PdEPF1 regulates water‐use efficiency and drought tolerance by modulating stomatal density in poplar

Water deficiency is a critical environmental condition that is seriously reducing global plant production. Improved water‐use efficiency (WUE) and drought tolerance are effective strategies to address this problem. In this study, PdEPF1, a member of the EPIDERMAL PATTERNING FACTOR (EPF) family, was isolated from the fast‐growing poplar clone NE‐19 [Populus nigra × (Populus deltoides × Populus nigra)]. Significantly, higher PdEPF1 levels were detected after induction by dehydration and abscisic acid. To explore the biological functions of PdEPF1, transgenic triploid white poplars (Populus tomentosa ‘YiXianCiZhu B385’) overexpressing PdEPF1 were constructed. PdEPF1 overexpression resulted in increased water deficit tolerance and greater WUE. We confirmed that the transgenic lines with greater instantaneous WUE had approximately 30% lower transpiration but equivalent CO₂ assimilation. Lower transpiration was associated with a 28% reduction in abaxial stomatal density. PdEPF1 overexpression not only strongly enhanced WUE, but also greatly improved drought tolerance, as measured by the leaf relative water content and water potential, under limited water conditions. In addition, the growth of these oxPdEPF1 plants was less adversely affected by reduced water availability than plants with a higher stomatal density, indicating that plants with a low stomatal density may be well suited to grow in water‐scarce environments. Taken together, our data suggest that PdEPF1 improves WUE and confers drought tolerance in poplar; thus, it could be used to breed drought‐tolerant plants with increased production under conditions of water deficiency.     

PeCHYR1, a ubiquitin E3 ligase from Populus euphratica,enhances drought tolerance via ABA‐induced stomatal closure by ROS production in Populus

Drought, a primary abiotic stress, seriously affects plant growth and productivity. Stomata play a vital role in regulating gas exchange and drought adaptation. However, limited knowledge exists of the molecular mechanisms underlying stomatal movement in trees. Here, PeCHYR1, a ubiquitin E3 ligase, was isolated from Populus euphratica, a model of stress adaptation in forest trees. PeCHYR1 was preferentially expressed in young leaves and was significantly induced by ABA (abscisic acid) and dehydration treatments. To study the potential biological functions of PeCHYR1, transgenic poplar 84K (Populus alba × Populus glandulosa) plants overexpressing PeCHYR1 were generated. PeCHYR1 overexpression significantly enhanced H₂O₂ production and reduced stomatal aperture. Transgenic lines exhibited increased sensitivity to exogenous ABA and greater drought tolerance than that of WT (wild‐type) controls. Moreover, up‐regulation of PeCHYR1 promoted stomatal closure and decreased transpiration, resulting in strongly elevated WUE (water use efficiency). When exposed to drought stress, transgenic poplar maintained higher photosynthetic activity and biomass accumulation. Taken together, these results suggest that PeCHYR1 plays a crucial role in enhancing drought tolerance via ABA‐induced stomatal closure caused by hydrogen peroxide (H₂O₂) production in transgenic poplar plants.     

Nitrogen fertilization enhances water‐use efficiency in a saline environment

Effects of salinity and nutrients on carbon gain in relation to water use were studied in the grey mangrove, Avicennia marina, growing along a natural salinity gradient in south‐eastern Australia. Tall trees characterized areas of seawater salinities (fringe zone) and stunted trees dominated landward hypersaline areas (scrub zone). Trees were fertilized with nitrogen (+N) or phosphorus (+P) or unfertilized. There was no significant effect of +P on shoot growth, whereas +N enhanced canopy development, particularly in scrub trees. Scrub trees maintained greater CO₂ assimilation per unit water transpired (water‐use efficiency, WUE) and had lower nitrogen‐use efficiency (NUE; CO₂ assimilation rate per unit leaf nitrogen) than fringe trees. The CO₂ assimilation rates of +N trees were similar to those in other treatments, but were achieved at lower transpiration rates, stomatal conductance and intercellular CO₂ concentrations. Maintaining comparable assimilation rates at lower stomatal conductance requires greater ribulose 1·5‐bisphosphate carboxylase/oxygenase activity, consistent with greater N content per unit leaf area in +N trees. Hence, +N enhanced WUE at the expense of NUE. Instantaneous WUE estimates were supported by less negative foliar δ¹³C values for +N trees and scrub control trees. Thus, nutrient enrichment may alter the structure and function of mangrove forests along salinity gradients.     

