Acid-induced changes in the in vivo wall-yielding properties of hypocotyl sections of Vigna unguiculata

Elongation growth of hypocotyl sections of Vigna unguiculata under xylem perfusion was significantly enhanced when acid was applied by acid-aerosol to an abraded hypocotyl surface in the air. The in vivo wall extensibility (φ) and the effective turgor (Pⁱ– Y), both of which were determined by the pressure-jump method, increased during acid-induced growth as observed in IAA-induced growth. The intracellular pressure (Pⁱ), however, decreased significantly at the beginning of acid-induced growth whereas Pⁱ scarcely changed in IAA-induced growth. This result indicates that protons increase the effective turgor by decreasing the yield threshold as IAA does. There seems to be no essential difference between proton and auxin in the effects on the in vivo mechanical properties of the surface cell wall.     

Effect of cycloheximide on IAA-induced proton excretion and IAA-induced growth in abraded Avena coleoptiles

IAA-induced proton excretion in peeled or abraded oat ( Avena saliva L. cv. Victory) coleoptiles is closely associated with IAA-induced growth. It was attempted to separate these two processes by using cycloheximide to inhibit them differentially. Growth of abraded coleoptile segments was measured by a shadow graphic method, and their IAA-induced acidification of the external solution was monitored with a pH meter. IAA stimulated proton excretion in abraded Avena coleoptile segments after a 13 min lag. IAA-induced proton excretion was inhibited within 5 min by cycloheximide at concentrations of 1.8 × 10⁻⁶, 3.6 × 10 or 3.6 × 10⁻⁵ M. Cycloheximide at these concentrations, added within 4 min of IAA, prevented IAA-induced acidification of the medium for at least 60 min. However, it did not prevent IAA-induced growth during this time. It is concluded that some of the initial IAA-induced growth seen in Avena coleoptiles is independent of detectable IAA-induced proton excretion.     

Effects of UV-C radiation on extension growth, H+-extrusion and transmembrane electric potential in maize coleoptile segments

The effect of 253.7 nm ultraviolet radiation on elongation growth, medium acidification and changes in electric potential difference between vacuole and external medium in cells of maize ( Zea mays L.) coleoptile segments was investigated. It was found that irradiation with 390, 1170, 3900 and 5 850 J m⁻² UV-C (ultraviolet radiation 253.7 nm) inhibited elongation growth, whereas at 195 J m⁻² stimulation of growth was observed. The administration of IAA (10⁻⁵ M ) to the incubation medium of coleoptile segments partially abolished the inhibitory effect of UV-C. The pH of the incubation medium, measured simultaneously with growth, showed that the exposure of the segments to UV-C caused inhibition of H⁺-extrusion (or stimulation of H⁺ uptake). The presence of IAA (10⁻⁵ M ) in the incubation medium promoted (except after 5850 J m⁻² irradiation) H⁺-extrusion to a level comparable with that produced by IAA in non-irradiated segments. In UV-C irradiated segments the potential difference underwent significant alterations. Irradiation of coleoptile segments with 390 J m⁻² caused a transient depolarization, which was fully reversible within 30 min, while at higher doses depolarization was irreversible. The hyperpolarization of the membrane potential (MP) in cells of maize coleoptile induced by IAA was completely nullified by subsequent irradiation with UV-C. It is suggested that UV-C inhibited IAA-induced growth by a mechanism independent of cell wall acidification.     

Roles of auxin and gibberellic acid in growth and maturation of epicotyls of Vigna angularis: Cell wall changes

The effects of auxin and gibberellic acid on cell wall composition in various regions of epicotyls of azuki bean ( Vigna angularis Ohwi and Ohashi cv. Takara) were investigated with the following results. (1) Young segments excised from apical regions of the epicotyl elongated in response to added 10⁻⁴ M indole-3-acetic acid (IAA). When the segments were supplied with 50 m M sucrose, the IAA-induced segment growth was accompanied by enhanced overall synthesis of cell wall polysaccharides, such as xyloglucans, polyuronides and cellulose. This IAA effect on the cell wall synthesis is a consequence of extension growth induced by IAA. Gibberellic acid (GA) at 10⁻⁴ M synergistically enhanced the IAA-induced cell wall synthesis as well as IAA-induced extension growth, although GA by itself neither stimulated the cell wall synthesis nor extension growth. In the absence of sucrose, cell wall synthesis was not induced by IAA or GA. (2) In mature segments excised from basal regions of the epicotyl, no extension growth was induced by IAA or GA. GA enhanced the synthesis of xylans and cellulose when the segments were supplied with 50 m M sucrose. IAA had no effect on the cell wall synthesis. These findings indicate that synthesis of polyuronides, xyloglucans and cellulose, which occurs during extension growth of the apical region of the epicotyl, is regulated chiefly by auxin whereas synthesis of xylans and cellulose during cell maturation in the basal region of the epicotyl is regulated by GA.     

