Nitrate control of root hydraulic properties in plants: translating local information to whole plant response

The sessile lifestyle of plants constrains their ability to acquire mobile nutrients such as nitrate. Whereas proliferation of roots might help in the longer term, nitrate-rich patches can shift rapidly with mass flow of water in the soil. A mechanism that allows roots to follow and capture this source of mobile nitrogen would be highly desirable. Here, we report that variation in nitrate concentration around roots induces an immediate alteration of root hydraulic properties such that water is preferentially absorbed from the nitrate-rich patch. Further, we show that this coupling between nitrate availability and water acquisition results from changes in cell membrane hydraulic properties and is directly related to intracellular nitrate concentrations. Split-root experiments in which nitrate was applied to a portion of the root system showed that the response is both localized and reversible, resulting in rapid changes in water uptake to the portions of the roots exposed to the nitrate-rich patch. At the same time, water uptake by roots not supplied with nitrate was reduced. We believe that the increase in root hydraulic conductance in one part causes a decline of water uptake in the other part due to a collapse in the water potential gradient driving uptake. The translation of local information, in this case nitrate concentration, into a hydraulic signal that can be transmitted rapidly throughout the plant and thus coordinate responses at the whole plant level, represents an unexpected, higher level physiological interaction that precedes the level of gene expression.     

Nitrate induction of root hydraulic conductivity in maize is not correlated with aquaporin expression

Some plant species can increase the mass flow of water from the soil to the root surface in response to the appearance of nitrate in the rhizosphere by increasing root hydraulic conductivity. Such behavior can be seen as a powerful strategy to facilitate the uptake of nitrate in the patchy and dynamically changing soil environment. Despite the significance of such behavior, little is known about the dynamics and mechanism of this phenomenon. Here we examine root hydraulic response of nitrate starved Zea mays (L.) plants after a sudden exposure to 5 mM NO₃ ⁻ solution. In all cases the treatment resulted in a significant increase in pressure-induced (pressure gradient ~ 0.2 MPa) flow across the root system by ~50% within 4 h. Changes in osmotic gradient across the root were approximately 0.016 MPa (or 8.5%) and thus the results could only be explained by a true change in root hydraulic conductance. Anoxia treatment significantly reduced the effect of nitrate on xylem root hydraulic conductivity indicating an important role for aquaporins in this process. Despite a 1 h delay in the hydraulic response to nitrate treatment, we did not detect any change in the expression of six ZmPIP1 and seven ZmPIP2 genes, strongly suggesting that NO₃ ⁻ ions regulate root hydraulics at the protein level. Treatments with sodium tungstate (nitrate reductase inhibitor) aimed at resolving the information pathway regulating root hydraulic properties resulted in unexpected findings. Although this treatment blocked nitrate reductase activity and eliminated the nitrate-induced hydraulic response, it also produced changes in gene expression and nitrate uptake levels, precluding us from suggesting that nitrate acts on root hydraulic properties via the products of nitrate reductase.     

Plant morphology and root hydraulics are altered by nutrient deficiency in Pistacia lentiscus (L.)

The plants in arid and semiarid areas are often limited by water and nutrients. Morpho-functional adjustments to improve nutrient capture may have important implications on plant water balance, and on plant capacity to withstand drought. Several studies have shown that N and P deficiencies may decrease plant hydraulic conductance. Surprisingly, studies on the implications of nutrient limitations on water use in xerophytes are scarce. We have evaluated the effects of strong reductions in nitrogen and phosphorus availability on morphological traits and hydraulic conductance in seedlings of a common Mediterranean shrub, Pistacia lentiscus L.. Nitrogen deficiency resulted in a decrease in aboveground biomass accumulation, but it did not affect belowground biomass accumulation or root morphology. Phosphorus-deficient plants showed a decrease in leaf area, but no changes in aboveground biomass. Root length, root surface area, and specific root length were higher in phosphorus-deficient plants than in control plants. Nitrogen and phosphorus deficiency reduced both root hydraulic conductance and root hydraulic conductance scaled by total root surface area. On the other hand, nutrient limitations did not significantly affect root conductance per unit of foliar surface area. Thus, adaptation to low nutrient availability did not affect seedling capacity for maintaining water supply to leaves. The implications for drought resistance and survival during seedling establishment in semi-arid environments are discussed.     

