Red Beet Storage Tissue Research Articles

Effects of Prolonged Washing on Primary and Secondary Transport Processes at the Plasma Membrane in Red Beet Storage Tissue.

Changing patterns of enzyme activity and solute transport in response to washing were investigated in red beet (Beta vulgaris L.) storage tissue. Washing had a pronounced effect on the plasma membrane (PM) H+-ATPase with an increase in both hydrolytic and proton-pumping activities. Immunoblotting indicated that this may be due, in part, to a higher amount of this enzyme in the PM of washed tissue. Activities of the tonoplast (V)H+-ATPase and pyrophosphatase fluctuated during a 4-d washing period, but overall showed no marked change in activity. In tissue discs sucrose (Suc), glucose (Glc), and fructose uptakes increased significantly in response to washing. Cycloheximide, cordycepin, and tunicamycin inhibited both Glc- and Suc-inducible uptake. Monensin also strongly inhibited inducible Glc uptake, but the effect on Suc was less marked. N-Ethylmaleimide inhibited both Suc and Glc uptake, with its effects being more pronounced in fresh tissue. Other protein-modifying reagents showed no significant difference in their level of inhibition between fresh and washed tissue. Transport studies, carried out using energized PM vesicles from fresh and washed tissue, indicated that there was no rise in Suc and Glc uptake rates in response to washing. Results with a range of inhibitors indicated that there was no marked change in transporter sensitivity in vesicles isolated from fresh and washed tissue. The results indicate that the well-described enhancement of solute transport in washed storage tissue may be due to an increased PM H+-ATPase activity rather than to changes in PM carrier activity or to changes in metabolism such as invertase activity.

The Vacuolar ATPase of Red Beet Storage Tissue: Electron Microscopic Demonstration of the “Head‐and‐Stalk” Structure*

AbstractTonoplast membranes were prepared from tissue homogenates and from vacuoles isolated from beetroot storage tissue (Beta vulgaris L., ssp. conditiva) for transmission electron microscopic analysis of the structure of the beetroot vacuolar ATPase using the negative staining technique. By comparison of the specific inhibitor sensitivities of the ATPase activity, i.e. ATP hydrolysis and H+‐pumping, the purity of the tonoplast preparations with respect to contamination with mitochondrial inner membranes was assessed to avoid confusion with mitochondrial F1F0‐ATPase. Membranes prepared in Hepes/Tris or BTP/Mes‐containing media rarely showed typical head‐and‐stalk structures although characteristic nitrate‐ and bafilomycin A1‐sensitive ATP‐hydrolysis and H+‐pumping could be measured. However, typical head‐and‐stalk structures were observed regularly when these buffers were replaced by K‐phosphate buffer. Under these conditions, the beetroot vacuolar ATPase is characterized by a large head group with a central cleft, a thin stalk, connecting it to the membrane and by basal components projecting from the base of the stalks near the vacuolar membrane and forming a distinct layer of electron‐light particles between the vacuolar membrane and the layer of non‐stained head groups.

Inhibition and Ultraviolet-Induced Chemical Modification of UDP-Glucose:(1,3)-β-Glucan (Callose) Synthase by Chlorpromazine

UDP-glucose:(1,3)-beta-glucan (callose) synthase (CS) from storage tissue of red beet (Beta vulgaris L.) was strongly inhibited by the phenothiazine drug chlorpromazine (CPZ). In the absence of ultraviolet irradiation, CPZ was a noncompetitive inhibitor with 50% inhibitory concentration values for plasma membrane and solubilized CS of 100 and 90 mum, respectively. Both the Ca(2+)- and Mg(2+)- stimulated components of CS activity were affected. CPZ inhibition was partially alleviated at saturating levels of Ca(2+), but not Mg(2+), suggesting that CPZ interferes with the Ca(2+)-binding site of CS. Binding experiments with [(14)C]CPZ, however, showed strong non-specific partitioning of CPZ into the plasma membrane, providing evidence that perturbation of the membrane environment is probably the predominant mode of inhibition. Ultraviolet irradiation at 254 nm markedly enhanced CPZ inhibition, with complete activity loss following exposure to 4 mum CPZ for 2 min. Inhibition followed a pseudo-first order mechanism with at least three CPZ binding sites per CS complex. Under these conditions, [(3)H]CPZ was covalently incorporated into plasma membrane preparations by a free radical mechanism; however, polypeptide labeling profiles showed labeling to be largely nonspecific, with many polypeptides labeled even at [(3)H]CPZ levels as low as 1 mum, and with boiled membranes. Although CPZ is one of the most potent known inhibitors of CS, its use as a photolabel will require a homogeneous CS complex or establishment of conditions that protect against the interaction of CPZ with specific binding sites located on various polypeptide components of the CS complex.