Bioelectrical reactions of Nitellopsis obtusa induced by indole-3-acetic acid

The complex of bioelectrical paramenters (membrane potential, membrane resistance and capacitance) of internodal cells of Nitellopsis obtusa was measured over a wide range of IAA concentration (10⁻¹⁰ to 10⁻⁴ M ) with two intracellular microelectrodes. Primary effects of IAA at a concentration as low as 10⁻¹⁰ M were observed. The optimum range of IAA action was from 10⁻⁹ to 10⁻⁶ M . The type of IAA-induced electroresponse depended on the initial level of membrane potential, which characterized the energetic state of the plasmalemma. In the energized state (ca −200 mV) N. obtusa cells appeared to have 3 typical reactions: hyperpolarization (membrane potential less than K⁺-equilibrium potential), depolarization (membrane potential higher than K⁺-potential) and absence of response at K⁺-electrochemical equilibrium. Membrane capacitance was found constant at 0.74 ± 0.05 μF cm⁻², but membrane resistance increased up to 50% independently of the sign of the electrogenic reaction. Increase of membrance capacitance and decrease of the membrane resistance was a feature of the de-energized state (ca −135 mV) and may be explained by lower viscosity of membrane lipids, which interacted with IAA. The complex of parameter, including cytoplasmic steaming taken as an indicator of energy supply, is discussed as indicating slow IAA penetration combined with a primary action of IAA on the plasmalemma receptor sites.     

The Characteristics of the Adjustment in the pH of the Xylem Exudate of Segments of Vigna Hypocotyls during Xylem Perfusion

The xylem vessels of segments of Vigna hypocotyl were perfusedwith solutions of various pH values. With both acid and alkalineperfusion solutions, the pH of the xylem exudate tended to equilibrateat approximately 6 during the course of xylem perfusion. Thisphenomenon was observed both in air and under anoxia, and alsoin segments pretreated with methanol or preheated in a microwaveoven. Therefore, the adjustment in pH does not depend on therespiratory metabolism but originates in the buffering actionof the xylem wall apoplast. High concentrations of protons inducedthe release of other cations into the xylem exudate under anoxia,and high concentrations of K⁺ or Ca²⁺ ions induced the releaseof protons into the xylem under anoxia. These results indicatethat a cation-exchange reaction on the xylem cell wall was responsiblefor the buffering effect. The physiological role of the highbuffering capacity of the xylem wall apoplast is discussed inlight of the role of the proton pump of the plasmalemma in elongationgrowth. (Received September 25, 1990; Accepted January 21, 1991)     

Relationships between auxin- and acid-induced growth in Vigna radiata hypocotyl segments

The H⁺- and IAA-induced growth responses of isolated Vigna radiata (L.) Wilczek hypocotyl segments were investigated concurrently with IAA-induced H⁺ excretion. The effects of external pH on these reactions were also studied. Experiments were performed with intact, peeled and abraded segments. Only abraded segments reacted to H⁺ and to IAA. In short-term experiments, the cuticle prevented proton efflux and influx; however, it allowed gradual ion movements which become measurable after 1 h. Both phases of the IAA growth response reacted to external pH. The interactions between these two phases and their pH dependencies are discussed.     

Stimulation of proton extrusion by K+ and divalent cations (Ni2+, Co2+ Zn2+) in maize root segments

Entry of the divalent cations Ni²⁺, Co²⁺ and Zn²⁺ into cells of maize ( Zea mays L. cv. Dekalb XL 85) root tissue is accompanied by an acidification of the incubation medium, a decrease in both the pH of the cell sap and the level of malate in the cells, and by an inhibition of dark fixation of CO₂. K⁺, on the contrary, induces only a very low acidification of the incubation medium, does not change either the pH of the cell sap or the malate level in the cells, and induces an increase in CO₂ dark fixation. Different mechanisms are postulated for the stimulation of proton extrusion by divalent cations and K⁺.     

