Two water molds can grow without measurable turgor pressure

The water molds Achlya bisexualis Coker and Saprolegnia ferax (Gruithuisen) Thuret (Class: Oomycetes) normally grow in the form of slender hyphae with up to 0.8 MPa (8 bar) of internal pressure. Models of plant cell growth indicate that this turgor pressure drives the expansion of the cell wall. However, under conditions of prolonged osmotic stress, these species were able to grow in the absence of measurable turgor. Unpressurized cells of A. bisexualis grew in the form of a plasmodium-like colony on solid media, and produced a multinucleate yeast-like phase in liquid. By contrast, the morphology of S. ferax was unaffected by the loss of turgor, and the mold continued to generate tip-growing hyphae. Measurements of cell wall strength indicate that these microorganisms produce a very fluid wall in the region of surface growth, circumventing the usual requirement for turgor.Abbreviations DAPI 4,6-diamidino-2-phenylindole - PEG polyethylene glycol This work was supported by National Science Foundation grant DCB 90-17130.     

Lectin activity of the nucleocytoplasmic EUL protein from Arabidopsis thaliana

The Euonymus lectin (EUL) domain was recognized as the structural motif for a novel class of putative carbohydrate binding proteins. Confocal microscopy demonstrated that the lectin from Euonymus europaeus (EEA) as well as the EUL protein from Arabidopsis thaliana (ArathEULS3) are located in the nucleocytoplasmic compartment of the plant cell. ArathEULS3 as well as its EUL domain were successfully expressed in Pichia pastoris and purified. The EUL domain from Arabidopsis interacts with glycan structures containing Lewis Y, Lewis X and lactosamine, indicating that it can be considered a true lectin domain. Despite the high sequence identity between the EUL domains in EEA and ArathEULS3, both domains recognize different carbohydrate structures.     

Salt tolerance in the halophyte Suaeda maritima L. Dum.

Osmotic potentials and individual epidermal cell turgor pressures were measured in the leaves of seedlings of Suaeda maritima growing over a range of salinities. Leaf osmotic potentials were lower (more negative) the higher the salt concentration of the solution and were lowest in the youngest leaves and stem apices, producing a gradient of osmotic potential towards the apex of the plant. Epidermal cell turgor pressures were of the order of 0.25 to 0.3 MPa in the youngest leaves measured, decreasing to under 0.05 MPa for the oldest leaves. This pattern of turgor pressure was largely unaffected by external salinity. Calculation of leaf water potential indicated that the gradient between young leaves and the external medium was not altered by salinity, but with older leaves, however, this gradient diminished from being the same as that for young leaves in the absence of NaCl, to under 30% of this value at 400 mM NaCl. These results are discussed in relation to the growth response of S. maritima.     

Changes in turgor pressure in response to increases in external NaCl concentration in the gas-vacuolate cyanobacterium Microcystis sp.

Changes in cell turgor pressure have been followed in cells of Microcystis sp. transferred to culture medium containing added NaCl at osmolalities of 30–1,500 mosmol kg^-1 ( 74–3,680 kPa). Upon upshock turgor decreased, due to osmotically-induced water loss from the cell. However, partial recovery of turgor was then observed in illuminated cells, with maximum turgor regain in media containing 30–500 mosmol kg^-1 NaCl. The lightdependent recovery of turgor pressure was completed within 60 min, with no evidence of further changes in cell turgor up to 24 h. This is the first direct evidence that turgor regulation may occur in a prokaryotic organism. Short-term increases in cell K⁺ content were also observed upon upshock in NaCl, indicating that turgor regain may involve a turgorsensitive K⁺ uptake system. Estimation of internal K⁺ concentration in cells transferred to 250 mosmol kg^-1 NaCl showed that changes in cell K⁺ may account for at least half of the observed turgor regain up to 60 min.     

Turgor pressure and water transport properties of suspension-cultured cells of Chenopodium rubrum L.

