Response of cassava leaf area expansion to water deficit: cell proliferation, cell expansion and delayed development

BACKGROUND AND AIMS: Cassava (Manihot esculenta) is an important food crop in the tropics that has a high growth rate in optimal conditions, but also performs well in drought-prone climates. The objectives of this work were to determine the effects of water deficit and rewatering on the rate of expansion of leaves at different developmental stages and to evaluate the extent to which decreases in cell proliferation, expansion, and delay in development are responsible for reduced growth. METHODS: Glasshouse-grown cassava plants were subjected to 8 d of water deficit followed by rewatering. Leaves at 15 developmental stages from nearly full size to meristematic were sampled, and epidermal cell size and number were measured on leaves at four developmental stages. KEY RESULTS: Leaf expansion and development were nearly halted during stress but resumed vigorously after rewatering. In advanced-stage leaves (Group 1) in which development was solely by cell expansion, expansion resumed after rewatering, but not sufficiently for cell size to equal that of controls at maturity. In Group 2 (cell proliferation), relative expansion rate and cell proliferation were delayed until rewatering, but then recovered partially, so that loss of leaf area was due to decreased cell numbers per leaf. In Group 3 (early meristematic development) final leaf area was not affected by stress, but development was delayed by 4-6 d. On a plant basis, the proportion of loss of leaf area over 26 d attributed to leaves at each developmental stage was 29, 50 and 21 % in Group 1, 2 and 3, respectively. CONCLUSIONS: Although cell growth processes were sensitive to mild water deficit, they recovered to a large extent, and much of the reduction in leaf area was caused by developmental delay and a reduction in cell division in the youngest, meristematic leaves.     

Control of Leaf Expansion by Nitrogen Nutrition in Sunflower Plants : ROLE OF HYDRAULIC CONDUCTIVITY AND TURGOR

Nitrogen nutrition strongly affected the growth rate of young sunflower (Helianthus annuus L.) leaves. When plants were grown from seed on either of two levels of N availability, a 33% decrease in tissue N of expanding leaves was associated with a 75% overall inhibition of leaf growth. Almost all of the growth inhibition resulted from a depression of the daytime growth rate. Measurements of pressure-induced water flux through roots showed that N deficiency decreased root hydraulic conductivity by about half. Thus, N deficiency lowered the steady-state water potential of expanding leaves during the daytime when transpiration was occurring. As a result, N-deficient leaves were unable to maintain adequate turgor for growth in the daytime. N deficiency also decreased the hydraulic conductivity for water movement into expanding leaf cells in the absence of transpiration, but growth inhibition at night was much less than in the daytime. N nutrition had no detectable effects on plastic extensibility or the threshold turgor for growth.     

The expression pattern of plasma membrane aquaporins in maize leaf highlights their role in hydraulic regulation

Leaves are key organs for evaporation and photosynthesis and play a crucial role in plant growth and development. In order to function properly, they need to maintain a balanced water content. Water movement through a leaf occurs by a combination of different pathways: water can follow an apoplastic route through the cell wall or a cell-to-cell route via the symplastic and transcellular paths. As aquaporins (AQPs) play an important role in regulating transcellular water flow and CO(2) conductance, studies on AQP mRNA and protein expression in leaves are essential to better understand their role in these physiological processes. Here, we quantified and localized the expression of Zea mays plasma membrane aquaporins (ZmPIPs, plasma membrane intrinsic proteins) in the leaf using quantitative RT-PCR and immunodetection. All ZmPIP genes except ZmPIP2;7 were expressed in leaves. Expression was found to be dependent on the developmental stage of the leaf tissue, with, in general, an increase in expression at the end of the elongation zone and a decrease in mature leaf tissue. These data correlated with the cell water permeability, as determined using a protoplast swelling assay. The diurnal expression of ZmPIPs was also investigated and expression was found to be higher during the first hours of the light period than at night. Immunocytochemical localization of four ZmPIP isoforms indicated that they are involved in leaf radial water movement, in particular in vascular bundles and the mesophyll.     