The dose-response curves for IAA induced elongation growth and acidification of the incubation medium of Zea mays coleoptile segments

The initial dose-response curves for auxin-induced elongation growth of Zea mays L. coleoptile segments and simultaneously measured changes of pH of the incubation medium were studied. It was found that these curves are bell-shaped on all occasions and that at all IAA concentrations studied acidification of the incubation medium took place. The optimum response for IAA-induced elongation growth and acidification of the incubation medium was 10⁻⁵ and 10⁻⁴ M IAA, respectively. The regression curves and correlation coefficients between magnitude of the growth response and acidification of the incubation medium indicated a close relationship between these sets of data over a wide range of IAA concentrations.     

Ontogenetic changes in potassium transport in xylem of tomato

The influence of plant ontogeny on xylem exudate K⁺ concentrations and K⁺ transport to the shoot was studied in both nutrient-solution and field-grown tomato plants ( Lycopersicon esculentum ).
K⁺ concentrations in xylem exudate from decapitated plants decreased during tomato plant development from a high of 12 m M to a low of 5 m M . In the nutrient-solution plants, the most rapid decline occurred during the vegetative growth phase, while in field-grown plants, the xylem K⁺ concentrations remained high during an-thesis and then subsequently declined. The rapid decline in nutrient-solution plants might be related to a decrease in the absorptive efficiency of the root system. In field-grown plants, a reduction in the availability of assimilates to the root might account in part for the decrease in xylem exudate K⁺ concentrations. The volume (ml h⁻¹ plant⁻¹) and the net rates of K⁺ exudation (mmol h⁻¹ plant⁻¹) decreased dramatically as the fruits approached maturity. Since only a small reduction in xylem exudate K⁺ concentrations occurred during fruiting, the hydraulic conductivity of the root system decreased as the tomato plants aged. It is proposed that the ontogenetic changes in xylem transport of K⁺ contribute to a reduction in leaf free space K⁺ concentration which would explain the decline in tomato leaf K⁺ concentrations.     

Cation fluxes in the saps of Sinapis alba during the floral transition

Plants of Sinapis alba L. were induced to flower by either a single long day or a single displaced short day. The levels of three cations. Ca²⁺, Mg²⁺ and K⁺, were measured by atomic absorption spectrophotometry in exudates from roots, leaves and apical stem tips. The export of all three cations out of the root system (root exudate) was increased in induced as compared to non-induced plants. No changes were observed in cation export out of the mature leaves (leaf exudate). The supply of cations to the apical bud (apical exudate) did not originate from the phloem and, so, should mainly be of apoplastic origin. Only the supply of Ca²⁺ to the apical bud was increased, not the supply of Mg²⁺ or K⁺. The increase in Ca²⁺ supply was transient and occurred at about the same time as a conspicuous stimulation of cell division, previously detected in the apical bud.     

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.     

Effect of auxins on spermidine uptake into carrot protoplasts

The effect of an auxin, indole-3-acetic-acid (IAA), on spermidine uptake into protoplasts of carrot ( Daucus carota L. cv. Ingrid) was studied. In the presence of 1 m M Ca²⁺, IAA (10⁻⁷ to 10⁻⁴ M ) enchances [¹⁴C]-spermidine uptake into carrot protoplasts, while no stimulation occurs in the absence of Ca²⁺. The time course of the uptake with and without IAA is very rapid and reaches saturation within 1 to 2 min. Preincubation of protoplasts with IAA inhibits the spermidine uptake. La³⁺, known not to penetrate the plasmalemma, exerts the same effect as Ca²⁺, but gives lower uptake values than Ca²⁺. The application of vanadate, an ATPase inhibitor, strongly inhibits IAA-stimulated spermidine uptake, suggesting that an energy-dependent mechanism may be involved in this transport. Neither spermidine nor Ca²⁺ alone stimulate IAA uptake. The synthetic auxin 2,4-dichlorophenoxyacetic acid, yields the same results as IAA with regard to time course of spermidine uptake with and without preincubation while, unlike IAA, no significant effect was observed on the Ca²⁺ -induced increase of spermidine uptake.     