The turgor pressure and water relation parameters were determined in single photoautotrophically grown suspension cells and in individual cells of intact leaves of Chenopodium rubrum using the miniaturized pressure probe. The stationary turgor pressure in suspension-cultured cells was in the range of betwen 3 and 5 bar. From the turgor pressure relaxation process, induced either hydrostatically (by means of the pressure probe) or osmotically, the halftime of water exchange was estimated to be 20±10 s. No polarity was observed for both ex- and endosmotic water flow. The volumetric elastic modulus, , determined from measurements of turgor pressure changes, and the corresponding changes in the fractional cell volume was determined to be in the range of between 20 and 50 bar. increases with increasing turgor pressure as observed for other higher plant and algal cells. The hydraulic conductivity, Lp, is calculated to be about 0,5–2·10^–6 cm s^–1 bar^–1. Similar results were obtained for individual leaf cells of Ch. rubrum. Suspension cells immobilized in a cross-linked matrix of alginate (6 to 8% w/w) revealed the same values for the half-time of water exchange and for the hydraulic conductivity, Lp, provided that the turgor pressure relaxation process was generated hydrostatically by means of the pressure probe. Thus, it can be concluded that the unstirred layer from the immobilized matrix has no effect on the calculation of Lp from the turgor pressure relaxation process, using the water transport equation derived for a single cell surrounded by a large external volume. By analogy, this also holds true for Lp-values derived from turgor pressure changes generated by the pressure probe in a single cell within the leaf tissue. The fair similarity between the Lp-values measured in mesophyll cells in situ and mesophyll-like suspension cells suggests that the water transport relations of a cell within a leaf are not fundamentally different from those measured in a single cell.     

Direct turgor pressure measurements in individual leaf cells of Tradescantia virginiana

The water relations of leaves of Tradescantia virginiana were studied using the miniaturized pressure probe (Hüsken, E. Steudle, Zimmermann, 1978 Plant Physiol. 61, 158–163). Under well-watered conditions cell turgor pressures, P _o, ranged from 2 to 8 bar in epidermal cells. In subsidiary cells P _o was about 1.5 to 4.5 bar and in mesophyll cells about 2 to 3.5 bar. From the turgor pressure, relaxation induced in individual cells by changing the turgor pressure directly by means of the pressure probe, the half-time of water exchange was measured to be between 3 and 100 s for the epidermal, subsidiary, and mesophyll cells. The volumetric elastic modulus, , of individual cells was determined by changing the cell volume by a defined amount and simultaneously measuring the corresponding change in cell turgor pressure. The values for the elastic modulus for epidermal, subsidiary, and mesophyll cells are in the range of 40 to 240 bar, 30 to 200 bar, and 6 to 14 bar, respectively. Using these values, the hydraulic conductivity, L _p, for the epidermal, subsidiary, and mesophyll cells is calculated from the turgor pressure relaxation process (on the basis of the thermodynamics of irreversible processes) to be between 1 and 55·10^-7 cm s^-1 bar^-1. The data for the volumetric elastic modulus of epidermal and subsidiary cells indicate that the corresponding elastic modulus for the guard cells should be considerably lower due to the large volume changes of these cells during opening or closing. Recalculation of experimental data obtained by K. Raschke (1979, Encycl. Plant Physiol. N.S., vol. 7, pp 383–441) on epidermal strips of Vicia faba indicates that the elastic modulus of guard cells of V. faba is in the order of 40–80 bar for closed stomata. However, with increasing stomatal opening, i.e., increasing guard cell volume, decreases. Therefore, in our opinion Raschke's results would indicate a relationship between guard cell volume and which would be inverse to that for plant cells known in the literature. assumes values between 20–40 bar when the guard cell colume is soubled.     

