The spatially variable inhibition by water deficit of maize root growth correlates with altered profiles of proton flux and cell wall pH

Growth of elongating primary roots of maize (Zea mays) seedlings was approximately 50% inhibited after 48 h in aerated nutrient solution under water deficit induced by polyethylene glycol 6000 at -0.5 MPa water potential. Proton flux along the root elongation zone was assayed by high resolution analyses of images of acid diffusion around roots contacted for 5 min with pH indicator gel. Profiles of root segmental elongation correlated qualitatively and quantitatively (r(2) = 0.74) with proton flux along the surface of the elongation zone from water-deficit and control treatments. Proton flux and segmental elongation in roots under water deficit were remarkably well maintained in the region 0 to 3 mm behind the root tip and were inhibited from 3 to 10 mm behind the tip. Associated changes in apoplastic pH inside epidermal cell walls were measured in three defined regions along the root elongation zone by confocal laser scanning microscopy using a ratiometric method. Finally, external acidification of roots was shown to specifically induce a partial reversal of growth inhibition by water deficit in the central region of the elongation zone. These new findings, plus evidence in the literature concerning increases induced by acid pH in wall-extensibility parameters, lead us to propose that the apparently adaptive maintenance of growth 0 to 3 mm behind the tip in maize primary roots under water deficit and the associated inhibition of growth further behind the tip are related to spatially variable changes in proton pumping into expanding cell walls.     

Physiological and morphological response to water deficit in seedlings of five provenances of Pinus canariensis: potential to detect variation in drought-tolerance

To assess the potential of short-term screenings for drought resistance at the seedling stage to detect ecotypic variation and predict field performance, we studied the responses to water deficit of seedlings of Pinus canariensis from five geographic origins under controlled conditions and compared these responses with the performance of provenances in a multi-site field trial. Leaf water potential, the osmotic component, leaf chlorophyll fluorescence and growth and biomass partitioning were measured as seedlings were subjected for 11 days to two levels of osmotic potential generated by polyethylene glycol (PEG 6000), −1 MPa (slowly imposed water deficit; S) and −1.5 MPa (fast imposed water deficit; F), and a control treatment (no PEG added to the nutrient solution; C). Leaf water potential declined to final mean values of −1.2, −2.7 and −4.7 MPa in the C, S and F treatments, respectively. The ratio of variable to maximum chlorophyll fluorescence declined to final mean values of 0.77, 0.66 and 0.40 in the C, S and F treatments, respectively, with no differences amongst provenances. All provenances showed an active osmotic adjustment (OA) in response to water deficit which varied depending on the drying rate. A slow imposition of water deficit favoured solute accumulation. Pooling all treatments, the index of OA ranged from 0.28 to 0.40, but rose considerably when only C and S treatments were considered (0.56 to 0.70). There was a positive and significant correlation between the overall index of OA (all treatments pooled) and the drought period in the site of origin, suggesting ecotypic variation in OA as a result of drought duration. Seedlings allocated more dry matter to roots than shoots when subjected to moderate and slowly imposed water deficit; only one provenance showed no increase in the root to shoot ratio at the end of the treatment period compared with control seedlings. Responses to controlled water deficits were only qualitatively related to performance (survival and growth) of provenances in several field sites, indicating the involvement of complex mechanisms to cope with drought under natural conditions. However, the provenance with the highest overall index of OA outgrew and outsurvived the other provenances in the most arid site, and the only provenance not modifying the root to shoot ratio in response to water deficit survived the least in all field sites. Acclimation of root to shoot ratio and net solute accumulation to water deficit could hence favour drought-tolerance beyond the seedling stage and be used as preliminary predictors of field performance.     