Effects of multiple stresses on radish growth and resource allocation

Summary Acclimation of wild radish plants to a simultaneous combination of SO₂ fumigation and decreasing nitrate availability was investigated. Plants were grown for 24 d under continuous daytime (10h) exposure to 0 or 0.4 ppm SO₂ and were grown in a nutrient solution with stable nitrate concentrations of 100 M for the first 15 d, 50 M from day 15 to day 19, and 25 M from day 19 to day 24. Analysis of relative growth rates (RGR) showed that radish plants responded rapidly to changes in nitrate availability and that SO₂ treatment affected those responses. Shoot RGR of plants from both treatments and root RGR of control plants showed rapid declines and subsequent recoveries in response to decreasing nitrate availability. Root RGR of SO₂-treated plants declined rapidly in response to decreased nitrate availability, but did not recover as quickly or completely as root RGR of control plants. Analysis of specific leaf weights and tissue nitrogen concentrations showed that control plants had significantly higher amounts of nitrogen in tissues after nitrate availability was lowered, and had higher rates of nitrate uptake in comparison to SO₂-treated plants; especially when nitrate availability was highest. Furthermore, control plants had temporarily higher rates of root respiration in comparison to SO₂-treated plants, suggesting that control plants temporarily allocated more resources to physiological processes occurring in roots, such as nutrient uptake. Although SO₂-induced changes in growth and resource allocation of plants were relatively small, it was probable that SO₂ treatment of radish plants affected plant nitrogen balance, and subsequently affected the ability of plants to respond to decreased nitrate availibility, by affecting resource partitioning to nitrate uptake and root growth.     

Effects of synthetic plant growth retardants and abscisic acid on root functions of Brassica rapa plants exposed to low root-zone temperature

Possible interactions of two synthetic plant-growth retardants during the short-term response of Brassica rapa L. ssp. oleifera (DC.) Metzger plants to low root-zone temperature were investigated by pretreating with mefluidide or paclobutrazol. Water and solute transfers were studied by measuring xylem sap volume flow (under root pressure exudation) and ion flow from the roots. Relations with nitrate uptake rate were also considered. Root pretreatment with paclobutrazol strongly restricted the cold-inducible processes which normally restore water and solute flow from the root xylem. Paclobutrazol decreased the rates of nitrate uptake and exudation flow from the root xylem (principally by reducing root hydraulic conductivity) with dramatic consequences for ion flow, especially that of nitrate.
The effects of root ABA pretreatment on plant response to root cooling were then studied separately or in association with a pretreatment with paclobutrazol. Despite a slight decrease in nitrate uptake rate, ABA pretreatment of the roots enabled the plant to develop rapid mechanisms for adaptation to cold constraint at the root level. Moreover, this action of exogenous ABA greatly reduced the effect of a simultaneous paclobutrazol pretreatment and partly restored water and solute flows.
Thus, the improvement of plant resistance to cold conditions brought about by treatments with mefluidide and paclobutrazol (previously shown in long-term experiments) cannot simply be explained by their short-term effects.     

Statistical modeling of nitrogen-dependent modulation of root system architecture in Arabidopsis thaliana

Plant root development is strongly affected by nutrient availability. Despite the importance of structure and function of roots in nutrient acquisition,statistical modeling approaches to evaluate dynamic and temporal modulations of root system architecture in response to nutrient availability have remained as widely open and exploratory areas in root biology. In this study,we developed a statistical modeling approach to investigate modulations of root system architecture in response to nitrogen availability. Mathematical models were designed for quantitative assessment of root growth and root branching phenotypes and their dynamic relationships based on hierarchical con figuration of primary and lateral roots formulating the fishbone-shaped root system architecture in Arabidopsis thaliana. Time-series datasets reporting dynamic changes in root developmental traits on different nitrate or ammonium concentrations were generated for statistical analyses. Regression analyses unraveled key parameters associated with:(i) inhibition of primary root growth under nitrogen limitation or on ammonium;(ii) rapid progression of lateral root emergence in response to ammonium; and(iii) inhibition of lateral root elongation in the presence of excess nitrate or ammonium. This study provides a statistical framework for interpreting dynamic modulation of root system architecture,supported by metaanalysis of datasets displaying morphological responses of roots to diverse nitrogen supplies.     