Syringomycin-Stimulated Phosphorylation of the Plasma Membrane H+-ATPase from Red Beet Storage Tissue

The syringomycin-stimulated in vitro protein phosphorylation of the plasma membrane H(+)-ATPase of red beet (Beta vulgaris L.) storage tissue was investigated. Peptides representing the H(+)-ATPase N and C termini and nucleotide binding site (P-2, P-3, and P-1, respectively) were synthesized, and rabbit antisera against each were produced. In western immunoblots of purified plasma membranes, these antisera immunoreacted with the 100-kilodalton polypeptide of the H(+)-ATPase and with other smaller polypeptides. The smaller polypeptides appeared to be degraded forms of the intact 100-kilodalton polypeptide. Immunoprecipitation experiments showed that plasma membranes treated with syringomycin had increased protein phosphorylation rates of the 100-kilodalton polypeptide. Optimal phosphorylation levels were achieved with 25 micromolar free Ca(2+). Phosphoserine and phosphothreonine were detected in the immunoprecipitates. Washed immunoprecipitates generated with anti-P-1 possessed protein phosphorylation activity. This immunoprecipitate activity was not stimulated by syringomycin, but it was inhibited when plasma membranes were treated with sodium deoxycholate before immunoprecipitation. The findings show that syringomycin stimulates the phosphorylation of the plasma membrane H(+)-ATPase and that specific protein kinase(s) are probably associated with the enzyme.

Rapid Enrichment of CHAPS-Solubilized UDP-Glucose: (1,3)-β-Glucan (Callose) Synthase from Beta vulgaris L. by Product Entrapment

Rapid enrichment of CHAPS-solubilized UDP-glucose:(1,3)-beta-glucan (callose) synthase from storage tissue of red beet (Beta vulgaris L.) is obtained when the preparation is incubated with an enzyme assay mixture, then centrifuged and the enzyme released from the callose pellet with a buffer containing EDTA and CHAPS (20-fold purification relative to microsomes). When centrifuged at high speed (80,000g), the enzyme can also be pelleted in the absence of substrate (UDP-Glc) or synthesis of callose, due to nonspecific aggregation of proteins caused by excess cations and insufficient detergent in the assay buffer. True time-dependent and substrate-dependent product-entrapment of callose synthase is obtained by low-speed centrifugation (7,000-11,000g) of enzyme incubated in reaction mixtures containing low levels of cations (0.5 millimolar Mg(2+), 1 millimolar Ca(2+)) and sufficient detergent (0.02% digitonin, 0.12% CHAPS), together with cellobiose, buffer, and UDP-Glc. Entrapment conditions, therefore, are a compromise between preventing nonspecific precipitation of proteins and permitting sufficient enzyme activity for callose synthesis. Further enrichment of the enzyme released from the callose pellet was not obtained by rate-zonal glycerol gradient centrifugation, although its sedimentation rate was greatly enhanced by inclusion of divalent cations in the gradient. Preparations were markedly cleaner when product-entrapment was conducted on enzyme solubilized from plasma membranes isolated by aqueous two-phase partitioning rather than by gradient centrifugation. Product-entrapped preparations consistently contained polypeptides or groups of closely-migrating polypeptides at molecular masses of 92, 83, 70, 57, 43, 35, 31/29, and 27 kilodaltons. This polypeptide profile is in accordance with the findings of other callose synthase enrichment studies using a variety of tissue sources, and is consistent with the existence of a multi-subunit enzyme complex.

Purification and characterization of the proton translocating plasma membrane ATPase of red beet storage tissue