Stress relaxation of cell walls and the yield threshold for growth

Theory predicts that, for growing plant cells isolated from a supply of water, stress relaxation of the cell wall should decrease cell turgor pressure (P) until the yield threshold for cell expansion is reached. This prediction was tested by direct P measurements of pea (Pisum sativum L.) stem cortical cells before and after excision of the growing region and isolation of the growing tissue from an external water supply. Cell P was measured with the micro-pressure probe under conditions which eliminated transpiration. Psychrometric measurements of water potential confirmed the pressureprobe measurements. Following excision, P of the growing cells decreased in 1 h by an average of 1.8 bar to a mean plateau value of 2.8 bar, and remained constant thereafter. Treatment with 10^-5 M indole-3-acetic acid or 10^-5 M fusicoccin (known growth stimulants) accelerated the rate of P relaxation, whereas various treatments which inhibit growth slowed down or completely stopped P relaxation in apical segments. In contrast, P of basal (nongrowing) segments gradually increased because of absorption of solutes from the cell-wall free space of the tissue. Such solute absorption also occurred in apical segments, but wall relaxation held P at the yield threshold in those segments which were isolated from an external water supply. These results provide a new and rapid method for measuring the yield threshold and they show that P in intact growing pea stems exceeds the yield threshold by about 2 bar. Wall relaxation is shown here to affect the water potential and turgor pressure of excised growing segments. In addition, solute release and absorption upon excision may influence the water potential and turgor pressure of nongrowing excised plant tissues.Abbreviations and symbols IAA indole-3-acetic acid - P turgor pressure - SE standard error of the mean - water potential     

Derivation of a weighted-average reflection coefficient for mesophyll cell membranes of Kalanchoë daigremontiana

In a study with the crassulacean acid metabolism (CAM) plant Kalanchoë daigremontiana Hamet et Perr. using the pressure probe, Rygol et al. (1987, Planta 172, 487–4493) calculated a value for the reflection coefficient () for malate of 0.6. This value was derived from the relationship between measured changes in cell turgor pressure and malic-acid concentration, and would imply that malate was a relatively ineffective osmoticum. Here we show that the calculation of Rygol et al. (1987) involved the implicit assumption that xylem tension was constant with changing cell turgor pressure and osmotic pressure. This has been shown not to be the case using the pressure-chamber technique. We present an alternative method of deriving a weighted-mean value of a for K. daigremontiana and show that it is not significantly different from 1.0.Part of this work was carried out at the University of Edinburgh, to whom we are grateful for facilities, with funding from the Agricultural and Food Research Council, UK. Murphy is grateful to the board of management of the Glasstone Benefaction for financial support at the University of Oxford. We thank Prof. U. Zimmermann for his comments on an earlier version of this paper.     

Replicase proteins as determinants of phloem-dependent long-distance movement of tobamoviruses in tobacco

Summary The turgor pressure of growing pollen tubes of the lily (Lilium longiflorum Thunb.) has been recorded using a turgor pressure probe. Insertion of the probe's micropipette was routinely accomplished, providing recording periods of 20 to 30 min. Probe insertion did not affect tube growth. The stable turgor values ranged between 0.1 and 0.4 MPa, the mean value being 0.209 ± 0.064 MPa (n=106). A brief increase in turgor, generated by injection of oil through the pressure probe, caused the tube to burst at its tip. Burst pressures ranged between 0.19 and 0.58 MPa, that is, individual lily pollen tubes do not withstand turgor pressure approaching twice their regular turgor pressure. In contrast, parallel experiments using the incipient plasmolysis technique yielded a mean putative turgor pressure of 0.79 MPa either using sucrose (n=24) or mannitol (n=25). Surprisingly, turgor pressure was not significantly correlated with tube growth rate which ranged from zero to 13 m/min. Nor did it correlate with tube length over the tested range of 100 to 1600 m. In addition the influence of the medium's osmolality was surprisingly low: raising the external osmotic pressure from 0.36 to 1.08 MPa, with sucrose or mannitol, only caused mean turgor pressure to decline from 0.27 to 0.18 MPa. We conclude that growing lily pollen regulates its turgor pressure remarkably well despite substantial variation in tube growth rate, tube length, and osmotic milieu.     