Possible involvement of protein phosphorylation in aluminum-responsive malate efflux from wheat root apex

In many plants, efflux of organic anions from roots has been proposed as one of the major Al resistance mechanisms. However it remains unknown how plants regulate efflux of organic anions in response to Al. In this study, the regulatory mechanisms of Al-responsive malate efflux in wheat (Triticum aestivum) were characterized focusing on the role of protein phosphorylation. Al-resistant wheat (cv Atlas) initiated malate efflux at 5 min after addition of Al, and this response was sensitive to temperature. K-252a, a broad range inhibitor of protein kinases, effectively blocked the Al-induced malate efflux accompanied with an increased accumulation of Al and intensified Al-induced root growth inhibition. A transient activation of a 48-kD protein kinase and an irreversible repression of a 42-kD protein kinase were observed preceding the initiation of malate efflux, and these changes were canceled by K-252a. Malate efflux was accompanied with a rapid decrease in the contents of organic anions in the root apex, such as citrate, succinate, and malate but with no change in the contents of inorganic anions such as chloride, nitrate, and phosphate. These results suggest that protein phosphorylation is involved in the Al-responsive malate efflux in the wheat root apex and that the organic anion-specific channel might be a terminal target that responds to Al signaling mediated by phosphorylation.     

Role of root systems of eastern larch and white spruce in response to flooding

Abstract. Soil flooding causes rapid reductions in transpiration, stomatal conductance and photosynthesis of many woody plants, which can decrease growth and ultimately result in plant death. This study was conducted to determine the role of the root system in the flooding response. Eastern larch ( Larix laricina ) seedlings were grown in Plexiglas tubes in which water uptake by flooded and unflooded roots was measured independently. Further flooding studies were conducted with eastern larch and white spruce ( Picea glauca ) in which stems were girdled. Root hydraulic properties were analysed using pressure-flow relationships. Transpiration rates of partially flooded plants declined more slowly than fully-flooded plants. Water uptake by unflooded roots of partially flooded seedlings increased momentarily with flooding. After lOd, flooding caused little change in root hydraulic conductance, a decrease in root system reflection coefficient, and an increase in osmotic permeability. Stem girdling had little effect on stomatal conductance and transpiration in comparison to flooding effects. The response of plant tops to flooding appears to be xylem-mediated and in proportion to the amount of root system flooded. Root hydraulic conductance appears to be unaffected by flooding except for a possible temporary increase on the first day following flooding treatments.     

Distinct roles of electric and hydraulic signals on the reaction of leaf gas exchange upon re-irrigation in Zea mays L

The hypothesis that electric and hydraulic long-distance signals modify photosynthesis and stomatal aperture upon re-irrigation in intact drought-stressed plants was examined. Maize plants (Zea mays L.) were exposed to drought conditions by decreasing the soil water content to 40-50% of field capacity. The decrease in water content resulted in a decline in stomatal conductance to 50-60% of the level in well-watered plants. Re-irrigation of the plants initiated both hydraulic and electric signals, followed by a two-phase response of the net CO2 uptake rate and stomatal conductance of leaves. The transitional first phase (phase 1) is characterized by a rapid decrease in both levels. In the second phase (phase 2), both parameters gradually increase to levels above those of drought-stressed plants. Elimination of either the hydraulic signal by compensatory pressure application to the root system, or of the electric signal by cooling of the leaf blade gave evidence that the two signals (1) propagated independently from each other and (2) triggered the two-phase response in leaf gas exchange. The results provided evidence that the hydraulic signal initiated a hydropassive decrease in stomatal aperture and for the involvement of electric signals in the regulation of photosynthesis of drought-stressed plants.     