Implication of ABA and proline on cell membrane injury of water deficit stressed barley seedlings

The aim of this work was to examine the ability of ABA and proline to counteract the deleterious effect of water deficit stress on cell membrane injuries. Six-day-old seedlings of two barley genotypes (cv. Aramir, line R567) were treated with ABA (2·10⁻⁴ M) or proline (0.1 M) for 24 h, and then subjected to osmotic stress for 24h, by immersing their roots in polyethylene glycol (PEG 6000) solution of osmotic potential of −1.0 MPa and −1.5 MPa or by submerging the leaf pieces in PEG solution of osmotic potential of −1.6 MPa. Pretreatment of plants with ABA and proline caused an increase of free proline level in the leaves. Plants treated with ABA exhibited a lower membrane injury index under water stress conditions than those untreated even when no effect of this hormone on RWC in the leaves of stressed plants was observed. Pretreatment of plants with proline prevented to some extent membrane damage in leaves of the stressed seedlings, but only in the case when stress was imposed to roots. Improvement in water status of leaves was also observed in seedlings pretreatment with proline. The protective effect of both ABA and proline was more pronounced in line R567 that exhibited higher membrane injury under water deficit stress conditions.     

Water Deficit Rapidly Stimulates the Activity of a Protein Kinase in the Elongation Zone of the Maize Primary Root

The mechanisms by which plants detect water deficit and transduce that signal into adaptive responses is unknown. In maize (Zea mays L.) seedlings, primary roots adapt to low water potentials such that substantial rates of elongation continue when shoot growth is completely inhibited. In this study, in-gel protein kinase assays were used to determine whether protein kinases in the elongation zone of the primary root undergo activation or inactivation in response to water deficit. Multiple differences were detected in the phosphoprotein content of root tips of water-stressed compared with well-watered seedlings. Protein kinase assays identified water-deficit-activated protein kinases, including a 45-kD, Ca2+-independent serine/threonine protein kinase. Water-deficit activation of this kinase occurred within 30 min after transplanting seedlings to conditions of low water potential and was localized to the elongation zone, was independent of ABA accumulation, and was unaffected by cycloheximide-mediated inhibition of protein translation. These results provide evidence that the 45-kD protein kinase acts at an early step in the response of maize primary roots to water deficit and is possibly involved in regulating the adaptation of root growth to low water potential.     

Changes in leaf water relations, gas exchange, growth and flowering quality in potted geranium plants irrigated with different water regimes

Geranium plants are an important part of urban green areas but suffer from drought, especially when grown in containers with a limited volume of medium. In this experiment, we examined the response of potted geraniums to different irrigation levels. Geranium (Pelargoniumxhortorum L.) seedlings were grown in a growth chamber and exposed to three irrigation treatments, whereby the plants were irrigated to container capacity (control), 60% of the control (moderate deficit irrigation, MDI), or 40% of the control (severe deficit irrigation, SDI). Deficit irrigation was maintained for 2 months, and then all the plants were exposed to a recovery period of 112 month. Exposure to drought induced a decrease in shoot dry weight and leaf area and an increase in the root/shoot ratio. Height and plant width were significantly inhibited by the SDI, while flower color parameters were not affected by deficit treatment. The number of wilting and yellow leaves increased, coinciding with the increase in the number of inflorescences and open flowers. Deficit irrigation led to a leaf water potential of about -0.8MPa at midday, which could have caused an important decrease in stomatal conductance, affecting the photosynthetic rate (Pn). Chlorophyll fluorescence (Fvm) values of 0.80 in all treatments throughout the experiment demonstrate the lack of drought-induced damage to PSII photochemistry. Pressure-volume analysis revealed low osmotic adjustment values of 0.2MPa in the SDI treatment, accompanied by increases in the bulk tissue elastic modulus (epsilon, wall rigidity) and resulting in turgor loss at lower leaf water potential values (-1.38MPa compared with -1.0MPa for the control). Leaf water potential values throughout the experiment below those for Psitlp were not found at any sampling time. By the end of the recovery period, the leaf water potential, stomatal conductance and net photosynthesis had recovered. We infer from these results that moderate deficit irrigation in geranium reduced the consumption of water, while maintaining the good overall quality of plants. However, when SDI was applied, a reduction in the number of flowers per plant was observed.     