Water‐use efficiency in a semi‐arid woodland with high rainfall variability

As the ratio of carbon uptake to water use by vegetation, water‐use efficiency (WUE) is a key ecosystem property linking global carbon and water cycles. It can be estimated in several ways, but it is currently unclear how different measures of WUE relate, and how well they each capture variation in WUE with soil moisture availability. We evaluated WUE in an Acacia‐dominated woodland ecosystem of central Australia at various spatial and temporal scales using stable carbon isotope analysis, leaf gas exchange and eddy covariance (EC) fluxes. Semi‐arid Australia has a highly variable rainfall pattern, making it an ideal system to study how WUE varies with water availability. We normalized our measures of WUE across a range of vapour pressure deficits using g₁, which is a parameter derived from an optimal stomatal conductance model and which is inversely related to WUE. Continuous measures of whole‐ecosystem g₁ obtained from EC data were elevated in the 3 days following rain, indicating a strong effect of soil evaporation. Once these values were removed, a close relationship of g₁ with soil moisture content was observed. Leaf‐scale values of g₁ derived from gas exchange were in close agreement with ecosystem‐scale values. In contrast, values of g₁ obtained from stable isotopes did not vary with soil moisture availability, potentially indicating remobilization of stored carbon during dry periods. Our comprehensive comparison of alternative measures of WUE shows the importance of stomatal control of fluxes in this highly variable rainfall climate and demonstrates the ability of these different measures to quantify this effect. Our study provides the empirical evidence required to better predict the dynamic carbon–water relations in semi‐arid Australian ecosystems.     

Ecological interactions and the fitness effect of water‐use efficiency: Competition and drought alter the impact of natural MPK12 alleles in Arabidopsis

The presence of substantial genetic variation for water‐use efficiency (WUE) suggests that natural selection plays a role in maintaining alleles that affect WUE. Soil water deficit can reduce plant survival, and is likely to impose selection to increase WUE, whereas competition for resources may select for decreased WUE to ensure water acquisition. We tested the fitness consequences of natural allelic variation in a single gene (MPK12) that influences WUE in Arabidopsis, using transgenic lines contrasting in MPK12 alleles, under four treatments; drought/competition, drought/no competition, well‐watered/competition, well‐watered/no competition. Results revealed an allele × environment interaction: Low WUE plants performed better in competition, resulting from increased resource consumption. Contrastingly, high WUE individuals performed better in no competition, irrespective of water availability, presumably from enhanced water conservation and nitrogen acquisition. Our findings suggest that selection can influence MPK12 evolution, and represents the first assessment of plant fitness resulting from natural allelic variation at a single locus affecting WUE.     

Chloride as a macronutrient increases water‐use efficiency by anatomically driven reduced stomatal conductance and increased mesophyll diffusion to CO2

Chloride (Cl^?) has been recently described as a beneficial macronutrient, playing specific roles in promoting plant growth and water‐use efficiency (WUE). However, it is still unclear how Cl^? could be beneficial, especially in comparison with nitrate (NO₃^?), an essential source of nitrogen that shares with Cl^? similar physical and osmotic properties, as well as common transport mechanisms. In tobacco plants, macronutrient levels of Cl^? specifically reduce stomatal conductance (g_s) without a concomitant reduction in the net photosynthesis rate (A_N). As stomata‐mediated water loss through transpiration is inherent in the need of C₃ plants to capture CO₂, simultaneous increase in photosynthesis and WUE is of great relevance to achieve a sustainable increase in C₃ crop productivity. Our results showed that Cl^?‐mediated stimulation of larger leaf cells leads to a reduction in stomatal density, which in turn reduces g_s and water consumption. Conversely, Cl^? improves mesophyll diffusion conductance to CO₂ (g_m) and photosynthetic performance due to a higher surface area of chloroplasts exposed to the intercellular airspace of mesophyll cells, possibly as a consequence of the stimulation of chloroplast biogenesis. A key finding of this study is the simultaneous improvement of A_N and WUE due to macronutrient Cl^? nutrition. This work identifies relevant and specific functions in which Cl^? participates as a beneficial macronutrient for higher plants, uncovering a sustainable approach to improve crop yield.     

