Reduction In Turgor Research Articles

Optimization of Osmotic Dehydration of Sapodilla (Achras zapota L.).

Sapodilla (Achras zapota L.) is a fruit with a great nutritional potential; however, its perishable nature is a great obstacle for commercialization/exportation. Herein, osmotic dehydration was applied to sapodilla to reduce post-harvest losses and obtain a stable product with acceptable sensorial characteristics. Initially, a 2³ full-factorial design was performed to determine the effect of temperature (30–50 °C), sucrose concentration (40–60% °Brix) and immersion time (90–240 min) on the moisture loss (ML), solid gain (SG) and dehydration efficiency index (DEI). The samples with higher DEI values were subjected to sensory analysis, followed by physicochemical, microbiological and structural analyses. The temperature and the concentration of the osmotic solution had significant influence (p < 0.05) on ML and SG, whereas DEI was significantly influenced (p < 0.05) by the concentration of osmotic solution and the immersion time. The sample produced by osmotic dehydration using the optimized conditions (40 °C, 50 °Brix; 165 min) obtained higher scores on the sensorial attributes, greater compliance with microbiological standards and generated turgor reduction and ruptures of sapodilla cell walls.

Conditional stomatal closure in a fern shares molecular features with flowering plant active stomatal responses

Stomata evolved as plants transitioned from water to land, enabling carbon dioxide uptake and water loss to be controlled. In flowering plants, the most recently divergent land plant lineage, stomatal pores actively close in response to drought. In this response, the phytohormone abscisic acid (ABA) triggers signaling cascades that lead to ion and water loss in the guard cells of the stomatal complex, causing a reduction in turgor and pore closure. Whether this stimulus-response coupling pathway acts in other major land plant lineages is unclear, with some investigations reporting that stomatal closure involves ABA but others concluding that closure is passive. Here, we show that in the model fern Ceratopteris richardii active stomatal closure is conditional on sensitization by pre-exposure to either low humidity or exogenous ABA and is promoted by ABA. RNA-seq analysis and de novo transcriptome assembly reconstructed the protein-coding complement of the C.richardii genome, with coverage comparable to other plant models, enabling transcriptional signatures of stomatal sensitization and closure to be inferred. In both cases, changes in abundance of homologs of ABA, Ca2+, and ROS-related signaling components were observed, suggesting that the closure-response pathway is conserved in ferns and flowering plants. These signatures further suggested that sensitization is achieved by lowering the threshold required for a subsequent closure-inducing signal to trigger a response. We conclude that the canonical signaling network for active stomatal closure functioned in at least a rudimentary form in the stomata of the last common ancestor of ferns and flowering plants.

Direct evidence for dynamics of cell heterogeneity in watercored apples: turgor-associated metabolic modifications and within-fruit water potential gradient unveiled by single-cell analyses

Watercore is a physiological disorder in apple (Malus × domestica Borkh.) fruits that appears as water-soaked tissues adjacent to the vascular core, although there is little information on what exactly occurs at cell level in the watercored apples, particularly from the viewpoint of cell water relations. By combining picolitre pressure-probe electrospray-ionization mass spectrometry (picoPPESI-MS) with freezing point osmometry and vapor pressure osmometry, changes in cell water status and metabolisms were spatially assayed in the same fruit. In the watercored fruit, total soluble solid was lower in the watercore region than the normal outer parenchyma region, but there was no spatial difference in the osmotic potentials determined with freezing point osmometry. Importantly, a disagreement between the osmotic potentials determined with two methods has been observed in the watercore region, indicating the presence of significant volatile compounds in the cellular fluids collected. In the watercored fruit, cell turgor varied across flesh, and a steeper water potential gradient has been established from the normal outer parenchyma region to the watercore region, retaining the potential to transport water to the watercore region. Site-specific analysis using picoPPESI-MS revealed that together with a reduction in turgor, remarkable metabolic modifications through fermentation have occurred at the border, inducing greater production of watercore-related volatile compounds, such as alcohols and esters, compared with other regions. Because alcohols including ethanol have low reflection coefficients, it is very likely that these molecules would have rapidly penetrated membranes to accumulate in apoplast to fill. In addition to the water potential gradient detected here, this would physically contribute to the appearance with high tissue transparency and changes in colour differences. Therefore, it is concluded that these spatial changes in cell water relations are closely associated with watercore symptoms as well as with metabolic alterations.