Abscisic acid accumulation maintains maize primary root elongation at low water potentials by restricting ethylene production

Previous work showed that primary root elongation in maize (Zea mays L.) seedlings at low water potentials (psi(w)) requires the accumulation of abscisic acid (ABA) (R.E. Sharp, Y. Wu, G.S. Voetberg, I.N. Saab, M.E. LeNoble [1994] J Exp Bot 45: 1743-1751). The objective of the present study was to determine whether the inhibition of elongation in ABA-deficient roots is attributable to ethylene. At a psi(w) of -1.6 MPa, inhibition of root elongation in dark-grown seedlings treated with fluridone to impose ABA deficiency was largely prevented with two inhibitors of ethylene synthesis (aminooxyacetic acid and aminoethoxyvinylglycine) and one inhibitor of ethylene action (silver thiosulfate). The fluridone treatment caused an increase in the rate of ethylene evolution from intact seedlings. This effect was completely prevented with aminooxyacetic acid and also when ABA was supplied at a concentration that restored the ABA content of the root elongation zone and the root elongation rate. Consistent results were obtained when ABA deficiency was imposed using the vp5 mutant. Both fluridone-treated and vp5 roots exhibited additional morphological symptoms of excess ethylene. The results demonstrate that an important role of ABA accumulation in the maintenance of root elongation at low psi(w) is to restrict ethylene production.     

Increased endogenous abscisic Acid maintains primary root growth and inhibits shoot growth of maize seedlings at low water potentials

Roots of maize (Zea mays L.) seedlings continue to grow at low water potentials that cause complete inhibition of shoot growth. In this study, we have investigated the role of abscisic acid (ABA) in this differential growth sensitivity by manipulating endogenous ABA levels as an alternative to external applications of the hormone. An inhibitor of carotenoid biosynthesis (fluridone) and a mutant deficient in carotenoid biosynthesis (vp 5) were used to reduce the endogenous ABA content in the growing zones of the primary root and shoot at low water potentials. Experiments were performed on 30 to 60 hour old seedlings that were transplanted into vermiculite which had been preadjusted to water potentials of approximately −1.6 megapascals (roots) or −0.3 megapascals (shoots). Growth occurred in the dark at near-saturation humidity. Results of experiments using the inhibitor and mutant approaches were very similar. Reduced ABA content by either method was associated with inhibition of root elongation and promotion of shoot elongation at low water potentials, compared to untreated and wild-type seedlings at the same water potential. Elongation rates and ABA contents at high water potential were little affected. The inhibition of shoot elongation at low water potential was completely prevented in fluridone-treated seedlings during the first five hours after transplanting. The results indicate that ABA accumulation plays direct roles in both the maintenance of primary root elongation and the inhibition of shoot elongation at low water potentials.     

Scale of Physiological Processes Sensitivity to PEG-Induced Water Stress in Scots Pine Seedlings

The idea that water deficit strengthening induces concerted changes of plant physiological parameters is rather widespread. However, such changes are often difficult to identify due to challenges in establishments and maintenance of required water stress intensities using solid substrates. Therefore, we exposed Scots pine (Pinus sylvestris L.) seedlings to the range of water potentials from–0.15 to–1.5 MPa in PEG-water culture to identify the series of physiological parameters differently sensitive to water stress. We observed that even mild water stress (–0.15 MPa) inhibited root elongation, which could be one of the main pine seedlings vulnerabilities under drought. Active accumulation of osmolytes was already induced by mild water deficit and further increased with water stress severity. Plant fresh biomass growth sensitivity was more related to changes of relative water content (RWC) than to changes in tissue water content or dry weight accumulation. Plants were able to grow and accumulate dry weight down to–0.5 MPa, but lower medium water potentials (–1.0 and–1.5 MPa) suppressed growth and heavily damaged root cells, as judged from many-fold increase of Ca²⁺ content in roots. Chlorophyll a content was surprisingly sensitive to water stress, while carotenoids level was increased under severe stress conditions. In conclusion, the experimental system with stepwise water potential values allowed us to analyze the sensitivity scale of a range of P. sylvestris physiological processes to water stress. It was largely similar to those described earlier for other plant species, but its peculiarities were high sensitivity of root elongation, marked resistance of biomass growth to water deficit and well-developed ability to osmotic adjustment.     

