested that sugar transport and translocation may be used as biochemical criteria for rapid screening of effective yield enhancing bioregulators.     

Kinetics of Ion Uptake in Higher Plants

Single, multiphasic mechanisms located in the plasmalemma and the tonoplast are proposed to mediate ion uptake in cells of higher plants. There are no parallel uptake mechanisms and, normally, little or no free diffusion across the plasmalemma. The plasmalemma controls the rate of uptake at low external salt concentrations, while the tonoplast may become rate-limiting at high concentrations.     

吲哚乙酸对向日葵下胚轴生长区单向K~+(~(86)Rb~+)通量的影响

植物生长素在刺激某些植物组织生长的同时促进K~+吸收和H~+分泌(Hager等1971,Cleland1975,赖寿鹏和倪晋山1983),可能是由于生长素刺激位于植物细胞质膜上的H~+泵ATP酶(Scherer 1984,赖寿鹏和倪晋山1985)。但以往的文献中只测定了生长素促进的植物组织对K~+的净吸收速率,即组织中K~+的累积速率或吸收溶液中K~+的减少速率。自从MacRobbie和Dainty(1958)首先运用放射性     

Sucrose transport and hexose release in the maize scutellum

Since hexoses readily diffuse from maize scutellum cells, it should be possible to detect them if they are produced during sucrose transport at the tonoplast or the plasmalemma. To test this idea, scutellum slices were placed in dinitrophenol (DNP) (which inhibits hexose utilization while greatly increasing utilization of vacuolar sucrose), and the utilization, uptake and leakage of sugars were measured. Only negligible amounts of hexose appeared in the DNP solution during a 5-hr incubation during which the slices metabolized 72μmol of sucrose. Glucose and fructose, added at a concentration of 2 mM, were taken up by the slices at rates 33% and 14% (respectively) of the rate of vacuolar sucrose utilization. It is suggested, therefore, that sucrose transport at the tonoplast does not release free hexose into the cytoplasm. Sucrose transport at the plasmalemma was studied using DNP- and mannose-treated slices. During incubation of these slices in sucrose, the disappearance of sucrose resulted in the appearance of significant quantities of glucose and fructose in the bathing solution. Evidence is presented that sucrose is split into glucose and fructose during transport across the plasmalemma. It is concluded that free hexose is not normally a product of this splitting but is a result of an uncoupling in the transport system caused by the DNP or mannose treatments.     

Ca2+ regulation of vascular smooth muscle

Regulation of intracellular free Ca2+ concentrations in vascular smooth muscle is accomplished mainly by Ca2+ channels and ATP-dependent Ca2+ pumps in the plasmalemma and sarcoplasmic reticulum (SR). Ca2+ entry through the plasmalemma is apparently mediated by four different pathways: leak; receptor-operated Ca2+ channels; potential sensitive Ca2+ channels; and stretch-activated channels. The agonist releasable intracellular Ca2+ store appears to be identical with the SR. Evidence for the involvement of Ca2+-induced Ca2+ release and inositol-1,4,5-trisphosphate in the release of SR Ca2+ is discussed. Smooth muscle contractions induced by certain agonists may be further enhanced by inhibition of Ca2+ uptake by the SR and of active Ca2+ extrusion across the plasmalemma. At the moment it is not clear from a consideration of the Ca2+ regulatory mechanisms present in vascular smooth muscle how dietary Ca2+ affects vascular tone. The increased Ca2+ permeation through smooth muscle cell membranes of resistance arteries taken from spontaneously hypertensive rats may be relevant to this problem.     