Dynamic Relation between Expansion and Cellular Turgor in Growing Grape (Vitis vinifera L.) Leaves

Measurements of the growth and water relations of expanding grape (Vitis vinifera L.) leaves have been used to determine the relationship between leaf expansion rate and leaf cell turgor. Direct measurement of turgor on the small (approximately 15 micrometer diameter) epidermal cells over the midvein of expanding grape leaves was made possible by improvements in the pressure probe technique. Leaf expansion rate and leaf water status were perturbed by environmentally induced changes in plant transpiration. After establishing a steady state growth rate, a step decrease in plant transpiration resulted in a rapid and large increase in leaf cell turgor (0.25 megapascal in 5 minutes), and leaf expansion rate. Subsequently, leaf expansion rate returned to the original steady state rate with no change in cell turgor. These results indicate that the expansion rate of leaves may not be strongly related to the turgor of the leaf cells, and that substantial control of leaf expansion rate, despite changes in turgor, may be part of normal plant function. It is suggested that a strictly physical interpretation of the parameters most commonly used to describe the relationship between turgor and growth in plant cells (cell wall extensibility and yield threshold) may be inappropriate when considering the process of plant cell expansion.     

Experimental Studies of the Factors Controlling Transpiration: III. THE INTERRELATIONS BETWEEN TRANSPIRATION RATE, STOMATAL MOVEMENT, AND LEAF-WATER CONTENT

Transpiration rates of single leaves of Pelargonium and wheatwere measured under constant conditions of light, temperature,and air flow. Concurrently, stomatal movement was followed withthe resistance porometer during cycles of changing water contentof the leaf and changes induced by light and darkness. Stomatalmovement was found to exert a large controlling influence onthe transpiration rate, whereas water content had an extremelysmall or negligible effect. An approximately inverse linearrelation between transpiration rate and logarithm of resistanceto viscous flow through the leaf is believed to be the resultantof an inverse curvilinear relationship between the diffusiveconductance of the stomata and log. leaf resistance and thedecreasing difference of vapour pressure arising from the highertranspiration rates with increasing stomatal conductances. Nevertheless,the relation demonstrates that the transpiration rate is influencedby the degree of stomatal opening throughout its entire range. There was some evidence of lower transpiration rates duringand after recovery from wilting than before wilting. This isattributed to a decrease in a cell-wall conductance, the evaporatingsurface being located within the cell wall. During wilting partiallyirreversible contraction of the cell wall occurs. There wasalso evidence of slow changes in cell volume at full turgidityattributable to plastic flow. These occurred when the leaf wastransferred from environments of a high to low potential forevaporation. Extensive movement of the stomata followed changes in leaf water,passive opening resulting from decrease and closure from increaseof leaf water. It is suggested that the direction and extentof stomatal changes induced by water deficits is a consequenceof the rate of change of leaf water content and not of the absolutevalues. The stomata also showed an enhanced tendency to closein dry moving air following a period of wilting even after theleaf had regained turgidity.     

Effects of nitrogen deprivation on cell division and expansion in leaves of Ricinus communis L.

The effects of nitrogen deprivation on leaf extension, cell numbers and epidermal cell size were followed in leaves of Ricinus communis L. The extent to which reductions in final cell number or final epidermal cell size contributed to the reduction in final leaf size depended on the developmental stage of the leaf at the time of N deprivation. In leaves which already had their full complement of cells (leaf 2), the reduction in final leaf size following nitrogen deprivation was associated with a reduction in final cell size. In leaves that were at earlier stages of development at the onset of N deprivation (leaves 3 and 4), the reduction in final leaf size was greater than in leaf 2. In these younger leaves, the final cell size was even smaller than in leaf 2, but the greatest contribution to reduced final leaf size was a reduction in the number of cells produced. This accounted for approximately 80% of the reduction in final leaf size in leaf 4. During leaf development, the contribution from different tissue layers to the total cell number changed. In the smallest leaf sizes, the contribution from upper and lower epidermis and spongy parenchyma was greater than that from palisade parenchyma. As the leaf size increased, cells in the palisade parenchyma continued to divide for longer than in the other layers. At final leaf size, the contribution from the different tissue layers to total cell number was the same for leaves 2, 3 and 4, irrespective of N treatment. In these final leaf structures, palisade parenchyma contributed 60% of the total cell number. Thus, although nitrogen deprivation affected leaf size variously through cell division and cell expansion, depending on leaf developmental stage at the time of nitrogen deprivation, the ratio of cell numbers and sizes in different tissue layers, at final leaf size, was unaffected.     