Water stress induced by PEG decreases the maximum growth pressure of the roots of pea seedlings

Roots of plants growing in dry soil often experience large mechanical impedance because the decreased soil water content is associated with increased in soil strength. The combined effect of mechanical impedance and water stress hinders the establishment of seedlings in many soils, but little is known about the interaction between these two stresses. A method has been designed that, for the first time, measured the maximum axial force exerted by a root growing under controlled water stress. Using this technique the axial force exerted by a pea radicle was measured using a shear beam, while the seedling was suspended in an aerate solution of polyethylene glycol 20 000 at osmotic potentials between 0 and -0.45 MPa. The maximum growth force was then divided by the cross-sectional area of the root to give the maximum axial growth pressure. The value of maximum axial growth pressure decreased linearly from 0.66 and 0.35 MPa as the osmotic potentials of the solution of PEG decreased from 0 to -0.45 MPa. In dry soil, therefore, the maximum strength of soil that a root can penetrate is decreased because of the decrease in maximum growth pressure. The elongation rates of unimpeded roots were similar whether the roots were subject to either a matric potential in soil or to an osmotic potential in a solution of PEG.Key words: Pisum sativum L, pea, mechanical impedance, axial growth pressure, water stress, PEG 20 000.      

Does proline accumulated in leaves of water deficit stressed barley plants confine cell membrane injuries? II. Proline accumulation during hardening and its involvement in reducing membrane injuries in leaves subjected to severe osmotic stress

The purpose of this research was to examine whether proline accumulation in leaves of barley under conditions of mild water deficit (PEG — 0.75 MPa imposed on roots) may modify membrane injuries caused by subsequent severe osmotic stress (PEG — 1.6 MPa imposed on leaves). Six-day-old seedlings of four barley genotypes were used in the experiments. Substantial and different proline accumulation was found in the leaves of mild water deficit-stressed plants of the most investigated genotypes. This stress factor caused rather a small decrease in RWC and did not lead to membrane injuries. Severe osmotic stress imposed on leaves caused considerable membrane injuries in all the genotypes investigated. Leaves of plants pre-stressed with mild water deficit and then subjected to severe osmotic stress exhibited about a 50% lower membrane injury than those of not pre-stressed plants. A possible role of proline accumulated in the leaves of pre-stressed plants in the process of alleviating cell membrane injuries in the leaves subsequently exposed to severe water deficit is discussed.     

Influence of temperature and water potential on root growth of white oak

Root growth of white oak ( Quercus alba L.) was observed under field conditions using a rhizotron. The effects of temperature, soil water potential, and leaf water potential were evaluated on three measures of root growth and development: root elongation rate, number of growing roots, and root growth intensity (sum of projected root area compared to the total root viewing area). Root elongation rate was linearly related to changes in soil temperature and soil water potential. At soil temperatures less than 17deg;C, temperature was the dominant factor affecting rate of growth, bat at temperatures greater than 17°C soil water potential became the important factor. Unlike root elongation rate, the number of growing roots and root growth intensity increased at cold soil temperatures (8°C) and at soil water potentials of-0.3 to -0.8 MPa. At high soil water potentials (-0.1 MPa) root elongation rate reached a maximum while the number of growing roots and root growth intensity were low. These differences showed that root growth and development were not exclusively affected by the soil environment. In addition, the relationship between root growth and predawn leaf water potential suggested that root growth was a contributing factor to the drought resistance of white oak.     

Membrane permeability and micro- and macroelement accumulation in spring wheat cultivars during the short-term effect of salinity- and PEG-induced water stress

The comparative responses of ten spring wheat cultivars to water stress were investigated. Wheat plants were cultured under hydroponics conditions (Hoagland nutrient) to the stage of three-leaf seedlings. Then, the water medium was supplemented with PEG (drought) or NaCl (salinity) to obtain a water status equal to −1.5 MPa. After a 2-day treatment, the changes in the following parameters were determined: fresh and dry weight, macro- and microelement accumulation, membrane injury (electrolyte leakage, lipid peroxidation) and fatty acid content of the phospholipid fraction of plasmalemma (in comparison to plants not stressed, taken as a control). Generally, the plants were more significantly influenced by water stress stimulated by PEG than by NaCl treatment, as compared to the plants cultivated in the control media. The results of the decrease in water content in leaves and electrolyte leakage from cells corresponded well with the intensity of lipid peroxidation (determined by malondialdehyde—MDA-content) and were chosen for the selection of investigated genotypes for tolerance to both stresses. The more tolerant genotypes exhibited the opposite changes in phospholipid fatty acid unsaturation for two applied stresses i.e. NaCl treatment caused a decrease in unsaturation whereas in PEG-treated plants an increase in unsaturation was observed. These changes were reversed for less tolerant plants, i.e. NaCl treatment influenced an increase in fatty acid unsaturation whereas in PEG-treated plants a decrease in unsaturation was measured. The ratio of U/S (unsaturated to saturated fatty acids) correlated with the total amount of accumulated macroelements. The content of Mg, Ca and S in leaves of plants undergoing both stress factors (NaCl and PEG) dropped whereas the K and P content increased in leaves of wheat seedlings cultured on media containing NaCl only. For microelements, a decrease in the accumulation of these nutrients was detected in all investigated seedlings. However, a greater reduction in the level of these elements occurred in seedlings grown on media with PEG in comparison to those grown on NaCl containing media.     