Evidence of changing intrinsic water‐use efficiency under rising atmospheric CO2 concentrations in Boreal Fennoscandia from subfossil leaves and tree ring δ13C ratios

Investigating the many internal feedbacks within the climate system is a vital component of the effort to quantify the full effects of future anthropogenic climate change. The stomatal apertures of plants tend to close and decrease in number under elevated CO₂ concentrations, increasing water‐use efficiency (WUE) and reducing canopy evapotranspiration. Experimental and modelling studies reveal huge variations in these changes such that the warming associated with reduced evapotranspiration (known as physiological forcing) is neither well understood or constrained. Palaeo‐observations of changes in stomatal response and plant WUE under rising CO₂ might be used to better understand the processes underlying the physiological forcing feedback and to link measured changes in plant WUE to a specific physiological change in stomata. Here we use time series of tree ring (Pinus sylvestris L.) δ¹³C and subfossil leaf (Betula nana L.) measurements of stomatal density and geometry to derive records of changes in intrinsic water‐use efficiency (iWUE) and maximum stomatal conductance in the Boreal zone of northern Finland and Sweden. We investigate the rate of change in both proxies, over the recent past. The independent lines of evidence from these two different Boreal species indicate increased iWUE and reduced maximum stomatal conductance of similar magnitude from preindustrial times (ca. ad 1850) to around ad 1970. After this maximum stomatal conductance continues to decrease to ad 2000 in B. nana but iWUE in P. sylvestris reaches a plateau. We suggest that northern boreal P. sylvestris might have reached a threshold in its ability to increase WUE as CO₂ rises.     

Contrasting water sources and water‐use efficiency in coexisting desert plants in two saline‐sodic soils in northwest China

• Soil degradation resulting from various types of salinity is a major environmental problem, especially in arid and semiarid regions. Exploring the water‐related physiological traits of halophytes is useful for understanding the mechanisms of salt tolerance. This knowledge could be used to rehabilitate degraded arid lands.
• To investigate whether different types of salinity influence the water sources and water‐use efficiency of desert plants (Karelinia caspia, Tamarix hohenackeri, Nitraria sibirica, Phragmites australis, Alhagi sparsifolia, Suaeda microphylla, Kalidium foliatum) in natural environments, we measured leaf gas exchange, leaf carbon and xylem oxygen isotope composition and soil oxygen isotope composition at neutral saline‐sodic site (NSS) and alkaline saline‐sodic site (ASS) in northwest China.
• The studied plants had different xylem water oxygen isotope compositions (δ¹⁸O) and foliar carbon isotope compositions (δ¹³C), indicating that desert plants coexist through differentiation in water use patterns. Compared to that at the NSS site, the stem water in K. caspia, A. sparsifolia and S. microphylla was depleted in ¹⁸O at the ASS site, which indicates that plants can switch to obtain water from deeper soil layers when suffering environmental stress from both salinity and alkalinisation. Alhagi sparsifolia had higher δ¹³C at the ASS site than at the NSS site, while K. caspia and S. microphylla had lower δ¹³C, which may have resulted from interspecific differences in plant alkali and salt tolerance ability.
• Our results suggest that under severe salinity and alkalinity, plants may exploit deeper soil water to avoid ion toxicity resulting from high concentrations of soluble salts in the superficial soil layer. In managed lands, it is vital to select and cultivate different salt‐tolerant or alkali‐tolerant plant species in light of local conditions.