Phospholipid composition and metabolism in mouse muscular dystrophy

1. The uptake of monosaccharides and polyols in the obligatory aerobic yeast Rhodotorula gracilis (glutinis) was accompanied by proton uptake. 2. The half-saturation constant of transport, KT, depended on pH, changing from about 2mM at pH 4.5 to 80mM at pH8.5 for D-xylose; this change of the effective carrier affinity was reversible. 3. The apparent dissociation constant of the monosaccharide carrier was estimated at pKa 6.75. 4. At pH8.5, when the pH gradient across the cell membrane vanished, no sugar accumulation was demonstrable. 5. The half-saturation constants of sugar uptake and H+ co-transport were very similar to each other, the latter obviously being controlled by the former. 6. The H+/sugar stoicheiometry remained constant under various physiological conditions; it amounted to one H+ ion per sugar molecule taken up. 7. The data are interpreted as a strong piece of evidence in favour of the active monosaccharide transport in R. gracilis (glutinis) being an H+-symport energized by the electrochemical gradient of H+ across the plasma membrane of the yeast.     

Calcium Transport in Protoplasts Isolated from ml-o Barley Isolines Resistant and Susceptible to Powdery Mildew

Free cytoplasmic calcium has been postulated to play a role in preventing powdery mildew in a series of homozygous ml-o mutants of barley, Hordeum vulgare L. Protoplasts isolated from 7-day-old plants of the ml-o resistant-susceptible (R-S) barley isolines, Riso 5678/3^* × Carlsberg II R and S, were used to test for differences in fluxes of Ca²⁺ across the plasmalemma. Greater influx or lesser efflux might account for a higher free cytosolic Ca²⁺ postulated to exist in ml-o R mutants. Uniform patterns of uptake were maintained for 3 hours from solutions of 0.2 and 2 millimolar Ca²⁺. Washout curves of ⁴⁵Ca²⁺ from R and S protoplasts revealed three compartments—presumed to represent release from the vacuole, organelles, and the cytoplasm (which included bound as well as free Ca²⁺). Uptake and washout did not differ between isolines. On the basis of recent determinations of submicromolar levels of free cytoplasmic Ca²⁺ and our initial rates of ⁴⁵Ca-labeled Ca²⁺ uptake, we show that measurement of the unidirectional influx of Ca²⁺ across the plasmalemma is not feasible because the specific activity of the pool of free cytoplasmic calcium increases almost instantaneously to a level that would result in a significant, but unknown, efflux of label. Similarly, measurement of the efflux of Ca²⁺ across the plasmalemma is not possible since the activity of the pool of free cytoplasmic calcium is a factor of 350 smaller than the most rapid component of the washout experiment. This pool of cytoplasmic free Ca²⁺ will wash out too rapidly and be too small to detect under the conditions of these experiments.     

Some Ionic and Bioelectric Properties of the Ameba Chaos chaos

Ionic relationships in the giant ameba Chaos chaos were studied by analyzing bulk preparations of ground cytoplasm for K, Na, and Cl. Ion levels under normal conditions were compared with the levels in cells exposed to varying concentrations of different ions, for varying times and at different temperatures. By standard intracellular electrode techniques, the bioelectric potential, electrical resistance, and rectifying properties of the plasmalemma were studied on intact cells in media of different composition. The results obtained, when related to evidence from other studies on ion fluxes and osmotic relationships, suggest the following concept of ionic regulation in Chaos chaos. In the absence of active membrane uptake, the plasmalemma is essentially impermeable to anions but permeable to both K and Na, which enter passively. In the cold the cell does not discriminate between K and Na, the cytoplasmic level of K + Na is determined by a Donnan distribution, and osmotic imbalance leads to slow swelling. At normal temperatures active processes are added: Na and water are pumped out by the contractile vacuole system; Cl is accumulated, along with the colloid components of the cytoplasm, only during feeding and growth, which depend upon membrane uptake and intracellular membrane transformations. There is no evidence for active transport of any ion species directly across the plasmalemma.     