Stomatal limitation of photosynthesis as affected by water stress and CO2 concentration

A water stress effect on photosynthesis and transpiration of wheat seedlings at 50-500 µmol(CO2) mol-1 was measured in an open gas exchange system. The limitation of photosynthesis by stomatal conductance was quantified by a stomatal control coefficient of the net photosynthetic rate. The stomatal control coefficient increased linearly as the water potential of root media decreased to -1 MPa, and it decreased with increasing CO2 concentration.     

Nodule gas exchange and water potential response to rapid imposition of water deficit

The permeability (P) of the gaseous diffusion barrier in the nodules of soybean [Glycine max (L.) Merr.] decreases when water deficits are extended over a 7 to 10 d period. The mechanism controlling P changes is unclear, but may result from the release of water to intercellular pathways, and an associated change in the nodule water potential. The purpose of these experiments was to impose water deficit treatments rapidly in order to determine the early sequence of the responses of nodule water potential and nodule gas exchange without the complications that arise from long-term water deficit treatments. A vertical, split-root system was used to separate nodule drying effects from plant water deficits by replacing humidified air that was passed over upper root nodules in well-watered plants with dry air, or by replacing the nutrient solution that surrounded lower roots with -1.0 MPa polyethylene glycol (PEG) solution, or by a combination of the dry air and PEG treatments. The PEG treatment caused large decreases in both the components of nodule water potential and nodule relative water content, but there was no indication that these factors had immediate, direct effects on either nitrogenase activity or P. After 7 h of the PEG treatment a significant decrease in nitrogenase activity was found but no decrease in P was detected. These results indicate that changes in nitrogenase activity in response to water deficits precede decreases in P. Exposure of nodules to dry air in well-watered plants had no significant effect on either nitrogenase activity or P during the 7 h treatment.     

Carbendazim alleviates effects of water stress on chickpea seedlings

Carbendazim (methyl-2-benzimidazole carbamate) promoted root growth of chickpea (Cicer arietinum L.) seedlings subjected to polyethylene glycol (PEG, osmotic potential −0.5 MPa) induced water stress. The relative water content, membrane stability index, 2,3,5-triphenyltetrazolium chloride reduction and contents of some osmolytes (proline, sucrose, glucose and fructose) enhanced significantly while the contents of lipid peroxides and hydrogen peroxide diminished effectively by addition of 0.05 % carbendazim into PEG solution. This revised version was published online in September 2005 with the corrected author information.     

Effect of osmo- and hydropriming of chickpea seeds on seedling growth and carbohydrate metabolism under water deficit stress

Seven-day-old seedlings obtained from seeds primed with mannitol (4%)and water showed three to four fold more growth with respect to root and shootlength in comparison with seedlings obtained from non-primed seeds. Seedlingswere grown under water deficit stress conditions created by 15% polyethyleneglycol (PEG) 6000 in the medium. Priming of chickpea seeds with NaCl and PEGwasnot effective in increasing seedling growth under these water deficit stressconditions. The activities of amylase, invertases (acid and alkaline), sucrosesynthase (SS) and sucrose phosphate synthase (SPS) were higher in shoots ofprimed seedlings. An increase in the activities of SS, and both the acid andalkaline invertases was also observed in roots of primed seedlings. The twofoldincrease in specific activity of sucrose phosphate synthase was observed incotyledons of primed seedlings. The higher amylase activity in shoots of primedseedlings enhanced the rapid hydrolysis of transitory starch of the shootleading to more availability of glucose for shoot growth and this was confirmedby the low level of starch in shoots of primed seedlings.     