Transcriptome Responses of Ripe Cherry Tomato Fruit Exposed to Chilling and Rewarming Identify Reversible and Irreversible Gene Expression Changes.

Tomato fruit stored below 12°C lose quality and can develop chilling injury upon subsequent transfer to a shelf temperature of 20°C. The more severe symptoms of altered fruit softening, uneven ripening and susceptibility to rots can cause postharvest losses. We compared the effects of exposure to mild (10°C) and severe chilling (4°C) on the fruit quality and transcriptome of ‘Angelle’, a cherry-type tomato, harvested at the red ripe stage. Storage at 4°C (but not at 10°C) for 27 days plus an additional 6 days at 20°C caused accelerated softening and the development of mealiness, both of which are commonly related to cell wall metabolism. Transcriptome analysis using RNA-Seq identified a range of transcripts encoding enzymes putatively involved in cell wall disassembly whose expression was strongly down-regulated at both 10 and 4°C, suggesting that accelerated softening at 4°C was due to factors unrelated to cell wall disassembly, such as reductions in turgor. In fruit exposed to severe chilling, the reduced transcript abundances of genes related to cell wall modification were predominantly irreversible and only partially restored upon rewarming of the fruit. Within 1 day of exposure to 4°C, large increases occurred in the expression of alternative oxidase, superoxide dismutase and several glutathione S-transferases, enzymes that protect cell contents from oxidative damage. Numerous heat shock proteins and chaperonins also showed large increases in expression, with genes showing peak transcript accumulation after different times of chilling exposure. These changes in transcript abundance were not induced at 10°C, and were reversible upon transfer of the fruit from 4 to 20°C. The data show that genes involved in cell wall modification and cellular protection have differential sensitivity to chilling temperatures, and exhibit different capacities for recovery upon rewarming of the fruit.

Perilipin LDP1 coordinates lipid droplets formation and utilization for appressorium-mediated infection in Magnaporthe oryzae.

Lipid droplets (LDs) serve as one of the major reservoirs in conidia of Magnaporthe oryzae and are quickly utilized during appressorium formation. Here, we identified a gene, LDP1, encoding a perilipin that is important for LD formation and utilization during appressorium maturation. LDP1 is highly expressed in conidium and immature appressorium. Disruption mutants of LDP1 were significantly reduced in virulence, due to appressorial turgor reduction and difficulty in penetration. LDs were significantly reduced in the Δldp1 mutant, indicating LDP1 was required for LDs formation. LDP1 was colocalized with the LDs in conidium and immature appressorium but was gradually separated during appressorium maturation. A typical intracellular triacylglycerol lipase, TGL1-2, was clearly separated with LDs in conidium and immature appressorium but was well colocalized with LDs during appressorium maturation. The subcellular localization of TGL1-2 was affected by LDP1. These data suggested that LDP1 was bound to LDs for protecting from utilization in conidia and at the early appressorium stage but was separated from LDs for lipase entering and degradation. LDP1 was phosphorylated by CPKA at Thr96, which was essential for its localization and functions. These data indicate perilipin LDP1 can coordinate LD formation and utilization for appressorium-mediated infection of M. oryzae.

Heat-stress triggers MAPK crosstalk to turn on the hyperosmotic response pathway

Cells make decisions based on a combination of external and internal signals. In yeast, the high osmolarity response (HOG) is a mitogen-activated protein kinase (MAPK) pathway that responds to a variety of stimuli, and it is central to the general stress response. Here we studied the effect of heat-stress (HS) on HOG. Using live-cell reporters and genetics, we show that HS promotes Hog1 phosphorylation and Hog1-dependent gene expression, exclusively via the Sln1 phosphorelay branch, and that the strength of the activation is larger in yeast adapted to high external osmolarity. HS stimulation of HOG is indirect. First, we show that HS causes glycerol loss, necessary for HOG activation. Preventing glycerol efflux by deleting the glyceroporin FPS1 or its regulators RGC1 and ASK10/RGC2, or by increasing external glycerol, greatly reduced HOG activation. Second, we found that HOG stimulation by HS depended on the operation of a second MAPK pathway, the cell-wall integrity (CWI), a well-known mediator of HS, since inactivating Pkc1 or deleting the MAPK SLT2 greatly reduced HOG activation. Our data suggest that the main role of the CWI in this process is to stimulate glycerol loss. We found that in yeast expressing the constitutively open channel mutant (Fps1-Δ11), HOG activity was independent of Slt2. In summary, we suggest that HS causes a reduction in turgor due to the loss of glycerol and the accompanying water, and that this is what actually stimulates HOG. Thus, taken together, our findings highlight a central role for Fps1, and the metabolism of glycerol, in the communication between the yeast MAPK pathways, essential for survival and reproduction in changing environments.