Plasma membranes were prepared from red beet ( Beta vulgaris L.) storage tissue by partition in an aqueous two-phase system. A highly active proton-translocating ATPase was purified from these membranes by lysophosphatidylcholine extraction and glycerol density gradient centrifugation. The ATPase activity was inhibited by vanadate or dicyclohexyl carbodiimide, but was insensitive to azide, nitrate and molybdate at concentrations which inhibit the F 1ATPase, the tonoplast ATPase, and acid phosphatase. Inhibition by vanadate was consistent with a non-competitive mechanism, with K i = 10 μ M . The K m for Mg-ATP was about 1 mM, magnesium ions were required, and the activity was stimulated by KCI and by lysophosphatidylcholine. The optimal pH was 6.5. The molecular mass by gel filtration in the presence of 2 g/liter octyl glucoside was 600 kDa, while dodecyl sulfate gel electrophoresis gave a polypeptide molecular mass of 100 kDa. After blotting onto nitrocellulose, the purified enzyme did not bind concanavalin A, although a concanavalin A-binding peptide of the plasma membrane runs to nearly the same position on the gel and showed some tendency to co-purify with the ATPase. Phospholipid vesicles into which the purified ATPase had been incorporated by the freeze-thaw technique showed vanadate-sensitive, ATP-dependent proton uptake. When the ATPase was reconstituted into lipid membranes at high protein to lipid ratios and incubated with ATP, two-dimensionally crystalline arrays of protein molecules were formed.

Separation and Immunological Characterization of Membrane Fractions from Barley Roots

Tonoplast and plasma membranes (PM) were isolated from barley roots (Hordeum vulgare L. cv California Mariout 72) using sucrose step gradients. The isolation procedure yielded sufficient quantities of PM and tonoplast vesicles that were sealed and of the right orientation to measure ATP-dependent proton transport in vitro. The proteins of the endoplasmic reticulum, tonoplast-plus-Golgi membrane (TG) and PM fractions were separated on sodium dodecyl sulfate gels, and immunoblots were used to test for cross-contamination between the fractions. Proteins that cross-reacted with antibodies to the PM ATPase from corn roots and Neurospora were greatly enriched in the PM fraction, as were proteins that cross-reacted with monoclonal antibodies to an arabinogalactan protein from the PM of tobacco cells. Proteins that cross-reacted with antibodies to the 58- and 72-kilodalton subunits of the tonoplast ATPase of red beet storage tissue were greatly enriched in the TG fraction. The results with immunoblots and enzyme assays indicated that there was little cross-contamination between the tonoplast and PM vesicles. The molecular weights and isoelectric points of the PM ATPase and the tonoplast ATPase subunits were also determined using immunoblots of two-dimensional gels of the PM and TG proteins.

Bacterial phytotoxin, syringomycin, induces a protein kinase-mediated phosphorylation of red beet plasma membrane polypeptides

Syringomycin, a peptide toxin and a virulence factor produced by the bacterial phytopathogen Pseudomonas syringae pv. syringae, stimulated the phosphorylation of several plasma membrane polypeptides of red beet storage tissue. Among these was a 100-kDa polypeptide, which corresponds in size to the proton pump ATPase. The phosphorylations were insensitive to hydroxylamine, indicating that the polypeptide phosphorylated intermediates involved phosphate ester bonds characteristic of protein kinase-mediated phosphorylation. Phosphorylation of the 100-kDa polypeptide and of most of the other polypeptides was reduced or eliminated by extraction of the membranes with 0.1% (wt/vol) sodium deoxycholate, a treatment that also eliminated the ability of the toxin to stimulate ATPase activity. Phosphorylation of the 100-kDa polypeptide was highest with 10-20 mug of syringomycin; the same amounts gave the highest degree of ATPase activity stimulation. Phosphorylation of some of the polypeptides was eliminated or decreased by the Ca(2+) chelator EGTA. Addition of excess Ca(2+) restored the phosphorylation of most of these polypeptides. We conclude that syringomycin acts by stimulating an endogenous membrane protein kinase activity, which results in the phosphorylation of several membrane polypeptides. One of the phosphorylated polypeptides corresponds in size to the proton pump ATPase.

Sucrose Phosphatase Associated with Vacuole Preparations from Red Beet, Sugar Beet, and Immature Sugarcane Stem

The specific phosphatase, sucrose phosphate phosphohydrolase (sucrose phosphatase, EC 3.1.3.24) was present in vacuole preparations from storage tissue of red beet (Beta vulgaris L.), sugar beet (Beta vulgaris L. cultivar Kawemono), and immature sugarcane (Saccharum spp. hybrid, cultivar NCO 310). In red beet vacuole preparations the specific activity of sucrose phosphatase, using the naturally occurring vacuole marker, betanin, as reference, was higher than the specific activity of cytoplasmic markers, phosphoenolpyruvate carboxylase and glucose 6-phosphate dehydrogenase, suggesting that sucrose phosphatase is associated with the vacuoles. High speed centrifugation of lysed vacuoles did not result in precipitation of the enzyme indicating that the enzyme is not tightly bound to the tonoplast. Sucrose phosphatase was more sensitive to inhibition by sodium vanadate and less sensitive to ammonium molybdate than was the nonspecific phosphatase which was also present in the extracts. Sucrose phosphatase might be part of the group translocator proposed recently to operate in the tonoplast of sugarcane and red beet.