Water relations in competitive interactions of Mediterranean grasses and shrubs

In Mediterranean ecosystems, competition between opportunistic grasses and slower-growing woody species may affect the speed and path of ecosystem recovery and the success of restoration plantings after natural or human-induced disturbance. In this experiment, competitive interactions between Mediterranean annual and perennial grass species (Avena fatua and Brachypodium retusum, respectively) and an important Mediterranean shrub (Rosmarinus offlcinalis) were examined under semi-controlled conditions simulating wet and dry Mediterranean rainfall regimes. The identity of the grass competitor and the level of water availability in the plots interacted to produce differing rates of R. offlcinalis growth but similar levels of mortality. In particular, competition with the perennial grass resulted in very low rates of R. offlcinalis growth at both irrigation levels. Measurements of soil water content showed that both grasses reduced soil moisture to low levels, though this effect was temporary in the case of the winter annual grass. Resistance to hydraulic flow in roots was highest in the perennial grass, smaller but of similar magnitude in the shrub, and much lower in the annual grass. Transpirational response to decreasing leaf water potential was a quick, sharp drop in conductance in R. offlcinalis, in contrast to a moderated decline from much lower initial transpiration rates in B. retusum. The annual grass largely maintained both leaf water potential and transpiration through leaf-tip senescence and death. Quantification of the rate of hydric recuperation of leaves after irrigation of drought-stressed plants showed that the perennial grass recovered at a rate four times that of R. offlcinalis, suggesting a strategy for making quick use of rare summer rains that may contribute to its competitive success. The appropriateness of planting or suppressing grasses in restoration of disturbed sites in Mediterranean Spain is discussed.     

Abscisic acid triggers whole-plant and fruit-specific mechanisms to increase fruit calcium uptake and prevent blossom end rot development in tomato fruit

Calcium (Ca) uptake into fruit and leaves is dependent on xylemic water movement, and hence presumably driven by transpiration and growth. High leaf transpiration is thought to restrict Ca movement to low-transpiring tomato fruit, which may increase fruit susceptibility to the Ca-deficiency disorder, blossom end rot (BER). The objective of this study was to analyse the effect of reduced leaf transpiration in abscisic acid (ABA)-treated plants on fruit and leaf Ca uptake and BER development. Tomato cultivars Ace 55 (Vf) and AB2 were grown in a greenhouse environment under Ca-deficit conditions and plants were treated weekly after pollination with water (control) or 500 mg l(-1) ABA. BER incidence was completely prevented in the ABA-treated plants and reached values of 30-45% in the water-treated controls. ABA-treated plants had higher stem water potential, lower leaf stomatal conductance, and lower whole-plant water loss than water-treated plants. ABA treatment increased total tissue and apoplastic water-soluble Ca concentrations in the fruit, and decreased Ca concentrations in leaves. In ABA-treated plants, fruit had a higher number of Safranin-O-stained xylem vessels at early stages of growth and development. ABA treatment reduced the phloem/xylem ratio of fruit sap uptake. The results indicate that ABA prevents BER development by increasing fruit Ca uptake, possibly by a combination of whole-plant and fruit-specific mechanisms.     

Sensitivity of soybean leaf development to water deficits

Abstract. Drought effects on the final leaf area of individual leaves were hypothesized to depend on the leaf developmental stage at which drought occurred. To evaluate this hypothesis, final leaf area and cell number were measured for soybean ( Glycine max (L.) Merr.) leaves that were at different stages of development when single or cyclical drought treatment was imposed. Leaf emergence rate from the meristem, as depicted by changes in the plastochron index, was not as sensitive as leaf expansion to cyclical droughts. For leaf expansion, small leaves, once they emerged from the meristem, suffered larger decreases in growth than leaves undergoing rapid leaf area expansion. Decreases in final leaf area as a result of a cyclical drought were correlated with decreases in final cell number. Decreases resulting from a single 8-d drought were dependent on the age of the leaf at the time of drought, because small leaves were found to have proportionately larger decreases in final cell number and area than larger leaves. These results indicated that age-dependent leaf responses to drought are based on the relative activity of cell division and expansion at the time stress was imposed.     