Development of screening method for drought tolerance in cotton genotypes based on ABA, chlorophyll stability index and drought tolerant index

Drought stress adversely affects the growth, development and ultimately yield of cotton. The growth and productivity of cotton plants depend largely on their vulnerability to environmental stress. Water stress is commonly attributed to situations where the water loss exceeds sufficient absorption intensity causing a decrease in plant water content, turgor reduction and, consequenctly, a decrease in cellular expansion and alterations of various essential physiological and biochemical processes that can affect growth or productivity. The experiment was conducted by adopting Factorial Randomized Block Design with three replications. The treatments comprised of water stress imposed at vegetative, squaring and boll development stages of crop growth.Withholding water at any growth stage significantly increased the leaf ABA content. Among the different treatments, stress at squaring had a major impact over the ABA quantity enhancement. Chlorophyll stability index is a measure of integrity of membrane or heat stability of the pigments under stress conditions. The genotype KC 2 X MCU 13 showed tolerance to water stress as it accumulated relatively higher ABA.

Constraints on floral water status of successively blossoming Mediterranean plants under natural conditions

The water relations of showy laminar floral tissues (petals and tepals) were studied in 20 Mediterranean plants successively blossoming under ambient conditions. The water potential and osmotic potential of floral tissues decline according to the succession of the median day of flowering of the selected plant species. The highest (least negative) value of floral water potential (–0.32 MPa), among the examined species, was measured in petals of Anemone coronaria in March, and the lowest value (–1.25 MPa) in petals of Coridothymus capitatus in June. Low values of water potential of floral tissues coincided with constraints in declining values of osmotic potential at the onset of the dry period in the Mediterranean region; this apparently resulted in a reduction of turgor of floral tissues of Mediterranean plants subjected to water shortage. The reduced osmotic potential was correlated with enhanced soluble sugar content of floral tissues, presumably contributing to the expansion and water status of flowers under water scarcity, by decreasing water requirements.

Osmotic stress adaptations in rhizobacteria

Rhizobacteria have been reported to be beneficial to the plants in many different ways. Increasing salinity in the coastal agricultural zones has been shown to be a threat to the plant and microbial life in the area. Exposure of microorganisms to high-osmolality environments triggers rapid fluxes of cell water along the osmotic gradient out of the cell, thus causing a reduction in turgor and dehydration of the cytoplasm. The microorganisms have developed various adaptations to counteract the outflow of water. The first response to osmotic up shifts and the resulting efflux of cellular water is uptake of K⁺ and cells start to accumulate compatible solutes. Yet another mechanism is by altering the cell envelope composition resulting in changes in proteins, periplasmic glucans, and capsular, exo and lipopolysaccharides. Bacteria also initiate a program of gene expression in response to osmotic stress by high NaCl concentrations, which are manifested as a set of proteins produced in increased amounts in response to the stress. Genomics, transcriptomics and proteomics approaches have revealed the key components in molecular basis of bacteria salt adaptation. Understanding the mechanisms of osmo-adaptation in rhizobacteria would also be relevant from an ecological and an applicative point of view.

