Regulation of K+ Channels in Maize Roots by Water Stress and Abscisic Acid

Root cortical and stelar protoplasts were isolated from maize (Zea mays L.) plants that were either well watered or water stressed, and the patch-clamp technique was used to investigate their plasma membrane K⁺ channel activity. In the root cortex water stress did not significantly affect inward- or outward-rectifying K⁺ conductances relative to those observed in well-watered plants. In contrast, water stress significantly reduced the magnitude of the outward-rectifying K⁺ current in the root stele but had little effect on the inward-rectifying K⁺ current. Pretreating well-watered plants with abscisic acid also significantly affected K⁺ currents in a way that was consistent with abscisic acid mediating, at least in part, the response of roots to water stress. It is proposed that the K⁺ channels underlying the K⁺ currents in the root stelar cells represent pathways that allow K⁺ exchange between the root symplasm and xylem apoplast. It is suggested that the regulation of K⁺ channel activity in the root in response to water stress could be part of an important adaptation of the plant to survive drying soils.     

Inhibition of Ion Transport in Excised Barley Roots by Abscisic Acid; Relation to Water Permeability of the Roots

Previous papers have shown that abscisic acid can inhibit transportof ions across the root to the xylem vessels, resulting in reducedexudation from excised roots or inhibiting guttation from intactplants. However, it has not been established whether the inhibitionwas due to a reduction in salt transport (J_s) or in permeabilityof the roots to water (L_p). This paper investigates the effectof ABA on L_p and J_s separately. It is shown that L_p increasedin ABA and then fell, but was about the same as in control rootswhen transport was inhibited. The effect of ABA on exudationtherefore appeared to be mainly due to reduction in J_s. Inhibitionof J_s was also present in intact, transpiring plants and sowas not due to reduced water flow. The inhibition of ion releaseto the xylem affected Na⁺, Mg²⁺, Ca²⁺, and phosphate as wellas the major ion in the exudate, K⁺. It is concluded that ABAinhibits salt transport to the shoot by acting on ion transportinto the xylem, and not by reducing water flow coupled withsalt transport.     

Temperature-Dependent Water and Ion Transport Properties of Barley and Sorghum Roots : II. Effects of Abscisic Acid

Water flux through excised roots (J_v) is determined by root hydraulic conductance (L_p) and the ion flux to the xylem (J_i) that generates an osmotic gradient to drive water movement. These properties of roots are strongly temperature dependent. Abscisic acid (ABA) can influence J_v by altering L_p, J_i, or both. The effects of root temperature on responses to ABA were determined in two species differing in their temperature tolerances. In excised barley (Hordeum vulgare L.) roots, J_v was maximum at 25°C; 10 micromolar ABA enhanced J_v, primarily by increasing L_p, at all temperatures tested (15-40°C). In sorghum (Sorghum bicolor L.) roots, J_v peaked at 35°C; ABA reduced this optimum temperature for J_v to 25°C by increasing L_p at low temperatures and severely inhibiting J_i (dominated by fluxes of K⁺ and NO₃⁻) at warm temperatures. The inhibition of K⁺ flux by ABA at high temperature was mostly independent of external K⁺ availability, implying an effect of ABA on ion release into the xylem. In sorghum, ABA enhanced water flux through roots at nonchilling low temperatures but at the expense of tolerance of warm temperatures. These effects imply that ABA may shift the thermal tolerance range of roots of this heat-tolerant species toward cooler temperatures.     

The Metabolism and Transport of Abscisic Acid During Grain Fill in Wheat

The metabolism and transport of (±)-l2-¹⁴Clabscisic acidand its metabolites was investigated during the period of graindevelopment in wheat. Forty-five hours after feeding the hormoneinto the flag leaf blade, or after injection into the grains,nine metabolites could be extracted with acetone. Four of thesecompounds have been identified. They are phaseic acid, dihydrophaseicacid, abscisyl-ß-D-glucopyranoside and the polar metabolite.As well as the acetone-soluble metabolites a number of othershave been found which are insoluble in acetone. These appearto be conjugated to lipids, ‘gluten-like’ proteins,and carbohydrates. ABA and its metabolites were transported to all parts of theplant above the flag leaf node when the radioactive hormonewas introduced into the blade. However, when it was injectedinto the grain the radioactivity remained there until the pointwhen dry matter accumulation ceased, after which it was foundonly in the peduncle. The results suggest that the increasing level of ABA duringthe period of dry matter accumulation is due to biosynthesiswithin the grain and that the decrease after dry matter accumulationceases is due to both metabolism and redistribution within theplant.     