Photosynthesis and water-relation traits of the summer annual C4 grasses, Eleusine indica and Digitaria adscendens, with contrasting trampling tolerance

Two summer annual C₄ grasses with different trampling susceptibilities were grown as potted plants, and diurnal leaf gas exchange and leaf water potential in each grass were compared. The maximum net photosynthetic rate, leaf conductance and transpiration rate were higher in the trampling-tolerant Eleusine indica (L.) Gaertn. than in trampling sensitive Digitaria adscendens (H. B. K.) Henr. Leaf water potential was much lower in E. indica than in D. adscendens. There were no differences in soil-to-leaf hydraulic conductance and leaf osmotic potential at full turgor as obtained by pressure–volume analysis. However, the bulk modulus of elasticity in cell walls was higher in E. indica leaves than in D. adscendens leaves. This shows that the leaves of E. indica are less elastic. Therefore, the rigid cell walls of E. indica leaves reduced leaf water potential rapidly by decreasing the leaf water content, supporting a high transpiration rate with high leaf conductance. In trampled habitats, such lowering of leaf water potential in E. indica might play a role in water absorption from the compacted soil. In contrast, the ability of D. adscendens to colonize dry habitats such as coastal sand dunes appears to be due to its lower transpiration rate and its higher leaf water potential which is not strongly affected by decreasing leaf water content.     

Phytochrome regulates cellular response plasticity and the basic molecular machinery of leaf development

Plants are plastic organisms that optimize growth in response to a changing environment. This adaptive capability is regulated by external cues, including light, which provides vital information about the habitat. Phytochrome photoreceptors detect far-red light, indicative of nearby vegetation, and elicit the adaptive shade-avoidance syndrome (SAS), which is critical for plant survival. Plants exhibiting SAS are typically more elongated, with distinctive, small, narrow leaf blades. By applying SAS-inducing end-of-day far-red (EoD FR) treatments at different times during Arabidopsis (Arabidopsis thaliana) leaf 3 development, we have shown that SAS restricts leaf blade size through two distinct cellular strategies. Early SAS induction limits cell division, while later exposure limits cell expansion. This flexible strategy enables phytochromes to maintain control of leaf size through the proliferative and expansion phases of leaf growth. mRNAseq time course data, accessible through a community resource, coupled to a bioinformatics pipeline, identified pathways that underlie these dramatic changes in leaf growth. Phytochrome regulates a suite of major development pathways that control cell division, expansion, and cell fate. Further, phytochromes control cell proliferation through synchronous regulation of the cell cycle, DNA replication, DNA repair, and cytokinesis, and play an important role in sustaining ribosome biogenesis and translation throughout leaf development.

Phytochrome regulates leaf blade plasticity through two alternative cellular response strategies and concertedly coordinates gene expression of the basic cellular machinery of leaf development.     

Multiple steps of leaf thickening during sun‐leaf formation in Arabidopsis

Plant morphological and physiological traits exhibit plasticity in response to light intensity. Leaf thickness is enhanced under high light (HL) conditions compared with low light (LL) conditions through increases in both cell number and size in the dorsoventral direction; however, the regulation of such phenotypic plasticity in leaf thickness (namely, sun‐ or shade‐leaf formation) during the developmental process remains largely unclear. By modifying observation techniques for tiny leaf primordia in Arabidopsis thaliana, we analysed sun‐ and shade‐leaf development in a time‐course manner and found that the process of leaf thickening can be divided into early and late phases. In the early phase, anisotropic cell elongation and periclinal cell division on the adaxial side of mesophyll tissue occurred under the HL conditions used, which resulted in the dorsoventral growth of sun leaves. Anisotropic cell elongation in the palisade tissue is triggered by blue‐light irradiation. We discovered that anisotropic cell elongation processes before or after periclinal cell division were differentially regulated independent of or dependent upon signalling through blue‐light receptors. In contrast, during the late phase, isotropic cell expansion associated with the endocycle, which determined the final leaf thickness, occurred irrespective of the light conditions. Sucrose production was high under HL conditions, and we found that sucrose promoted isotropic cell expansion and the endocycle even under LL conditions. Our analyses based on this method of time‐course observation addressed the developmental framework of sun‐ and shade‐leaf formation.     