Do pH changes in the leaf apoplast contribute to rapid inhibition of leaf elongation rate by water stress? Comparison of stress responses induced by polyethylene glycol and down‐regulation of root hydraulic conductivity

We have dissected the influences of apoplastic pH and cell turgor on short-term responses of leaf growth to plant water status, by using a combination of a double-barrelled pH-selective microelectrodes and a cell pressure probe. These techniques were used, together with continuous measurements of leaf elongation rate (LER), in the (hidden) elongating zone of the leaves of intact maize plants while exposing roots to various treatments. Polyethylene glycol (PEG) reduced water availability to roots, while acid load and anoxia decreased root hydraulic conductivity. During the first 30 min, acid load and anoxia induced moderate reductions in leaf growth and turgor, with no effect on leaf apoplastic pH. PEG stopped leaf growth, while turgor was only partially reduced. Rapid alkalinization of the apoplast, from pH 4.9 ± 0.3 to pH 5.8 ± 0.2 within 30 min, may have participated to this rapid growth reduction. After 60 min, leaf growth inhibition correlated well with turgor reduction across all treatments, supporting a growth limitation by hydraulics. We conclude that apoplastic alkalinization may transiently impair the control of leaf growth by cell turgor upon abrupt water stress, whereas direct hydraulic control of growth predominates under moderate conditions and after a 30-60 min delay following imposition of water stress.

Turgor Closely Relates to Postharvest Fruit Softening and Can Be a Useful Index to Select a Parent for Producing Cultivars with Good Storage Potential in Apple

Changes in turgor and flesh firmness during storage at 20 °C were investigated using 27 apple (Malus ×domestica Borkh.) cultivars for 2 years. Flesh firmness was measured using a penetrometer, and turgor was determined using a thermocouple psychrometer. Firmness and turgor of fruit decreased during storage. The cultivars with little softening during storage had low rates of reduction in turgor. The softening rates in mealy cultivars were high, but there were cultivars with low rates of turgor reduction. When the rates of reduction in turgor after harvest were low, the mealy cultivars of the fruit tended to develop severe mealiness during storage. Therefore, a low rate of reduction in turgor could contribute to cultivars with both good shelf life and severe mealiness. The reduction rates of turgor in progeny cultivars were nearly identical to the mean reduction rates of turgor of their parents. This suggests that a cultivar with a low reduction rate of turgor, although it can be mealy, has the potential to produce a progeny with a low reduction rate of turgor.

Reduction of Turgor Is Not the Stimulus for the Sensor Kinase KdpD ofEscherichia coli

Stimulus perception by the KdpD/KdpE two-component system of Escherichia coli is still controversial with respect to the nature of the stimulus that is perceived by the sensor kinase KdpD. Limiting potassium concentrations in the medium or high osmolality leads to KdpD/KdpE signal transduction, resulting in kdpFABC expression. It has been hypothesized that changes in turgor are sensed by KdpD through alterations in the physical state of the cytoplasmic membrane. However, in this study the quantitative determination of expression levels of the kdpFABC operon revealed that the system responds very effectively to K(+)-limiting conditions in the medium but barely and to various degrees to salt and sugar stress. Since the current view of stimulus perception calls for mainly intracellular parameters, which might be sensed by KdpD, we set out to test the cytoplasmic concentrations of ATP, K(+), Na(+), glutamate, proline, glycine, trehalose, putrescine, and spermidine under K(+)-limiting conditions. As a first result, the determination of the cytoplasmic volume, which is a prerequisite for such measurements, revealed that a transient shrinkage of the cytoplasmic volume, which is indicative of a reduction in turgor, occurred only under osmotic upshift but not under K(+)-limiting conditions. Furthermore, the intracellular ATP concentration significantly increased under osmotic upshift, whereas only a slight increase occurred after a potassium downshift. Finally, the cytoplasmic K(+) concentration rose severalfold only after an osmotic upshock. For the first time, these data indicate that stimulus perception by KdpD correlates neither with changes in the cytoplasmic volume nor with changes in the intracellular ATP or K(+) concentration or those of the other solutes tested. In conclusion, we propose that a reduction in turgor cannot be the stimulus for KdpD.