The Effect of VA Mycorrhizal Infection and Phosphorus Status on Sunflower Hydraulic and Stomatal Properties

Koide, R. 1985. The effect of VA mycorrhizal infection and phosphorusstatus on sunflower hydraulic and stomatal properties.—J. exp. Bot. 36: 1087–1098. Mycorrhizal (M) and non-mycorrhizal (NM) sunflower plants weregrown in a soil of low phosphorus availability (with and withoutphosphorus amendment) and in a soil of moderate phosphorus availability(without phosphorus amendment). Using the Ohm's law analogyand measured leaf water potentials, stem water potentials, andtranspiration rates, hydraulic resistances were calculated forthe whole plant, leaf, and below leaf components. Mycorrhizalinfection (as high as 89%) was shown to have no effect on theintrinsic hydraulic properties of the soil/plant system overa wide range of transpiration rates in either soil when M andNM plants of equivalent root length were compared. When grownin the soil of moderate phosphorus availability, calculatedhydraulic resistances under given environmental conditions werethe same for M and NM plants, as were stomatal resistances andtranspiration rates. When grown in the soil of low phosphorusavailability, calculated values of hydraulic resistance werelower for M plants than for NM plants under given sets of environmentalconditions. These differences in calculated hydraulic resistancewere not due to a difference in the intrinsic hydraulic propertiesof M and NM plants. The differences were evident because stomatalresistances were lower and transpiration rates higher for Mplants and because hydraulic resistance varied inversely withtranspiration rate. When plants of significantly greater rootlength were compared to plants of lesser root length, the calculatedhydraulic resistances under given environmental conditions weremuch lower for the plants of greater root length. This differencewas largely due to a difference in the intrinsic hydraulic propertiesbetween large and small plants, and not because of differencesin transpiration rate. The elevated transpiration rates exhibitedby M plants were attributed to an enhanced phosphorus status.Short term phosphorus amendments made to phosphorus-deficientNM plants improved transpiration; transpiration rates were similarfor M and NM plants before NM plants became phosphorus-deficient,and phosphorus-amended M and NM plants had similar transpirationrates. The data are discussed in relation to other reports ofmycorrhizal influence on hydraulic and stomatal resistances.Possible mechanisms for the influence of infection on stomatalresistance are also briefly discussed. Key words: Hydraulic resistance, stomatal resistance, mycorrhizas     

Nitrate and phosphate availability and distribution have different effects on root system architecture of Arabidopsis

Plant root systems can respond to nutrient availability and distribution by changing the three-dimensional deployment of their roots: their root system architecture (RSA). We have compared RSA in homogeneous and heterogeneous nitrate and phosphate supply in Arabidopsis. Changes in nitrate and phosphate availability were found to have contrasting effects on primary root length and lateral root density, but similar effects on lateral root length. Relative to shoot dry weight (DW), primary root length decreased with increasing nitrate availability, while it increased with increasing phosphate supply. Lateral root density remained constant across a range of nitrate supplies, but decreased with increasing phosphate supply. In contrast, lateral root elongation was suppressed both by high nitrate and high phosphate supplies. Local supplies of high nitrate or phosphate in a patch also had different effects. Primary root growth was not affected by a high nitrate patch, but growth through a high phosphate patch reduced primary root growth after the root left the patch. A high nitrate patch induced an increase in lateral root density in the patch, whereas lateral root density was unaffected by a high phosphate patch. However, both phosphate- and nitrate-rich patches induced lateral root elongation in the patch and suppressed it outside the patch. This co-ordinated response of lateral roots also occurs in soil-grown plants exposed to a nutrient-rich patch. The auxin-resistant mutants axrl, axr4 and aux1 all showed the wild-type lateral root elongation responses to a nitrate-rich patch, suggesting that auxin is not required for this response.     