Control of plant growth by nitrogen: differences between cereals and broadleaf species

Abstract. In four dicotyledonous species low levels of N strongly inhibited leaf expansion during the day but had little or no effect at night. In contrast, daytime and night-time expansion were equally affected in four cereal species. The results support the general concept that in dicotyledons, N controls leaf expansion through its effects on hydraulic conductivity. In such N-limited plants, water deficits generated by transpiration may inhibit daytime cell expansion. In cereals, cell expansion and transpiration occur in separate zones of the leaf and are apparently unrelated.
Growth analysis showed that low levels of N inhibited leaf area growth more strongly in dicotyledons than in cereals, but had similar effects on net assimilation rates of plants in the two groups. As a result, dry matter production was more efficient in cereals than in dicotyledons when N was limiting.     

Water relations and leaf expansion: importance of time scale

The role of leaf water relations in controlling cell expansion in leaves of water-stressed maize and barley depends on time scale. Sudden changes in leaf water status, induced by sudden changes in humidity, light and soil salinity, greatly affect leaf elongation rate, but often only transiently. With sufficiently large changes in salinity, leaf elongation rates are persistently reduced. When plants are kept fully turgid throughout such sudden environmental changes, by placing their roots in a pressure chamber and raising the pressure so that the leaf xylem sap is maintained at atmospheric pressure, both the transient and persistent changes in leaf elongation rate disappear. All these responses show that water relations are responsible for the sudden changes in leaf elongation rate resulting from sudden changes in water stress and putative root signals play no part. However, at a time scale of days, pressurization fails to maintain high rates of leaf elongation of plants in either saline or drying soil, indicating that root signals are overriding water relations effects. In both saline and drying soil, pressurization does raise the growth rate during the light period, but a subsequent decrease during the dark results in no net effect on leaf growth over a 24 h period. When transpirational demand is very high, however, growth-promoting effects of pressurization during the light period outweigh any reductions in the dark, resulting in a net increase in growth of pressurized plants over 24 h. Thus leaf water status can limit leaf expansion rates during periods of high transpiration despite the control exercised by hormonal effects on a 24 h basis.     

Foliar water uptake: Processes,pathways, and integration into plant water budgets

Nearly all plant families, represented across most major biomes, absorb water directly through their leaves. This phenomenon is commonly referred to as foliar water uptake. Recent studies have suggested that foliar water uptake provides a significant water subsidy that can influence both plant water and carbon balance across multiple spatial and temporal scales. Despite this, our mechanistic understanding of when, where, how, and to what end water is absorbed through leaf surfaces remains limited. We first review the evidence for the biophysical conditions necessary for foliar water uptake to occur, focusing on the plant and atmospheric water potentials necessary to create a gradient for water flow. We then consider the different pathways for uptake, as well as the potential fates of the water once inside the leaf. Given that one fate of water from foliar uptake is to increase leaf water potentials and contribute to the demands of transpiration, we also provide a quantitative synthesis of observed rates of change in leaf water potential and total fluxes of water into the leaf. Finally, we identify critical research themes that should be addressed to effectively incorporate foliar water uptake into traditional frameworks of plant water movement.     

Water relations of cotton flower petals and fruit

Water needed for expansion is believed to enter plant tissue in response to a growth-induced water potential gradient that occurs as turgor is reduced during relaxation of cell walls or in response to increased solutes. Under water stress, the cotton flower petal continues to expand when all leaves on the plant are wilted and new leaf expansion has ceased in the shoot tips. This study was undertaken to determine if water for expansion entered the petal in response to a gradient or to increased solutes. Water potentials of cotton petal, leaf, bract and fruit were determined pre-dawn and midday in dryland and irrigated field plots. The mechanism by which petal expansion occurs appears not to be associated with a growth-induced water potential gradient or to increased solutes because the gradient is reversed from that needed to drive expansion. The water potential of the petal tissues was consistently higher than that of the subtending leaves and bracts both during and after anthesis, and under different water stress conditions. How this reversal in water potential gradient is established and maintained should provide insight into mechanisms involved in growth during water stress.     