Influence of internal sugar levels on apoplasmic retrieval of exogenous sucrose in source leaf tissue

Sugar levels in Beta vulgaris leaves were increased by heat-girdling the petiole and returning the plant to the controlled-environment chamber for 10 and 34 hours. After 10 hours, sucrose influx into the treated leaves was similar to the controls, although sucrose levels increased from 2.1 to 5.3 micromoles per milligram chlorophyll. However, after a 34-hour treatment, sucrose levels increased from 2.1 to 11.5 micromoles per milligram chlorophyll. In this instance, sucrose influx decreased relative to the untreated controls. Decreasing sugar levels by DCMU treatment resulted in a small stimulation of sucrose influx. A similar DCMU treatment applied to leaves of Allium cepa also resulted in an increase in sucrose influx. However, in A. cepa we could not attribute this increase to a lowering of sugar levels, as the kinetic profiles obtained from control leaves did not vary from each other throughout the day, despite considerable changes in sugar levels. Additionally, it appeared that sucrose uptake in onion may be set at some point and remains invariant throughout the day. Similar studies were also conducted on discs cut from mature leaves of Spinacia oleracea var America. Between 1 and 8 hours after the onset of the photoperiod, the sucrose content of the spinach leaves increased from 2.6 to 9.3 micromoles per milligram chlorophyll. A comparison of the kinetic profiles obtained from leaf discs, taken at these times, indicated that sucrose uptake was not influenced by these changes in internal sugar levels. The relationship between the above findings and `trans' inhibition of exogenous sucrose uptake is discussed. Although intermediate changes in sugar levels in sugar beet leaves did not appear to affect sucrose influx, autoradiographic studies revealed that these changes dramatically affected the partitioning of exogenously supplied [¹⁴C]sucrose. Our results indicate that while intermediate changes in internal sugar levels have little effect on sucrose influx across the plasmalemma, they may dramatically affect partitioning between the phloem and the mesophyll vacuole.     

Sugar Selectivity and Other Characteristics of Phloem Loading in Beta vulgaris L

The rate of phloem loading, its selectivity, and the disposition of labeled carbon were studied following application of (14)C-labeled sugars to the free space of source leaves of sugar beet (Beta vulgaris L.). Buffered 10 mm solutions of (14)C-labeled sucrose, fructose, stachyose, mannitol, 3-0-methyl glucose or l-glucose were applied to the abraded epidermis of source leaves held in the dark. Distribution of the labeled carbon from sugar taken up from the free space was studied by micro-densitometry of autoradiographs. Uptake of labeled sugar from the free space, partition between mesophyll and minor veins, metabolic conversions, export and respiration were followed during the 3-hr time course studies. Rates of sugar uptake into the minor veins, flux rates through the sieve element-companion cell complex membrane and concentration ratios between free space and the interior of the minor vein phloem cells were compared for the six sugars studied for evidence of active uptake. The composition of the free space solution in leaves photosynthesizing in (14)CO(2) was studied by vacuum infiltration of the source leaf air spaces and removal of the solution by centrifugation. Labeled compounds in this solution were compared to those in an aqueous ethanol extract of the same leaf pieces.The results in sugar beet source leaves support the concept of direct, active uptake of sucrose from free space into minor veins. This is not the case for fructose, 3-0-methyl glucose, mannitol, or stachyose. The latter two sugars, which are translocated in some plants, are not loaded into the minor veins at a rate sufficient to make them a significant component of the material translocated. The rate of phloem loading is controlled in part by mesophyll metabolism, especially as it affects the availability of sucrose to the free space. Both the rate and selectivity of export are controlled by uptake from the free space into the sieve element-companion cell complex of the minor veins.     

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.     

Ion fluxes and phytochrome protons in mung bean hypocotyl segments: I. Fluxes of potassium