H+-ATPase Activity from Storage Tissue of Beta vulgaris

The molecular weight and isoelectric point of the plasma membrane H(+)-ATPase from red beet storage tissue were determined using N,N'-dicyclohexylcarbodiimide (DCCD) and a H(+)-ATPase antibody. When plasma membrane vesicles were incubated with 20 micromolar [(14)C]-DCCD at 0 degrees C, a single 97,000 dalton protein was visualized on a fluorograph of a sodium dodecyl sulfate polyacrylamide gel. A close correlation between [(14)C]DCCD labeling of the 97,000 dalton protein and the extent of ATPase inhibition over a range of DCCD concentration suggests that this 97,000 dalton protein is a component of the plasma membrane H(+)-ATPase. An antibody raised against the plasma membrane H(+)-ATPase of Neurospora crassa cross-reacted with the 97,000 dalton DCCD-binding protein, further supporting the identity of this protein. Immunoblots of two-dimensional gels of red beet plasma membrane vesicles indicated the isoelectric point of the H(+)-ATPase to be 6.5.

Mechanism of Action of Pseudomonas syringae Phytotoxin, Syringomycin

Syringomycin, a peptide toxin produced by the phytopathogen Pseudomonas syringae pv syringae preferentially stimulated (2-fold) the vanadate-sensitive ATPase activity associated with the plasma membrane of red beet storage tissue. The toxin had a very slight effect on the tonoplast ATPase and had no detectable effect on the mitochondrial ATPase. Optimal stimulation was achieved with 10 to 50 micrograms of syringomycin per 25 micrograms of membrane protein. Treatment of membranes with 0.1% (weight/volume) deoxycholate eliminated the activation effect, and enzyme solubilized with Zwittergent 3-14 was not affected by syringomycin. ATPase activity was activated to the same extent at KCl concentrations ranging from 0 to 50 millimolar. Valinomycin, nigericin, carbonylcyanide p-trifluoromethoxyphenylhydrazone, and gramicidin did not increase the plasma membrane ATPase activity. However, these ionophores did not hinder the ability of syringomycin to stimulate the activity. We suggest that syringomycin does not increase ATPase activity by altering membrane ion gradients nor directly interacting with the enzyme, but possibly through regulatory effectors or covalent modification of the enzyme.

Kinetics of Ca2+/H+ Antiport in Isolated Tonoplast Vesicles from Storage Tissue of Beta vulgaris L.

Artificial pH gradients across tonoplast vesicles isolated from storage tissue of red beet (Beta vulgaris L.) were used to study the kinetics of a Ca(2+)/H(+) antiport across this membrane. Ca(2+)-dependent H(+) fluxes were measured by the pH-dependent fluorescence quenching of acridine orange. DeltapH-dependent Ca(2+) influx was measured radiometrically. Both H(+) efflux and Ca(2+) influx displayed saturation kinetics and an identical dependence on external calcium with apparent K(m) values of 43.9 and 41.7 micromolar, respectively. Calcium influx was unaffected by an excess of Mg(2+) but was inhibited by La(3+) > Mn(2+) > Cd(2+). The apparent K(m) for external calcium was greatly affected (5-fold) by internal pH in the range of 6.0 to 6.5 and a transmembrane effect of internal proton binding on the affinity for external calcium is suggested.

Electrogenic Proton Translocation by the Adenosine Triphosphatase of Intact Vacuoles Isolated from Beet ( Beta vulgaris L.)

Summary Changes in the transtonoplast pH gradient and the electrical membrane potential have been monitored in parallel on addition of ATP to intact vacuoles isolated from storage tissue of red beet ( Beta vulgaris L.). Proton translocation by the ATPase results in a positive displacement of the membrane potential by about 7 mV (estimated using a microelectrode-calibration of a potentiometric fluorescent dye), with no detectable pH change. When compensatory K + movement across the tonoplast was permitted by the presence of valinomycin there was a rapid ATP-driven acidification of the vacuole with an initial rate of about 0.1 pH unit·min −1 . The restraining influence of the pH gradient on the ATP-dependent change in membrane potential was apparent when the vacuole was acidified by an addition of nigericin.