A comparison of cell-wall-yielding properties for two developmental stages of Phycomyces sporangiophores

The yielding properties of the cell wall, irreversible wall extensibility (m) and yield threshold (Y), are determined for stage I sporangiophores of Phycomyces blakesleeanus from in-vivo creep experiments, and compared to the values of m and Y previously determined for stage IVb sporangiophores using the same pressureprobe method (Ortega et al., 1989, Biophys. J. 56, 465). In either stage the sporangiophore enlarges (grows) predominately in length, in a specific region termed the growing zone, but the growth rates of stage I (5–20 urn · min^–1) are smaller than those of stage IVb (30–70 m · min^–1). The results demonstrate that this difference in growth rate is the consequence of a smaller magnitude of m for stage I sporangiophores; the obtained values of P (turgor pressure), Y, and P-Y (effective turgor for irreversible wall extension) for stage I sporangiophores are slightly larger than those of stage IVb sporangiophores. Also, it is shown that the magnitude of m for the stage I sporangiophore is regulated by altering the length of the growing zone, L_g. A relationship between m and L_g is obtained which can account for the difference between values of m determined for stage I and stage IVb sporangiophores. Finally, it is shown that similar changes in the magnitude of m and (which have been used interchangeably in the literature as a measure of irreversible wall extensibility) may not always represent the same changes in the cell-wall properties.Abbreviations and Symbols L length - L_g length of growing zone - m irreversible wall extensibility - P turgor pressure - Y yield threshold - (P-Y) effective turgor for irreversible wall extension - relative irreversible wall extensibility - _g relative irreversible wall extensibility of the growing zone (m/L_g) This work was supported by National Science Foundation grant DCB-8801717 to J.K.E. Ortega.     

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.     

The effect of abscisic acid on cell turgor pressures,solute content and growth of wheat roots

Abscisic acid (ABA) was shown to influence turgor pressure and growth in wheat (Triticum aestivum L.) roots. At a concentrations of 25 mmol·m^-3, ABA increased the turgor pressure of cells located within 1 cm of the tip by up to 450 kPa. At 4 to 5 cm from the root tip this concentration of ABA reduced the turgor pressure of peripheral cells (epidermis and the first few cortical cell layers) to zero or close to zero while that of the inner cells was increased. Increases in sap osmolality were dependent on the concentration of ABA and the effect saturated at 5 mmol·m^-3 ABA. The increase in osmolality took about 4 h and was partly the result of reducing-sugar accumulation. Levels of inorganic cations were not affected by ABA. Root growth was inhibited at ABA concentrations that caused a turgor-pressure increase. The results show that while ABA can affect root cell turgor pressures, this effect does not result in increased root growth.Abbreviation ABA abscisic acid     

Turgor regulation inValonia macrophysa following acute osmotic shock

Summary The marine algaValonia macrophysa an inhabitant of shallow subtropical waters, is subjected to sudden dilutions of external seawater during rain showers. This study describes the mechanisms involved in turgor pressure regulation following acute hyposmotic shock. Turgor regulation is 88% effective and complete within 4 hr following hyposmotic shocks of up to –10 bar. Loss of vacuolar K⁺, Na⁺ and Cl^– accounts for the decrease in vacuolar osmotic pressure associated with turgor regulation. A novel mechanism of turgor regulation is exhibited byValonia macrophysa given hyposmotic shocks greater than about –4 bar. Such an osmotic shock causes cell wall tension to increase above a critical value of about 6×10⁵ dyne/cm, whereupon the protoplasm ruptures and the cell wall stretches irreversibly at a localized site. The protoplasm rupture is suggested by (1) a large abrupt increase in K⁺ efflux (as measured by⁸⁶Rb⁺), (2) a rapid decrease in turgor pressure as measured with a pressure probe, and (3) sudden depolarization of the vacuole potential. Evidence for an increase in cell wall permeability includes efflux from the vacuole of dextran (mol wt 70,000), which normally has a very low cell wall permeability, and scanning electron micrographs which show a trabeculated scar area in the cell wall. This mechanism of turgor regulation is physiologically important because 98% of the cells regained normal growth rate and turgor following acute osmotic shock.     