K⁺ [⁸⁶Rb⁺] uptake by Phaseolus aureus Roxb. hypocotyl segments cut immediately below the hook is inhibited by the active form of phytochrome (Pfr). Short load-short wash experiments indicate that the inhibition of uptake occurs across the plasmalemma. A maximal inhibition of short term uptake occurs in 10 to 50 millimolar KCI. Low temperature had only a small effect on influx and the inhibition of influx from 50 millimolar KCI. A consideration of the electrochemical gradient for K⁺ suggests that passive K⁺ fluxes may predominate under these conditions. Red light induces small depolarizations of membrane potential in subhook cells. Far red light antagonizes this effect. Pfr inhibits efflux of K⁺[⁸⁶Rb⁺] from subhook segments. This effect is also relatively insensitive to low temperature. This inhibition of efflux may reflect inhibition of a K⁺ -K⁺ exchange process, or reduced passive permeability of the plasmalemma to K⁺. In contrast, Pfr enhances short term uptake of K⁺[⁸⁶Rb⁺] in apical hypocotyl hook segments of Phaseolus aureus Roxb. Short load-short wash experiments indicate that fluxes across the plasmalemma are modified by Pfr. A maximal enhancement of short term influx occurs in 50 millimolar KCI. Influx and the red light enhancement of influx from 50 millimolar KCI are relatively insensitive to low temperature. Pfr also enhances efflux of K⁺[⁸⁶Rb⁺] from preloaded apical hook segments. This increased influx may reflect enhancement of a K⁺ -K⁺ exchange process or increased passive permeability of the plasmalemma to K⁺.     

Characterization of Paraquat Transport in Protoplasts from Maize (Zea mays L.) Suspension Cells

Protoplasts isolated from maize (Zea mays L.) suspension cells were used to study transport of paraquat. [14C]Paraquat uptake was measured in 400-[mu]L centrifuge tubes using silicon oil centrifugation techniques. Approximately 50% of accumulation from a 100 [mu]M paraquat solution occurred in the first 10 s, and net accumulation reached a maximum after about 10 min. Membrane binding accounted for about 30% of apparent accumulation. Concentration-dependent uptake kinetics were characterized by a non-saturating curve, which was resolved into a linear and a saturable component. The Km of the saturable component was 132 [mu]M, and the Vmax was 0.512 nmol [mu]L of protoplasts-1 min-1. In the absence of sucrose, the Vmax of the saturable component was reduced by 52%, suggesting that paraquat uptake across the plasmalemma is energy dependent. Measurement of concentration-dependent binding of paraquat to burst protoplasts showed a linear response. This suggests that the linear component from intact protoplast concentration kinetics represented paraquat binding to the plasmalemma surface. Calcium inhibited the saturable component, and this inhibition was shown by Lineweaver-Burk analysis to be noncompetitive. Putrescine, a divalent cationic polyamine with a charge distribution similar to that of paraquat, competitively inhibited paraquat uptake. These results show that paraquat transport characteristics at the plasmalemma of maize protoplasts are similar to those reported earlier for paraquat transport in roots of intact maize seedlings.     

The effect of CCCP on ion fluxes in the stele and cortex of maize roots

Summary The accumulation of ⁸⁶Rb labelled potassium by isolated stelar and cortical tissues from 7-day-old roots of Zea mays has been compared with the levels accumulated by these tissues in the intact root. Cortical tissues have similar uptake eapacities in these two conditions whereas stelar tissues only exhibit an uptake capacity in the intact root system. The uncoupler carbonylcyanide m-chlorophenylhydrazone caused a considerable decrease in the uptake of potassium by these tissues. In the intact root system it prevented ions from the bathing medium reaching the stelar tissues. The efflux pattern from preloaded isolated stelar and cortical tissues was considerably altered by the inhibitor, a promotion of the efflux occurring in both of these tissues.It is concluded that stelar tissues only accumulated ions when these are supplied through the root symplasm and that the stelar plasmalemma has only a limited uptake capacity per se. Stelar uptake is thus a reflection of vacuolar accumulation across the tonoplast. There is no evidence in the present study of a carrier-mediated active secretion of ions across the stelar plasmalemma. The fact that the efflux was promoted rather than depressed by the uncoupler supports the postulate that a passive leakage is the final stage in the transport of ions across the plant root.     

The regulation of sugar uptake and accumulation in bean pod tissue

The identity, localization and physiological significance of enzymes involved in sugar uptake and accumulation were determined for endocarp tissue of pods of Kentucky Wonder pole beans (Phaseolus vulgaris). An intracellular, alkaline invertase (pH optimum, 8) was assayed in extracted protein, as well as enzymes involved in sucrose synthesis, namely, uridinediphosphate (UDP-glucose pyrophosphorylase and UDP-glucose-fructose transglucosylase). Indirect evidence indicated the presence also of hexokinase, phosphohexoseisomerase and phosphoglucomutase. The data suggested that sucrose synthesis occurred in the cytoplasm, and that both sugar storage and an alkaline invertase occurred in the vacuole. The latter functions to hydrolyze accumulated sucrose. An outer space invertase (pH optimum, 4.0) was detected, but was variable in occurrence. Although its activity at the cell surface enhanced sucrose uptake, sucrose may be taken up unaltered.