Cytokinin action is coupled to ethylene in its effects on the inhibition of root and hypocotyl elongation in Arabidopsis thaliana seedlings.

Cytokinins have profound effects on seedling development in Arabidopsis thaliana. Benzyladenine (BA) inhibits root elongation in light- or dark-grown seedlings, and in dark-grown seedlings BA inhibits hypocotyl elongation and exaggerates the curvature of apical hooks. The latter are characteristic ethylene responses and, therefore, the possible involvement of ethylene in BA responses was examined in seedlings. It was found that the inhibitory effects of BA on root and hypocotyl elongation were partially blocked by the action of ethylene inhibitors or ethylene-resistant mutations (ein1-1 and ein2-1). Ethylene production was stimulated by submicromolar concentrations of BA and could account, in part, for the inhibition of root and hypocotyl elongation. It was demonstrated further that BA did not affect the sensitivity of seedlings to ethylene. Thus, the effect of cytokinin on root and hypocotyl elongation in Arabidopsis appears to be mediated largely by the production of ethylene. The coupling between cytokinin and ethylene responses is further supported by the discovery that the cytokinin-resistant mutant ckr1 is resistant to ethylene and is allelic to the ethylene-resistant mutant ein2.     

The pressure-jump technique shows maize leaf growth to be enhanced by increases in turgor only when water status is not too high

This study on expansive growth of the first leaf of maize has two goals: one is to determine how the sensitivity of growth to changes in water status varies with the initial water status of the leaf, and the other is to adapt the pressure-jump technique of Okamoto et al. (1989 , Plant and Cell Physiology 30, 979–985), developed for studying growth of excised stem segments, for use on whole seedlings. Initial water status was varied by using: transpiring vs. non-transpiring conditions, seedlings differing in emerged leaf length and hence transpiring area, and root medium without mannitol vs. medium with added mannitol (to –0·3 MPa). The results show that growth changed with changes in plant water status when the water status was low, but was unaffected when water status was very high. A stepwise change in hydrostatic pressure on the root medium was quickly and fully transmitted to the base of the leaf. The increase in leaf elongation due to a pressure step of 0·025 MPa was negligible under conditions of high plant water status and became substantial under conditions of low water status. In adapting the pressure-jump method to the whole seedling, there was some loss of resolution, and the yield threshold Y of the Lockhart equation could not be estimated directly. Nonetheless, the data were suitable for the calculation of volumetric extensibility m and the estimation of growth effective turgor (turgor above Y ). Extensibility was shown to increase 3- to 4-fold when leaf water status was reduced from the maximum to the point where elongation rate was halved, while growth effective turgor was calculated to diminish even more markedly.     

Ethylene production by loblolly pine seedlings associated with water stress

Effects of water stress on production of ethylene and its precursor, 1-aminocyclopropane-1-carboxylic acid (ACC), by loblolly pine ( Pinus taeda L.) seedlings from a Texas drought-hardy and a Virginia Coastal Plain source were investigated. Ethylene production rates in needles from the Virgnia source increased slightly with initial stress (-1.3 MPa), declined until water potential reached -1.6 MPa and then increased sharply at -2.5 MPa. The ethylene production rates in needles from the Texas also increased slightly with initial stress, then decreased with decreasing water potential. Ethylene production by root tissue was two to three times higher than needle tissue and decreased with decreasing water potential. ACC concentrations in needles of both seed sources decreased as water potential began decreasing. Below -1.4 MPa, ACC levels started increasing (Texas source) or remained constant until -2.8 MPa (Virginia source) at which time its level increased three-fold. Mean ACC levels in root tissue [122 nmol (g dry weight)⁻¹] were slightly higher than the mean levels in the needle tissue [92 nmol (g dry weight) ⁻¹]; roots apparently were more efficient in converting it to ethylene since ethylene production was two to three times higher than needle tissue. The modulation of ethylene synthesis by ACC synthase and ethyleneforming enzyme appeared to be influenced by stress level, organ and seed source.