Na+/H+ Antiport in Isolated Tonoplast Vesicles from Storage Tissue of Beta vulgaris

The pH-dependent fluorescence quenching of acridine orange was used to study the Na(+)- and K(+)-dependent H(+) fluxes in tonoplast vesicles isolated from storage tissue of red beet and sugar beet (Beta vulgaris L.). The Na(+)-dependent H(+) flux across the tonoplast membrane could be resolved into two components: (a) a membrane potential-mediated flux through conductive pathways; and (b) an electroneutral flux which showed Michaelis-Menten kinetics relationship to Na(+) concentration and was competitively inhibited by amiloride (K(i) = 0.1 millimolar). The potential-dependent component of H(+) flux showed an approximately linear dependence on Na(+) concentration. In contrast, the K(+)-dependent H(+) flux apparently consisted of a single component which showed an approximately linear dependence on K(+) concentration, and was insensitive to amiloride. Based on the Na(+)- and K(+)-dependent H(+) fluxes, the passive permeability of the vesicle preparation to Na(+) was about half of that to K(+).The apparent K(m) for Na(+) of the electroneutral Na(+)/H(+) exchange varied by more than 3-fold (7.5-26.5 millimolar) when the internal and external pH values were changed in parallel. The results suggest a simple kinetic model for the operation of the Na(+)/H(+) antiport which can account for the estimated in vivo accumulation ratio for Na(+) into the vacuole.

Proton-Translocating Inorganic Pyrophosphatase in Red Beet (Beta vulgaris L.) Tonoplast Vesicles

The substrate and ionic requirements of ATP and inorganic pyrophosphate (PPi) hydrolysis by tonoplast vesicles isolated from storage tissue of red beet (Beta vulgaris L.) were compared with the requirements of ATP-and PPi-dependent proton translocation by the same material. Both ATP hydrolysis and ATP-dependent proton translocation are most stimulated by Cl(-) and inhibited by NO(3) (-). NaCl and KCl support similar rates of ATP hydrolysis and ATP-dependent proton translocation while K(2)SO(4) supports lesser rates for both. PPi hydrolysis and PPi-dependent proton translocation are most stimulated by K(+). KCl and K(2)SO(4) support similar rates of PPi hydrolysis and PPi-dependent proton translocation but NaCl has only a small stimulatory effect on both. Since PPi does not inhibit ATP hydrolysis and ATP does not interfere with PPi hydrolysis, it is inferred that the two phosphohydrolase and proton translocation activities are mediated by different tonoplast-associated enzymes. The results indicate the presence of an energy-conserving proton-translocating pyrophosphatase in the tonoplast of red beet.

A Comparison of Methods for Measuring Turgor Pressures and Osmotic Pressures of Cells of Red Beet Storage Tissue

Two methods for measuring the turgor pressures of cells in discs of storage tissue of red beet (Beta vulgaris L.) were compared, and a centrifugation method for extracting sap from frozen and thawed tissue was evaluated. Turgor pressures were measured directly using a pressure probe, or indirectly using a vapour pressure osmometer. With the latter, discs were placed directly in the osmometer chamber and turgor was calculated as the difference in osmotic pressure before and after freezing and thawing. Turgor in freshly cut discs, measured with the pressure probe, was between 0 012 MPa and 0-118 MPa with a mean ±s.d. of 0 092 + 0-032 MPa (n = 24). That measured with the osmometer varied between 0-08 MPa and 0-12 MPa with a mean ±s.d. of 0-09 ±0-10 MPa(n = 54). After vacuum infiltration of discs with distilled water, the turgor measured with the pressure probe increased to 1-05 1-12 MPa. Turgor measured with the osmometer also increased after vacuum infiltration but was, on average, 12% lower than that measured with the pressure probe. Overall, the results suggest that for routine measurements, the osmometer can provide reasonable estimates of the turgor of cells in beet discs. This is because a number of factors that, potentially, could interfere with this method have only a small effect in this tissue. None of the measured turgors is indicative of that occurring in intact storage roots because both excision and vacuum infiltration of discs alter the concentrations of solutes in the extracellular space. The osmotic pressure of sap extracted by centrifugation from frozen and thawed discs was not significantly different from that measured by placing frozen and thawed discs directly in the osmometer. Solute concentrations in the sap were not significantly different from those measured by chemical extraction of discs.