Over a wide range of concentrations of exogenous glucose the sucrose/reducing sugar ratio of accumulated sugars remained unchanged at about 20. Synthesis of sucrose appears to be requisite to initial accumulation from glucose or fructose, as free hexoses do not increase at the apparent saturating concentration for uptake. Sucrose accumulation from exogenous hexose represents a steady-state value, in which sucrose is transported across the tonoplast into the vacuole at a rate equivalent to its rate of synthesis. Evidence indicates that this component of the accumulation process involves active transport of sucrose against a concentration gradient. The ratio of sucrose/reducing sugars in the accumulated sugars immediately after a period of uptake was inversely related to the level of inner space invertase. Within 16 hours after a period of accumulation, practically all of the sugar occurs as glucose and fructose.

The absence of competition among hexoses and sucrose indicated that a common carrier was not involved in their uptake. From a series of studies on the kinetics of uptake of glucose and fructose, including competition studies, the effects of inhibitors, radioactive assay of accumulated sugars and the distribution of label in accumulated sucrose it appeared that rate limitation for glucose or fructose uptake resides in the sequence of reactions leading to sucrose synthesis, rather than in a process mediated by a carrier protein.

     

Uptake of ammonium and nitrate by carob (Ceratonia siliqua) as affected by root temperature and inhibitors

Seedlings of carob ( Ceratonia siliqua L. cv. Mulata) were grown in nutrient solution culture for 5 weeks, with or without nitrogen at different root temperatures (10, 16, 22, 30, 35 or 40deg;C) and with the air temperature kept between 20 and 24°C. The nitrogen was given as either ammonium or nitrate. At all root temperatures studied, nitrogen-depleted plants developed higher net uptake rates for nitrogen than plants grown in the presence of nitrogen. Temperature affected the kinetic parameters of nitrate uptake more than those of ammonium uptake. With increasing root temperature, the K_m of ammonium uptake decreased, but to a lesser extent than the K_m for nitrate. The increase in V_max of ammonium uptake with temperature was also less noticeable than that for nitrate uptake. Ammonium and nitrate uptakes were inhibited in a similar way by respiratory or protein synthesis inhibitors. It may be noted that ammonium uptake in the presence of inhibitors at 40°C was higher than uptake at 10°C without inhibitors. Some similarities between the transport mechanisms for nitrate and ammonium are underlined in the present work. Components of both transport systems displayed saturation kinetics and depended on protein synthesis and energy. The following components of nitrate uptake were distinguished: (a) a passive net influx into the apparent free space; (b) a constitutive active uptake and (c) active uptake dependent on protein synthesis. We may similarly define three ammonium uptake systems: (a) a passive influx into the apparent free space; (b) passive diffusion uptake at high temperature and (c) active uptake dependent on protein synthesis. The possible role of the ratio between mechanism (c) and mechanism (b) as determinant of ammonium sensitivity is discussed.     

Sulphate Influx in Characean Cells: I. GENERAL CHARACTERISTICS

There is a significant sulphate influx in Chara australis, Nitellatranslucens, and Tolypella intricata but not in Nitellopsisobtusa. The uptake is active in Chara, the ions moving intothe vacuole against an electrochemical potential gradient. Influxrate in the light remains constant whereas a transient increasein rate occurs upon transfer to the dark. Both the light andthe dark influx rate respond similarly to temperature, and toexternal sulphate ion concentration. It is proposed that sulphateis pumped actively across the plasmalemma and that the increasein influx rate following transfer to the dark reflects a transientincrease in the rate at which the pump operates. There is anapparent light stimulation of the tonoplast flux.