Catabolism of indole-3-acetic acid to indole-3-methanol in a crude enzyme extract and in protoplasts from Scots pine (Pinus sylvestris)

The antagonistic effects of ethylene and Ag⁺ on the metabolism of [1-¹⁴C]indole-3-acetic acid (IAA) and on the rates of ethylene production were studied in tobacco leaf discs ( Nicotiana rustica var. Brasilia ). During the first 10 h of incubation, Ag⁺-pretreated leaf discs contained more free [¹⁴C]IAA than untreated ones due to decreased oxidative decarboxylation, and the discs also produced more ethylene. Exogenously supplied ethylene nullified these effects of Ag⁺. However, the most pronounced effect of Ag⁺ in increasing ethylene production, as well as the strongest antagonistic effect of exogenous ethylene, were found between 24 and 48 h of incubation. During this time span no effect on the level of free IAA and on its decarboxylation could be observed. It is suggested that ethylene exerted its autoinhibitory effect by a feedback control on the IAA-induced ethylene biosynthesis. Possible mechanisms for the autoinhibitory effect of ethylene are discussed.     

Potassium ion channels in the plasmalemma

The potassium ion is an indispensible cytosolic component of living cells and a key osmolyte of plant cells, crossing the plasmalemma to drive physiological processes like cell growth and motor cell activity. K⁺ transport across the plasmalemma may be passive through channels, driven by the electrochemical gradient, K⁺ equilibrium potential (E_K) – membrane potential (V_m), or secondary active by coupling through a carrier to the inward driving force of H⁺ or Na⁺. Known K⁺ channels are permeable to monovalent cations, a permeability order being K⁺ > Rb⁺ > NH₄⁺ > Na⁺≥ Li⁺ > Cs⁺. The macroscopic K⁺ currents across a cell or protoplast surface commonly show rectification, i.e. a V^m-dependent conductance which in turn, may be controlled by the cytosolic activity of Ca²⁺, of K⁺, of H⁺, or by the K⁺ driving force. Analysis by the patch clamp technique reveals that plant K⁺ channels are similar to animal channels in their single channel conductance (4 to 100 pS), but different in that a given channel population slowly activates and may not inactivate at all. Single-channel kinetics reveal a broad range of open times (ms to s) and closed times (up to 100 s). Further progress in elucidating plant K⁺ channels will critically depend on molecular cloning, and the availability of channel-specific (phyto)toxins.     

K/Na selectivity at the plasmalemma of cortical root cells and preferential release of sodium to the xylem vessels in roots of Atriplex hortensis

Using excised roots of Atriplex hortensis L., cv. Gelbe Gartenmelde, the uptake, accumulation and xylem transport of K⁺ and Na⁺ have been measured. Influx as well as xylem transport proved to discriminate little between K⁺ and Na⁺, when considered in relation to the external solution. Both K⁺ and Na⁺ inhibited the uptake and xylem transport of each other to about the same degree. Measurements of intracel-lular Na⁺ fluxes by means of compartment analysis indicated that the low degree of K/Na discrimination during uptake was due to low influx selectivity. Moreover, K⁺/Na⁺ exchange at the plasmalemma was not very efficient in Atriplex roots. In order to establish the basis of the low K/Na discrimination in xylem transport, the rates of K⁺ and Na⁺ transport were related to the cytoplasmic K⁺ and Na⁺ concentrations to yield the selectivity ratio of transport, S(transport) = (φ_cx(K) × [Na⁺]_c)/(φ_cx(Na) × [K⁺]_c). Under all conditions this ratio was far below one indicating that Na⁺ was favoured during xylem release in excised roots of Atriplex at low external concentrations. The implications of this discrimination in favour of Na+ are discussed with respect to salt tolerance of A. hortensis .     