Alteration of transpiration rate, by changing air vapour pressure deficit, influences leaf extension rate transiently in Miscanthus

A controlled environment chamber for whole plants is described in which vapour pressure deficit (VPD) and temperature can be controlled independently. Plant responses to changes in VPD at constant temperature were measured in terms of leaf extension and plant transpiration rates. Manipulation of VPD independently of temperature was shown to be capable of altering the leaf extension rates of the C4 grass Miscanthus x giganteus grown in hydroponics. The effects of VPD on leaf extension are attributed to changes in transpiration rate and hence leaf water status. It was found that, at a temperature of 20C, the influence of a fixed change in VPD was proportionally less than those observed at temperatures which are close to the threshold for growth (between 6 and 10C). These responses are discussed in relation to our current understanding of the mechanisms of cell growth. The fact that the VPD effects on leaf expansion rates were largely transient suggest that simple models driven by temperature alone are adequate to predict leaf expansion within the temperature range 6-20°C, for this genotype of Miscanthus, in the field.Key words: Leaf growth, Miscanthus x giganteus, temperature, vapour pressure deficit, C4 plants.      

Short-term responses of leaf growth rate to water deficit scale up to whole-plant and crop levels: an integrated modelling approach in maize

Physiological and genetic studies of leaf growth often focus on short-term responses, leaving a gap to whole-plant models that predict biomass accumulation, transpiration and yield at crop scale. To bridge this gap, we developed a model that combines an existing model of leaf 6 expansion in response to short-term environmental variations with a model coordinating the development of all leaves of a plant. The latter was based on: (1) rates of leaf initiation, appearance and end of elongation measured in field experiments; and (2) the hypothesis of an independence of the growth between leaves. The resulting whole-plant leaf model was integrated into the generic crop model APSIM which provided dynamic feedback of environmental conditions to the leaf model and allowed simulation of crop growth at canopy level. The model was tested in 12 field situations with contrasting temperature, evaporative demand and soil water status. In observed and simulated data, high evaporative demand reduced leaf area at the whole-plant level, and short water deficits affected only leaves developing during the stress, either visible or still hidden in the whorl. The model adequately simulated whole-plant profiles of leaf area with a single set of parameters that applied to the same hybrid in all experiments. It was also suitable to predict biomass accumulation and yield of a similar hybrid grown in different conditions. This model extends to field conditions existing knowledge of the environmental controls of leaf elongation, and can be used to simulate how their genetic controls flow through to yield.     

The role of auxin-binding protein 1 in the expansion of tobacco leaf cells

Tobacco leaf was used to investigate the mechanism of action of auxin-binding protein 1 (ABP1). The distributions of free auxin, ABP1, percentage of leaf nuclei in G2 and the amount of auxin-inducible growth were each determined in control tobacco leaves and leaves over-expressing Arabidopsis ABP1. These parameters were compared with growth of tobacco leaves, measured both spatially and temporally throughout the entire expansion phase. Within a defined window of leaf development, juvenile leaf cells that inducibly expressed Arabidopsis ABP1 prematurely advanced nuclei to the G2 phase. The ABP1-induced increase in cell expansion occured before the advance to the G2 phase, indicating that the ABP1-induced G2 phase advance is an indirect effect of cell expansion. The level of ABP1 was highest at the position of maximum cell expansion, maximum auxin-inducible growth and where the free auxin level was the lowest. In contrast, the position of maximum cell division correlated with higher auxin levels and lower ABP1 levels. Consistent with the correlations observed in leaves, tobacco cells (BY-2) in culture displayed two dose-dependent responses to auxin. At a low auxin concentration, cells expanded, while at a relatively higher concentration, cells divided and incorporated [3H]-thymidine. Antisense suppression of ABP1 in these cells dramatically reduced cell expansion with negligible effect on cell division. Taken together, the data suggest that ABP1 acts at a relatively low level of auxin to mediate cell expansion, whereas high auxin levels stimulate cell division via an unidentified receptor.