Transport and Compartmentation of Abscisic Acid in Roots of Hordeum distichon under Osmotic Stress

Using modified compartmental analysis the unidirectional fluxesof abscisic acid (ABA) and their cytoplasmic and vacuolar contentsin ³H-ABA preloaded barley root segments (Hordeum distichoncv. Aura) have been studied. When root segments were stressedosmotically with sorbitol (osmotic potential of the media ⁰= 0.2 MPa) cytoplasmic and vacuolar contents of ABA were enhanced.Under increased stress cytoplasmic and vacuolar contents weremuch lower than in the unstressed controls. ABA fluxes werevery sensitive to osmotic stress and ABA transport from thecytoplasm of the xylem parenchyma to the xylem vessels (_cx)was rapidly inhibited. The cultivar Aura has higher cytoplasmicand vacuolar ABA contents than the barley cultivar Kocherperle.This correlates well with the higher stress tolerance of theAura cultivar. Key words: Abscisic acid, Compartmentation, Osmotic stress     

Proline Accumulation in Maize (Zea mays L.) Primary Roots at Low Water Potentials (I. Requirement for Increased Levels of Abscisic Acid)

We have characterized sulfate transport in the unicellular green alga Chlamydomonas reinhardtii during growth under sulfur-sufficient and sulfur-deficient conditions. Both the Vmax and the substrate concentration at which sulfate transport is half of the maximum velocity of the sulfate transport (K1/2) for uptake were altered in starved cells: the Vmax increased approximately 10-fold, and the K1/2 decreased approximately 7-fold. This suggests that sulfur-deprived C. reinhardtii cells synthesize a new, high-affinity sulfate transport system. This system accumulated rapidly; it was detected in cells within 1 h of sulfur deprivation and reached a maximum by 6 h. A second response to sulfur-limited growth, the production of arylsulfatase, was apparent only after 3 h of growth in sulfur-free medium. The enhancement of sulfate transport upon sulfur starvation was prevented by cycloheximide, but not by chloramphenicol, demonstrating that protein synthesis on 80S ribosomes was required for the development of the new, high-affinity system. The transport of sulfate into the cells occurred in both the light and the dark. Inhibition of ATP formation by the antibiotics carbonylcyanide m-chlorophenylhydrazone and gramicidin-S and inhibition of either F- or P-type ATPases by N,N-dicyclohexylcarbodiimide and vanadate completely abolished sulfate uptake. Furthermore, nigericin, a carboxylate ionophore that exchanges H+ for K+, inhibited transport in both the light and the dark. Finally, uptake in the dark was strongly inhibited by valinomycin. These results suggest that sulfate transport in C. reinhardtii is an energy-dependent process and that it may be driven by a proton gradient generated by a plasma membrane ATPase.     

Action of Abscisic Acid on Auxin Transport and its Relation to Phototropism

The action of abscisic acid on the kinetics of auxin transport through Zea mays L. (cv. Goudster) coleoptiles has been investigated. Abscisic acid applied simultaneously with indoleacetic acid-2-¹⁴C in the donor block reduced the transport intensity without materially affecting the basipetal velocity or the uptake. No effect on acropetal transport was observed. The data have been used to discuss the similarities in effects of abscisic acid and visible radiation and a hypothesis is proposed to explain the phenomena of phototropism.     

Effect of Temperature on the Radial Exchange of Labelled Water in Maize Roots

The temperature dependence of the efflux kinetics of labelledwater in isolated maize roots has been studied. The purposeof these experiments was to obtain the energy of activation,E (kcal/mole), of the rate-limiting step in this radial exchangeprocess under various experimental conditions. Estimates ofE were obtained from linear relations between ln{D'_w} and thereciprocal of the absolute temperature; values of the apparentdiffusion coefficient, D'_w, of labelled water in the root werefound from an analytical treatment of the efflux data in termsof a cylindrical diffusion model. The energy of activation forlabelled-water exchange in normal roots was 14.9 kcal/mole.The corresponding value for ‘dead’ (boiled) rootswas 3.9 kcal/mole. These values of E substantiate the view thatin normal roots the penetration of water across the membranesof the root cells constitutes the rate-determining step in theefflux whereas in ‘dead’ roots extracellular diffusionof water is the source of rate-control. Similar temperature dependence studies were performed on theefflux kinetics from normal and ‘dead’ roots treatedwith 10^–5 M phenylmercuric acetate (PMA). The energiesof activation for labelled-water exchange in normal and ‘dead’roots under these conditions were respectively 15.5 and 5.3kcal/mole. Moreover, the results of the efflux experiments onPMA-treated roots were considered to indicate that this inhibitorproduces an alteration in some structural aspect of the rate-controlling‘membranes’.     