ACCUMULATION OF SULFURIC ACID IN DICTYOTALES (PHAEOPHYCEAE): TAXONOMIC DISTRIBUTION AND ION CHROMATOGRAPHY OF CELL EXTRACTS

Four species in the order Dictyotales ( Dictyopteris latiuscula (Okamura) Okamura, D. prolifera (Okamura) Okamura, D. repens (Okamura) Børgesen, and Spatoglossum crassum J. Tanaka) were found to be highly acidic as in some species of the order Desmarestiales (Phaeophyceae). The pH within their cells, presumably that of the vacuole, was estimated to be 0.5 to 0.9 by pH measurements of their cell extracts in distilled water. However, other species of these genera ( D. divaricata (Okamura) Okamura, D. undulata Holmes, and S. pacificum Yendo) did not show high acidity. Ion chromatography of the cell extracts showed that those species contained high concentrations of SO     within their cells, up to 10 times that in seawater but relatively low Cl⁻. The sum of cations examined (Na⁺, NH     , K⁺, Mg²⁺, Ca²⁺) was significantly lower than that of anions (Cl⁻, Br⁻, NO     , SO     ), and the difference is presumed to represent protons (H⁺), causing the extremely low cell sap pH. Estimated cellular proton concentrations calculated from the pH data roughly agreed with those calculated from differences between the sum of cations and anions and that of anions. Although certain other, nonacidic, dictyotalean species also contained high concentrations of SO     , these species contained high concentrations of Mg²⁺, and the sums of cations and anions were balanced.     

Regulation of plant elongation growth by surface and xylem proton pumps

The roles of plasmalemma electrogenic proton pumps in elongation growth of plant stems are discussed on the basis of growth-electrophysiological studies on hypocotyl segments ofVigna unguiculata. Plant stems usually have two spatially separated electrogenic proton pumps: the surface proton pump which is located on the surface membrane of the symplast and the xylem proton pump, on the cell membrane of the symplast/xylem apoplast boundary. The surface proton pump excretes protons into the surface cell wall layer and causes the loosening of the cell wall. The xylem proton pump excretes protons into the xylem apoplast and drives the uptake of solute and water into the symplastvia secondary and/or tertiary active mechanisms: the proton cotransport system and the apoplast canal system. Both the surface and the xylem proton pumps are active during elongation growth because both the yielding of cell wall loosening and the uptake of water are necessary for continued elongation growth.     

Accumulation and intracellular compartmentation of lithium ions in Saccharomyces cerevisiae

Abstract Accumulation of Li⁺ in Saccharomyces cerevisiae X2180-1B occured via an apparent stoichiometric relationship of 1: 1 (K⁺/Li⁺) when S. cerevisiae was incubated in the presence of 5 and 10 mM LiCl for 3 h. Other cellular cations (Mg²⁺, Ca²⁺ and Na⁺) did not vary on Li⁺ accumulation, although lithium chemistry dictates a degree of similarity to Group I and II metal cations. Compartmentation of Li⁺ was mainly in the vacuole which accounted for 85% of the Li⁺ accumulated after a 6-h incubation period. The remainder was located in the cytosol with negligible amounts being bound to cell fragments including the cell wall. Transmission electron microscopy of Li⁺-loaded cells revealed enlarged vacuoles compared with control cells. This asymmetric cellular distribution may therefore enhance tolerance of S. cerevisiae to Li⁺ and ensure that essential metabolic processes in the cytosol are not disrupted.     

Characterization of the plasmalemma ATPase activity from roots of Plantago major ssp. pleiosperma, purified by the two-phase partitioning method

A purified plasmalemma preparation from roots of Plantago major L. ssp. pleiosperma (Pilger) was obtained by the two-phase partitioning method, using 6.5% (w/w) of Dextran T-500 and polyethylene glycol 3350, respectively. The distribution of murker enzymes proved the purity of the plasmalemma fraction. The ATPase activity was characterized by determining its sensitivity to anions, cations and inhibitors. The Mg²⁺-dependent ATPase activity peaked at pH 7.25, K⁺-stimulation at pH 6.75, and the Cl⁻ -stimulation both at pH 6.75 and 7.5 (all in the presence of 3 m M MgSO₄). The plasmalemma preparations hydrolyzed preferentially ATP (in the presence of Mg²⁺), although they were less specific for ATP at pH 7.5 than at pH 6.75. The Cl⁻ - stimulated ATPase is probably associated with and located on the plasmalemma. The question if the Cl⁻ -stimulated activity is due to an ATPase distinct from the classical K⁺-stimulated ATPase is considered.