Short-term responses of Brassica rapa plants to low root temperature: Effects on nitrate uptake and its translocation to the shoot

Brassica rapa L. plants were grown hydroponically for 5 or 6 weeks at 20°C and then half batches of plants were transferred to tanks in which the root temperature was lowered decrementally over 1 h to 7°C. Changes in nitrate uptake rate (NUR) and nitrate transfer from roots were studied in relation to transpiration and root pressure xylem exudation flow rates over a 48- or 72-h period. The response of plants following the root temperature decrease was biphasic. During phase 1, NUR and water and solute flow rates through the root decreased sharply. Coping mechanisms came into operation during phase 2, and tended to offset the effects of low temperature. The 3-h cold-treated roots exhibited a very low NUR but 48-h cold-treated roots partly recovered their ability to absorb nitrate. Transpiration rate decreased more slowly (during 24 h) than both root xylem exudation and parameters of root conductivity (during 6 h). Beyond these respective times, transpiration rate was balanced while root xylem exudation clearly increased, but without returning to the level of control plants. Nitrate transfer to the root xylem was strongly and rapidly affected by low root temperature, but the subsequent readjustment was such that no or little difference compared with the control was apparent after 48 h. Water and solute flows were strongly decreased when nitrate was replaced by chloride in the culture solution during exudation sampling. The major role of nitrate in root hydraulic conductivity and root xylem exudation is discussed.     

Nitrate reduction and nitrogen fixation in symbiotic association Rhizobium-legumes

The inhibitory effect of nitrate on nitrogenase activity in root nodules of legume plants has been known for a long time. The major factor inducing changes in nitrogenase activity is the concentration of free oxygen inside nodules. Oxygen availability in the infected zone of nodule is limited, among others, by the gas diffusion resistance in nodule cortex. The presence of nitrate may cause changes in the resistance to O2 diffusion. The aim of this paper is to review literature data concerning the effect of nitrate on the symbiotic association between rhizobia and legume plants, with special emphasis on nitrogenase activity. Recent advances indicate that symbiotic associations of Rhizobium strains characterized by a high nitrate reductase activity are less susceptible to inhibition by nitrate. A thesis may be put forward that dissimilatory nitrate reduction, catalyzed by bacteroid nitrate reductase, significantly facilitates the symbiotic function of bacteroids.     

Modification of leaf apoplastic pH in relation to stomatal sensitivity to root-sourced abscisic acid signals

The confocal microscope was used to determine the pH of the leaf apoplast and the pH of microvolumes of xylem sap. We quantified variation in leaf apoplast and sap pH in relation to changes in edaphic and atmospheric conditions that impacted on stomatal sensitivity to a root-sourced abscisic acid signal. Several plant species showed significant changes in the pH of both xylem sap and the apoplast of the shoot in response to environmental perturbation. Xylem sap leaving the root was generally more acidic than sap in the midrib and the apoplast of the leaf. Increasing the transpiration rate of both intact plants and detached plant parts resulted in more acidic leaf apoplast pHs. Experiments with inhibitors suggested that protons are removed from xylem sap as it moves up the plant, thereby alkalinizing the sap. The more rapid the transpiration rate and the shorter the time that the sap resided in the xylem/apoplastic pathway, the smaller the impact of proton removal on sap pH. Sap pH of sunflower (Helianthus annuus) and Commelina communis did not change significantly as soil dried, while pH of tomato (Lycopersicon esculentum) sap increased as water availability in the soil declined. Increasing the availability of nitrate to roots also significantly alkalinized the xylem sap of tomato plants. This nitrogen treatment had the effect of enhancing the sensitivity of the stomatal response to soil drying. These responses were interpreted as an effect of nitrate addition on sap pH and closure of stomata via an abscisic acid-based mechanism.     

A role for auxin redistribution in the responses of the root system architecture to phosphate starvation in Arabidopsis