     

Co‐expression of tonoplast Cation/H+ antiporter and H+‐pyrophosphatase from xerophyte Zygophyllum xanthoxylum improves alfalfa plant growth under salinity,drought and field conditions

Salinity and drought are major environmental factors limiting the growth and productivity of alfalfa worldwide as this economically important legume forage is sensitive to these kinds of abiotic stress. In this study, transgenic alfalfa lines expressing both tonoplast NXH and H⁺‐PPase genes, ZxNHX and ZxVP1‐1 from the xerophyte Zygophyllum xanthoxylum L., were produced via Agrobacterium tumefaciens‐mediated transformation. Compared with wild‐type (WT) plants, transgenic alfalfa plants co‐expressing ZxNHX and ZxVP1‐1 grew better with greater plant height and dry mass under normal or stress conditions (NaCl or water‐deficit) in the greenhouse. The growth performance of transgenic alfalfa plants was associated with more Na⁺, K⁺ and Ca²⁺ accumulation in leaves and roots, as a result of co‐expression of ZxNHX and ZxVP1‐1. Cation accumulation contributed to maintaining intracellular ions homoeostasis and osmoregulation of plants and thus conferred higher leaf relative water content and greater photosynthesis capacity in transgenic plants compared to WT when subjected to NaCl or water‐deficit stress. Furthermore, the transgenic alfalfa co‐expressing ZxNHX and ZxVP1‐1 also grew faster than WT plants under field conditions, and most importantly, exhibited enhanced photosynthesis capacity by maintaining higher net photosynthetic rate, stomatal conductance, and water‐use efficiency than WT plants. Our results indicate that co‐expression of tonoplast NHX and H⁺‐PPase genes from a xerophyte significantly improved the growth of alfalfa, and enhanced its tolerance to high salinity and drought. This study laid a solid basis for reclaiming and restoring saline and arid marginal lands as well as improving forage yield in northern China.     

Drought response of mesophyll conductance in forest understory species – impacts on water‐use efficiency and interactions with leaf water movement

Regulation of stomatal (g_s) and mesophyll conductance (g_m) is an efficient means for optimizing the relationship between water loss and carbon uptake in plants. We assessed water‐use efficiency (WUE)‐based drought adaptation strategies with respect to mesophyll conductance of different functional plant groups of the forest understory. Moreover we aimed at assessing the mechanisms of and interactions between water and CO₂ conductance in the mesophyll. The facts that an increase in WUE was observed only in the two species that increased g_m in response to moderate drought, and that over all five species examined, changes in mesophyll conductance were significantly correlated with the drought‐induced change in WUE, proves the importance of g_m in optimizing resource use under water restriction. There was no clear correlation of mesophyll CO₂ conductance and the tortuosity of water movement in the leaf across the five species in the control and drought treatments. This points either to different main pathways for CO₂ and water in the mesophyll either to different regulation of a common pathway.     

Conformational preferences of a short Aib/Ala‐based water‐soluble peptide as a function of temperature

The amino acid Aib predisposes a peptide to be helical with context‐dependent preference for either 3₁₀‐ or α‐ or a mixed helical conformation. Short peptides also show an inherent tendency to be unfolded. To characterize helical and unfolded states adopted by water‐soluble Aib‐containing peptides, the conformational preference of Ac‐Ala‐Aib‐Ala‐Lys‐Ala‐Aib‐Lys‐Ala‐Lys‐Ala‐Aib‐Tyr‐NH₂ was determined by CD, NMR and MD simulations as a function of temperature. Temperature‐dependent CD data indicated the contribution of two major components, each an admixture of helical and extended/polyproline II structures. Both right‐ and left‐handed helical conformations were detected from deconvolution of CD data and ¹³C NMR experiments. The presence of a helical backbone, more pronounced at the N‐terminal, and a temperature‐induced shift in α‐helix/3₁₀‐helix equilibrium, more pronounced at the C‐terminal, emerged from NMR data. Starting from polyproline II, the N‐terminal of the peptide folded into a helical backbone in MD simulations within 5 ns at 60°C. Longer simulations showed a mixed‐helical backbone to be stable over the entire peptide at 5°C while at 60°C the mixed‐helix was either stable at the N‐terminus or occurred in short stretches through out the peptide, along with a significant population of polyproline II. Our results point towards conformational heterogeneity of water‐soluble Aib‐based peptide helices and the associated subtleties. The problem of analyzing CD and NMR data of both left‐ and right‐handed helices are discussed, especially the validity of the ellipticity ratio [θ]₂₂₂/[θ]₂₀₇, as a reporter of α‐/3₁₀‐ population ratio, in right‐ and left‐handed helical mixtures. Proteins 2009. © 2008 Wiley‐Liss, Inc.