High-Frequency Regeneration by Abscisic Acid (ABA) from Petiole Callus of Jatropha curcas

Jatropha curcas L. is attaining worldwide interest as an important biofuel crop. Experiments were conducted to improve the prevailing micropropagation technique as well as to develop a new ex vitro rooting method for J. curcas plant regeneration. Regeneration and ex vitro rooting efficiency was enhanced by augmenting the culture medium with abscisic acid (ABA). Different concentrations of 6-benzylaminopurine (BAP) and indole-3-butyric acid (IBA) were tested for callus generation from both in vitro and in vivo explants (leaf and petiole) on Murashige and Skoog (MS) medium. The best regenerative callus was achieved on MS medium supplemented with BAP (4.44 μM) and IBA (2.45 μM) from in vitro-cultured petioles. Highest regeneration (91%) was achieved by culturing petiole callus on MS medium supplemented with BAP (8.88 μM), IBA (0.49 μM), and ABA (1.9 μM), whereas 61% regeneration was obtained from in vitro leaf callus. Shoot proliferation and elongation was achieved on BAP (2.22 μM) and IAA (8.56 μM) with 10–13 shoots per explants. Highest rooting (65%) was achieved from M1 shoots (BAP, IAA, and ABA) on MS medium supplemented with IBA (2.45 μM), naphthaleneacetic acid NAA (0.54 μM), and 0.02% activated charcoal. Ex vitro rooting of 1-mo-old M1 shoots obtained from the charcoal-containing medium resulted optimum rooting (>72%) when transferred to polybags containing sterile sand. The plantlets were successfully acclimatized in soil with more than 98% survival rate in the greenhouse.     

Effects of Salinity on Water Transport of Excised Maize (Zea mays L.) Roots

The root pressure probe was used to determine the effects of salinity on the hydraulic properties of primary roots of maize (Zea mays L. cv Halamish). Maize seedlings were grown in nutrient solutions modified by additions of NaCl and/or extra CaCl₂ so that the seedlings received one of four treatments: Control, plus 100 millimolar NaCl, plus 10 millimolar CaCl₂, plus 100 millimolar NaCl plus 10 millimolar CaCl₂. The hydraulic conductivities (Lp_r) of primary root segments were determined by applying gradients of hydrostatic and osmotic pressure across the root cylinder. Exosmotic hydrostatic Lp_r for the different treatments were 2.8, 1.7, 2.8, and 3.4·10⁻⁷ meters per second per megapascals and the endosmotic hydrostatic Lp_r were 2.4, 1.5, 2.7, and 2.3·10⁻⁷ meters per second per megapascals, respectively. Exosmotic Lp_r of the osmotic experiments were 0.55, 0.38, 0.68, and 0.60·10⁻⁷ meters per second per megapascals and the endosmotic Lp_r were 0.53, 0.21, 0.56, and 0.54·10⁻⁷ meters per second per megapascals, respectively. The osmotic Lp_r was significantly smaller (4-5 times) than hydrostatic Lp_r. However, both hydrostatic and osmotic Lp_r experiments showed that salinization of the growth media at regular (0.5 millimolar) calcium levels decreased the Lp_r significantly (30-60%). Addition of extra calcium (10 millimolar) to the salinized media caused ameliorative effects on Lp_r. The low Lp_r values may partially explain the reduction in root growth rates caused by salinity. High calcium levels in the salinized media increased the relative availability of water needed for growth. The mean reflection coefficients of the roots using NaCl were between 0.64 and 0.73 and were not significantly different for the different treatments. The mean values of the root permeability coefficients to NaCl of the different treatments were between 2.2 and 3.5·10⁻⁹ meters per second and were significantly different only in one of four treatments. Cutting the roots successively from the tip and measuring the changes in the hydraulic resistance of the root as well as staining of root cross-sections obtained at various distances from the root tip revealed that salinized roots had mature xylem elements closer to the tip (5-10 millimeters) compared with the controls (30 millimeters). Our results demonstrate that salinity has adverse effects on water transport and that extra calcium can, in part, compensate for these effects.     