Occurrence of Localized Intense Cation Uptake Sites in the Vascular Rings of Red Beet Storage Tissue 2

Development and decline of cation uptake capacity in discs taken from the vascular and parenchyma rings of storage tissue of red table beet (Beta vulgaris L.) were observed during 12 d of ageing. Uptake capacity for Na+ and Rb+ showed a steady rise reaching maximums by the fourth to fifth days of ageing. Thereafter, there was a steady decline in the uptake rates. Vascular ring tissues were able to develop a greater uptake capacity for both Na+ and Rb+ than the tissues of parenchyma rings. This difference, which was more pronounced for Rb+ than for Na+ uptake, is attributed to a combination of variations in cell density and differences in the acquisition and retention of the cation uptake capacity. Respiration of tissue discs showed no significant rise during ageing, nor were there significant differences in the respiration of vascular and parenchyma tissues. Vascular tissues contained significantly more betacyanin than parenchyma tissues; and they retained their pigment, as well as their acquired cation uptake capacity, for a longer period during the ageing process.

Evidence for a β-Aspartyl Phosphate Residue in the Phosphorylated Intermediate of the Red Beet Plasma Membrane ATPase

A borohydride reduction method was used to identify the phosphorylated amino acid in the phospho-enzyme of the red beet (Beta vulgaris L.) plasma membrane ATPase. Plasma membrane fractions were phosphorylated with unlabeled ATP in the presence of MgSO(4) at pH 6.5 and then treated with sodium [(3)H]borohydride. The borohydride-treated samples were subjected to hydrolysis in 6 normal HCl at 110 degrees C for 22 hours and then analyzed by high voltage paper electrophoresis and thin layer chromatography. This analysis demonstrated the formation of labeled homoserine as the major reduction product when phosphorylated membrane samples were treated with sodium [(3)H]borohydride. This suggests that the phosphoryl group in the plasma membrane ATPase of red beet storage tissue is attached to the beta-carboxyl side chain of an aspartic acid residue in the active site of the enzyme.

Ca2+-Counteracted Changes of Membrane Functions Induced by Calmodulin Antagonists in Red Beet Storage Tissue, in Pea and Maize Tissues Treated with Optimal Fusicoccin Concentrations

Previous work shows that the two calmodulin antagonists trifluoperazine (TFP) and chlorpromazine (CP), applied in vivo to isolated segments from Pea stems maize coleoptiles and roots, enhance acid secretion and hyperpolarize the transmembrane electric potential difference (PD) (maize roots) in the absence and, more markedly, in the presence of a fusicoccin (FC) concentration inducing half maximal stimulation of electrogenic H+ extrusion. The present data show that, when the two drugs are supplied in the presence of optimal FC concentrations, their stimulating effect on acid secretion disappears and in the roots becomes inhibitory and is associated with a depolarization of the PD. A similar inhibition of acid secretion and depolarization of PD is induced by TFP and CP in red beet root disks. TFP and CP induce an increase of the leakage of ninhydrin-reacting substances in Pea stem and maize root segments and of betacyanine in red beet root disks both in the absence and in the presence of FC. These responses to the calmodulin antagonists are partially counteracted by 0.5 mM CaSO4. No correlation is observed between the changes in the leakage of amino acids induced by the drugs and the simultaneous effects of these drugs on acid secretion under the different conditions. These results suggest that the two calmodulin antagonists, when applied with PC at optimal concentration, inhibit electrogenic H+ extrusion. The question remains open, however, whether this effect depends on the inactivation of the Ca2+-calmodulin complex or on consequent changes of free Ca2+ concentration or on non-specific modifications of the membrane structure.

Unidirectional sodium fluxes in red beet storage tissue (Beta vulgaris L. cv. Platronde Egyptische): effects of the phytohormones indol‐3yl‐acetic acid and abscisic acid

Abstract. The influence of indol‐3yl‐acetic acid (IAA) and abscisic acid (ABA) on the capacities of the cytoplasm and vacuole and their effects on unidirectional sodium fluxes across the plasmalemma and the tonoplast of aged red beet storage tissue was investigated. After loading the tissue in a labelled NaCl solution the efflux of radio‐activity was measured in unlabelled NaCl. By means of compartmental analysis the capacities and fluxes were determined and compared with those obtained after loading and elution in the presence of IAA or ABA.It was established that both IAA and ABA affect sodium transport across the principal cell membranes. Both hormones inhibited the efflux across the plasma‐lemma, possibly by affecting a Na+ for H+ exchanging system. Efflux across the tonoplast was stimulated by IAA and influx across the same membrane was enhanced by ABA. It was suggested that IAA stimulated a proton pump at this level while the influence of ABA remained difficult to explain.