Day-night changes in leaf water relations associated with the rhythm of crassulacean acid metabolism in Kalanchoë daigremontiana

A study was made of the day-night changes under controlled environmental conditions in the bulk-leaf water relations of Kalanchoë daigremontiana, a plant showing Crassulacean acid metabolism. In addition to nocturnal stomatal opening and net CO₂ uptake, the leaves of well-watered plants showed high rates of gas exchange during the whole of the second part of the light period. Measurements with the pressure chamber showed that xylem tension increased during the night and then decreased towards a minimum at about midday; a significant increase in xylem tension was also seen in the late afternoon. Cell-sap osmotic pressure paralleled leaf malate content and was maximum at dawn and minimum at dusk. The relationship between these two variables indicated that the nocturnally synthesized malate was apparently behaving as an ideal osmoticum. To estimate bulk-leaf turgor pressure, values for water potential were derived by correcting the pressurechamber readings for the osmotic pressure of the xylem sap. This itself was found to depend on the malate content of the leaves. Bulk-leaf turgor pressure changed rhythmically during the day-night cycle; turgor was low during the late afternoon and for most of the night, but increased quickly to a maximum of 0.20 MPa around midday. In water-stressed plants, where net CO₂ uptake was restricted to the dark period, there was also an increase in bulk-leaf turgor pressure at the start of the light period, but of reduced magnitude. Such changes in turgor pressure are likely to be of considerable ecological importance for the water economy of crassulacean-acid-metabolism plants growing in their natural habitats.Abbreviation and symbols CAM Crassulacean acid metabolism - P turgor pressure - osmotic pressure - water potential Dedicated to Professor Dr. H. Ziegler on the occasion of his 60th birthday     

Arabidopsis calcium-binding mitochondrial carrier proteins as potential facilitators of mitochondrial ATP-import and plastid SAM-import

Chloroplasts and mitochondria are central to crucial cellular processes in plants and contribute to a whole range of metabolic pathways. The use of calcium ions as a secondary messenger in and around organelles is increasingly appreciated as an important mediator of plant cell signaling, enabling plants to develop or to acclimatize to changing environmental conditions. Here, we have studied the four calcium-dependent mitochondrial carriers that are encoded in the Arabidopsis genome. An unknown substrate carrier, which was previously found to localize to chloroplasts, is proposed to present a calcium-dependent S-adenosyl methionine carrier. For three predicted ATP/phosphate carriers, we present experimental evidence that they can function as mitochondrial ATP-importers.     

Hydraulic propagation of pressure along immature and mature xylem vessels of roots of Zea mays measured by pressure-probe techniques

Turgor pressure was measured in cortical cells and in xylem elements of excised roots and roots of intact plants of Zea mays L. by means of a cell pressure probe. Turgor of living and hence not fully differentiated late metaxylem (range 0.6–0.8 MPa) was consistently higher than turgor of cortical cells (range 0.4–0.6 MPa) at positions between 40 and 180 mm behind the root tip. Closer to the tip, no turgor difference between the cortex and the stele was measured. The turgor difference indicated that late-metaxylem elements may function as nutrient-storage compartments within the stele. Excised roots were attached to the root pressure probe to precisely manipulate the xylem water potential. Root excision did not affect turgor of cortical cells for at least 8 h. Using the cell pressure probe, the propagation of a hydrostatic pressure change effected by the root pressure probe was recorded in mature and immature xylem elements at various positions along the root. Within seconds, the pressure change propagated along both early and late metaxylems. The half-times of the kinetics, however, were about five times smaller for the early metaxylem, indicating they are likely the major pathway of longitudinal water flow. The hydraulic signal dissipated from the source of the pressure application (cut end of the root) to the tip of the root, presumably because of radial water movement along the root axis. The results demonstrate that the water status of the growth zone and other positions apical to 20 mm is mainly uncoupled from changes of the xylem water potential in the rest of the plant.Abbreviations and Symbols CPP cell pressure probe - EMX early metaxylem - LMX Late metaxylem - P_c cell turgor - P_r root pressure - RPP root pressure probe - t_1/2,c half-time of water exchange across a single cell - t_1/2 half-time of water exchange across multiple cells We thank Antony Matista for his expert assistance in the construction and modification of instruments. The work was supported by grant DCB8802033 from the National Science Foundation and grant 91-37100-6671 from USDA, and by the award of a Feodor Lynen-Fellowship from the Alexander von Humboldt-Foundation (Germany) to J.F.     