Abscisic Acid Translocation and Influence of Water Stress on Grain Abscisic Acid Content

The movement of foliar applied [1-¹⁴C]abscisic acid (ABA) inwheat plants (Triticum aestivum L., cv. Kolibri) was investigatedat two stages of grain development (1000 grains, weight 19 and24 g dry matter). [1–¹⁴C]ABA seemed to be readily translocated within 12h into the developing grains as well as in other plant parts.A subsequent rapid metabolism took place leading to a decreasedactivity of the ABA-containing chromatogram fraction in theyounger plants 48 h after application. The metabolism seemodto be less intensive in the older grains, where the activityrunning with the ABA increased over 64 h. Treating the leaves of barley plants (Hordeum vulgare, L., cv.Union) 2 weeks after anthesis with a gentle stream of warm air(36° C) resulted in a significant increase in the ABA contentof all parts of the ear. The results mentioned above indicatethat this may be partially due to translocation from other partsof the plant such as the leaves.     

Some Structural Changes During Ripening of Mangoes (Mangifera indica var. Alphonso) by Abscisic Acid Treatment

Abscisic acid at 10^–6 M concentration enhances ripeningof mangoes. The cells in the pulp of the fruit are large andparenchymatous and lose their integrity due to cell wall hydrolysisat the ripe stage. They contain abundant polysaccharides, consistingmainly of starch, which is degraded during ripening. The chloroplastsare transformed to chromoplasts containing red or yellow carotenoidpigment. Abscisic acid treatment enhances all of these processes.Mitochondria, on the other hand, retain their structural integritythroughout the ripening process in untreated and abscisic acid-treatedmangoes. Mangoes, ripening, abscisic acid, structure     

Regulation of the High-Affinity Nitrate Transport System in Wheat Roots by Exogenous Abscisic Acid and Glutamine

Nitrate is a major nitrogen （N） source for most crops. Nitrate uptake by root cells is a key step of nitrogen metabolism and has been widely studied at the physiological and molecular levels. Understanding how nitrate uptake is regulated will help us engineer crops with improved nitrate uptake efficiency. The present study investigated the regulation of the high-affinity nitrate transport system （HATS） by exogenous abscisic acid （ABA） and glutamine （Gin） in wheat （Triticum aestivum L.） roots. Wheat seedlings grown in nutrient solution containing 2 mmol/L nitrate as the only nitrogen source for 2weeks were deprived of N for 4d and were then transferred to nutrient solution containing 50 μmol/L ABA, and 1 mmol/L Gin in the presence or absence of 2 mmol/L nitrate for 0, 0.5, 1, 2, 4, and 8 h. Treated wheat plants were then divided into two groups. One group of plants was used to investigate the mRNA levels of the HATS components NRT2 and NAR2 genes in roots through semi-quantitative RT-PCR approach, and the other set of plants were used to measure high-affinity nitrate influx rates in a nutrient solution containing 0.2 mmol/L ^15N-labeled nitrate. The results showed that exogenous ABA induced the expression of the TaNRT2.1, TaNRT2.2, TaNRT2.3, TaNAR2.1, and TaNAR2.2 genes in roots when nitrate was not present in the nutrient solution, but did not further enhance the induction of these genes by nitrate. Glutamine, which has been shown to inhibit the expression of NRT2 genes when nitrate is present in the growth media, did not inhibit this induction. When Gin was supplied to a nitrate-free nutrient solution, the expression of these five genes in roots was induced. These results imply that the inhibition by Gin of NRT2 expression occurs only when nitrate is present in the growth media. Although exogenous ABA and Gin induced HATS genes in the roots of wheat, they did not induce nitrate influx.     

Effects of Abscisic Acid and of Hydrostatic Pressure Gradient on Water Movement through Excised Sunflower Roots

The effect of abscisic acid on the exudation rate from decapitated roots of sunflower plants (Helianthus annuus L.) was investigated in the presence and absence of an imposed hydrostatic pressure gradient. The magnitude of the abscisic acid effect was constant even when suctions up to 60 cm Hg were applied to the cut stumps.     