The changes in root system architecture (RSA) triggered by phosphate (P) deprivation were studied in Arabidopsis (Arabidopsis thaliana) plants grown for 14 d on 1 mM or 3 microM P. Two different temporal phases were observed in the response of RSA to low P. First, lateral root (LR) development was promoted between days 7 and 11 after germination, but, after day 11, all root growth parameters were negatively affected, leading to a general reduction of primary root (PR) and LR lengths and of LR density. Low P availability had contrasting effects on various stages of LR development, with a marked inhibition of primordia initiation but a strong stimulation of activation of the initiated primordia. The involvement of auxin signaling in these morphological changes was investigated in wild-type plants treated with indole-3-acetic acid or 2,3,5-triiodobenzoic acid and in axr4-1, aux1-7, and eir1-1 mutants. Most effects of low P on RSA were dramatically modified in the mutants or hormone-treated wild-type plants. This shows that auxin plays a major role in the P starvation-induced changes of root development. From these data, we hypothesize that several aspects of the RSA response to low P are triggered by local modifications of auxin concentration. A model is proposed that postulates that P starvation results in (1) an overaccumulation of auxin in the apex of the PR and in young LRs, (2) an overaccumulation of auxin or a change in sensitivity to auxin in the lateral primordia, and (3) a decrease in auxin concentration in the lateral primordia initiation zone of the PR and in old laterals. Measurements of local changes in auxin concentrations induced by low P, either by direct quantification or by biosensor expression pattern (DR5::beta-glucuronidase reporter gene), are in line with these hypotheses. Furthermore, the observation that low P availability mimicked the action of auxin in promoting LR development in the alf3 mutant confirmed that P starvation stimulates primordia emergence through increased accumulation of auxin or change in sensitivity to auxin in the primordia. Both the strong effect of 2,3,5-triiodobenzoic acid and the phenotype of the auxin-transport mutants (aux1, eir1) suggest that low P availability modifies local auxin concentrations within the root system through changes in auxin transport rather than auxin synthesis.     

Distribution of xylem hydraulic resistance in fruiting truss of tomato influenced by water stress

In this study xylem hydraulic resistances of peduncles (truss stalk), pedicels (fruit stalk) and the future abscission zone (AZ) halfway along the pedicel of tomato (Lycopersicon esculentum L.) plants were directly measured at different stages of fruit development, in plants grown under two levels of water availability in the root environment. The xylem hydraulic connection between shoot and fruits has previously been investigated, but contradictory conclusions were drawn about the presence of a flow resistance barrier in the pedicel. These conclusions were all based on indirect functional measurements and anatomical observations of water-conducting tissue in the pedicel. In the present study, by far the largest resistances were measured in the AZ where most individual vessels ended. Plants grown at low water availability in the root environment had xylem with higher hydraulic resistances in the peduncle and pedicel segments on both sides of the AZ, while the largest increase in hydraulic resistance was measured in the AZ. During fruit development hydraulic resistances in peduncle and pedicel segments decreased on both sides of the AZ, but tended to increase in the AZ. The overall xylem hydraulic resistance between the shoot and fruit tended to increase with fruit development because of the dominating role of the hydraulic resistance in the AZ. It is discussed whether the xylem hydraulic resistance in the AZ of tomato pedicels in response to water stress and during fruit development contributes to the hydraulic isolation of fruits from diurnal cycles of water stress in the shoot.     

Control of leaf expansion in sunflower (Helianthus annuus L.) by nitrogen nutrition

Seedlings of Helianthus annuus L. were grown at an initiallyhigh relative nitrate supply rate (0.27 mol N mol N^–1d^–1). The supply was subsequently reduced to a low rate(0.04 mol N mol N^–1 d^–1). The response of leaf areadevelopment to this abrupt decrease in nitrate availabilitywas characterized by following the expansion of the primaryand secondary leaf pairs. The timing of the drop in nitratesupply was when cell division in the epidermis of the primaryleaf pair was largely complete. Reducing the availability ofnitrate had a strong effect on leaf area expansion. The finalleaf size of the primary leaf pair was affected indicating aneffect of nitrate availability on cell expansion. By the endof the experiment the secondary leaf pair was only one-thirdthe area of that on control seedlings. The role of epidermalcell turgor pressure in this growth response was assessed bydirect measurements with a miniature cell pressure probe. Noreduction in cell turgor pressure following the decrease innitrate availability was detected. It is concluded that a reductionin turgor pressure was not responsible for the reduction inleaf area expansion and it is suggested that reduced cell expansionwas due to changes in cell wall properties. Concentrations ofleaf and root abscisic acid increased following the reductionin nitrate availability. Key words: Abscisic acid, cell size, cell turgor pressure, nitrate, nitrogen, relative rate of nitrate supply