Transpiration by an emergent macrophyte: source of water and implications for nutrient supply

The contribution of sediment interstitial water and the water column to the transpiration stream of Myriophyllum aquaticum (Vellozo) Verdcourt was determined to estimate the significance of mass flow in supply of sediment nutrients for plant growth. Sediment interstitial water accounted for about 2% of the water transpired over a 37 day period. Because of the small volume of water that originated in the sediment we concluded that mass flow did not significantly enhance nutrient supply to the roots of M. aquaticum. Relative growth rate (RGR) of adventitious, water roots was greater than whole plant RGR, and RGR of sediment roots was not significantly different from zero, indicating a shift in the biomass allocation after emergence of the apical meristem into the air. Water use, measured by the transpiration coefficient, averaged 260 ml H₂O mg DW^-1, which is similar to C-4 terrestrial plants. M. aquaticum has leaf characteristics commonly associated with xerophytic habitats. These characteristics may be necessary if a high transpiration rate and a mechanical requirement for high cell turgor pressure, required by a reliance upon hydrostatic pressure for support of the aerial stems, are mutually exclusive because of morphological constraints on hydraulic conductivity.     

In vivo extraction of Arabidopsis cell turgor pressure using nanoindentation in conjunction with finite element modeling

Turgor pressure in plant cells is involved in many important processes. Stable and normal turgor pressure is required for healthy growth of a plant, and changes in turgor pressure are indicative of changes taking place within the plant tissue. The ability to quantify the turgor pressure of plant cells in vivo would provide opportunities to understand better the process of pressure regulation within plants, especially when plant stress is considered, and to understand the role of turgor pressure in cellular signaling. Current experimental methods do not separate the influence of the turgor pressure from the effects associated with deformation of the cell wall when estimates of turgor pressure are made. In this paper, nanoindentation measurements are combined with finite element simulations to determine the turgor pressure of cells in vivo while explicitly separating the cell‐wall properties from the turgor pressure effects. Quasi‐static cyclic tests with variable depth form the basis of the measurements, while relaxation tests at low depth are used to determine the viscoelastic material properties of the cell wall. Turgor pressure is quantified using measurements on Arabidopsis thaliana under three pressure states (control, turgid and plasmolyzed) and at various stages of plant development. These measurements are performed on cells in vivo without causing damage to the cells, such that pressure changes may be studied for a variety of conditions to provide new insights into the biological response to plant stress conditions.     

Effect of cell turgor on hydraulic conductivity and elastic modulus of Elodea leaf cells

Water relation parameters of leaf cells of the aquatic plant Elodea densa have been measured using the pressure probe. For cells in both the upper and lower epidermis it was found that the elastic modulus () and the hydraulic conductivity (Lp) were dependent on cell turgor (P). Lp was (7.8±5.5)·10^-7 cm s^-1 bar^-1 (mean±SD; n=22 cells) for P>4 bar in cells of the upper epidermis and was increasing by a factor of up to three for P0 bar. No polarity of water movement or concentration dependence of Lp was observed. For cells of the lower epidermis the Lp-values were similar and the hydraulic conductivity also showed a similar dependence on turgor. No wall ingrowth or wall labyrinths (as in transfer cells) could be found in the cells of the lower epidermis. The elastic modulus () of cells of the upper epidermis could be measured over the whole pressure range (P=0–7 bar) by changing the osmotic pressure of the medium. increased linearly with increasing turgor and ranged between 10 and 150 bar. For cells of the lower epidermis the dependence of on P was similar, although the pressure dependence could not be measured on single cells. The Lp-values are compared with literature data obtained for Elodea by a nuclear magnetic resonance (NMR)-technique. The dependence of Lp on P is discussed in terms of pressure dependent structural changes of the cell membranes and interactions between solute and water transport.Abbreviations P cell turgor pressure - Lp hydraulic conductivity - volumetric elastic modulus - T _1/2 half-time of water exchange of individual cell