Water Transport in Onion (Allium cepa L.) Roots (Changes of Axial and Radial Hydraulic Conductivities during Root Development)

The hydraulic architecture of developing onion (Allium cepa L. cv Calypso) roots grown hydroponically was determined by measuring axial and radial hydraulic conductivities (equal to inverse of specific hydraulic resistances). In the roots, Casparian bands and suberin lamellae develop in the endodermis and exodermis (equal to hypodermis). Using the root pressure probe, changes of hydraulic conductivities along the developing roots were analyzed with high resolution. Axial hydraulic conductivity (Lx) was also calculated from stained cross-sections according to Poiseuille's law. Near the base and the tip of the roots, measured and calculated Lx values were similar. However, at distances between 200 and 300 mm from the apex, measured values of Lx were smaller by more than 1 order of magnitude than those calculated, probably because of remaining cross walls between xylem vessel members. During development of root xylem, Lx increased by 3 orders of magnitude. In the apical 30 mm (tip region), axial resistance limited water transport, whereas in basal parts radial resistances (low radial hydraulic conductivity, Lpr) controlled the uptake. Because of the high axial hydraulic resistance in the tip region, this zone appeared to be "hydraulically isolated" from the rest of the root. Changes of the Lpr of the roots were determined by measuring the hydraulic conductance of roots of different length and referring these data to unit surface area. At distances between 30 and 150 mm from the root tip, Lpr was fairly constant (1.4 x 10-7 m s-1 MPa-1). In more basal root zones, Lpr was considerably smaller and varied between roots. The low contribution of basal zones to the overall water uptake indicated an influence of the exodermal Casparian bands and/or suberin lamellae in the endodermis or exodermis, which develop at distances larger than 50 to 60 mm from the root tip.     

Three Classes of Abscisic Acid (ABA)-Insensitive Mutations of Arabidopsis Define Genes that Control Overlapping Subsets of ABA Responses

Wild type and three abscisic acid (ABA)-insensitive mutants of Arabidopsis (ABI1, ABI2, and ABI3) were compared for their ability to respond to ABA for a variety of ABA-inducible responses throughout the life cycle of the plants. The responses tested included effects on seedling growth, proline accumulation in seedlings, ABA-regulated protein synthesis in plantlets, and seed storage protein and lipid synthesis and accumulation. The abi1 and abi2 mutants showed reduced sensitivity to ABA for inhibition of seedling growth, induction of proline accumulation, and alterations in protein synthesis patterns during vegetative growth, but had wild type levels of storage reserves. In contrast, the abi3 mutant had wild type sensitivity for induction of proline accumulation and was only slightly less responsive to ABA with respect to effects on seedling growth and changes in patterns of protein synthesis. The major effects of this mutation were on seed development. Seeds of the abi3 mutant had two-thirds of the wild type level of storage protein and one-third the wild type level of eicosenoic acid, the major fatty acid component of storage lipids in wild type seeds. These results show that none of the abi mutants is insensitive for all ABA-inducible responses and that the abi3 effects are not seed-specific. Comparison of the degree of ABA sensitivity of monogenic mutant lines with that of digenic mutant lines carrying pairwise combinations of the abi mutations suggests that ABA responses in mature seeds are controlled by at least two parallel pathways.     

Timing of Kernel Development in Water-stressed Maize: Water Potentials and Abscisic Acid Concentrations

Maize (Zea mays L. cv. Pioneer 3925) subjected to post-anthesiswater stress during the first 2 weeks of kernel developmenthad lower leaf-water potentials and higher leaf-ABA concentrationsthan well-watered controls. There was a concomitant rise inABA concentration in kernel tissues 3 and 7 d after pollination(DAP), after which the concentration decreased to control levelsby 13 DAP. Kernel water potential, however, remained unchangedby the water stress. Radiolabelled ABA, fed to a leaf, was translocatedto kernels, where free ABA as well as several ABA metaboliteswere the major labelled fractions. This suggested that the stress-inducedkernel ABA was of maternal origin. Since ABA plays a putativerole in seed maturation of several crop species, and appliedABA or water stress often hastens seed development, we expectedthat a water-stress-induced rise in kernel ABA concentrationearly in grain development may serve to prematurely induce storage-productaccumulation. Zein, starch and several enzymes key to the starchsynthesis pathway followed the same course of induction throughoutthe experiment, with no difference between treatments Henceit was concluded that although water stress increased kernelABA independent of kernel water status, there was no apparenteffect of water stress or ABA on timing of early kernel developmentalprocesses. Zea mays L. cv. Pioncer 3925, maize, water stress, abscisic acid, endosperm development