Biophysical constraints on leaf expansion in a tall conifer

The physiological mechanisms responsible for reduced extension growth as trees increase in height remain elusive. We evaluated biophysical constraints on leaf expansion in old-growth Douglas-fir (Pseudotsuga menziesii (Mirb.) Franco) trees. Needle elongation rates, plastic and elastic extensibility, bulk leaf water (Psi(L)) and osmotic (Psi(pi)) potential, bulk tissue yield threshold and final needle length were characterized along a height gradient in crowns of > 50-m-tall trees during the period between bud break and full expansion (May to June). Although needle length decreased with increasing height, there was no height-related trend in leaf plastic extensibility, which was highest immediately after bud break (2.9%) and declined rapidly to a stable minimum value (0.3%) over a 3-week period during which leaf expansion was completed. There was a significant positive linear relationship between needle elongation rates and plastic extensibility. Yield thresholds were consistently lower at the upper and middle crown sampling heights. The mean yield threshold across all sampling heights was 0.12 +/- 0.03 MPa on June 8, rising to 0.34 +/- 0.03 MPa on June 15 and 0.45 +/- 0.05 MPa on June 24. Bulk leaf Psi(pi) decreased linearly with increasing height at a rate of 0.004 MPa m(-1) during the period of most rapid needle elongation, but the vertical osmotic gradient was not sufficient to fully compensate for the 0.015 MPa m(-1) vertical gradient in Psi(L), implying that bulk leaf turgor declined at a rate of about 0.011 MPa m(-1) increase in height. Although height-dependent reductions in turgor appeared to constrain leaf expansion, it is possible that the impact of reduced turgor was mitigated by delayed phenological development with increasing height, which resulted in an increase with height in the temperature during leaf expansion.

Growth responses of seminal roots of wheat seedlings to a reduction in the water potential of vermiculite

We examined the elongation rate, water status and solute accumulation in the seminal roots of wheat seedlings (Triticum aestivum L.) that were growing in vermiculite with a water potential (Ψw) ranging from −0 03 to −1 10 MPa. The elongation rate of the primary seminal root was similar to that of the first pair of seminal roots but that of the second pair of seminal roots was lower at all values of Ψw tested. The elongation rate was highest in vermiculite with a Ψw of −0.03 MPa but did not decrease significantly until the Ψw was reduced to −0.15 MPa. Further reductions in Ψw reduced the elongation rate markedly. The Ψw of mature tissues was always similar to that of vermiculite. The osmotic potential (Ψo) decreased to the same extent as the decrease in Ψw. Thus, the turgor pressure (Ψp) remained unchanged even in vermiculite with a low Ψw. In elongating tissues, Ψw and Ψo were far lower than they were in mature tissues and, thus, reductions in turgor were not significant. Even when the Ψw of vermiculite changed, there were no consistent changes in terms of a difference in Ψw between elongating plus mature tissues and vermiculite. There were also no consistent changes in levels of osmotica, calculated using the van’t Hoff’s law, in the elongating tissues but the levels in mature tissues increased in vermiculite with a low Ψw. Our results suggest that (1) reductions in root elongation in vermiculite with a low Ψw were caused by reductions in the extensibility and/or increases in the yield threshold of cell walls and by reductions in the hydraulic conductivity of the tissues; and (2) a seminal root regulates its growth to keep turgor pressure unchanged.

The role of mannitol in affecting the thermal transitions in carrot tissue

Dynamic mechanical thermal analysis (DMTA) and differential scanning calorimetry (DSC) were used to observe thermal events in osmotically manipulated carrot. A glass transition of the concentrated amorphous solution (CAS), recrystallisation of water and ice melting were shown in the tissue by DMTA. DSC showed the role of mannitol as a plasticiser in freeze-dried osmotically manipulated carrot tissue, lowering the cell wall rich and sugar rich phase glass transitions. This cautions against the use of mannitol as an osmoticum for texture measurement since a decrease in stiffness could be due to shift towards a more rubbery state or a reduction of turgor. Processing treatments of air-drying–rehydrating, freezing–thawing or heating–cooling without prior osmotic treatment, induced consistently lower stiffness of carrot tissue compared to that of fresh specimens. The order of stiffness decrease was: fresh>drying–rehydrating∼heating–cooling>freezing–thawing. When osmotically manipulated carrot tissue was processed, stiffness decreased in comparison with osmotically treated specimens alone for molarities less than isotonic. Stiffness could not be restored further to any process events after osmotic changes, demonstrating the irreversible effect of processing on stiffness and contrary to some earlier reports that a prior decrease in turgor would enable the mechanical properties to be recovered.

Yeast osmosensor Sln1 and plant cytokinin receptor Cre1 respond to changes in turgor pressure.

Very little is known about how cellular osmosensors monitor changes in osmolarity of the environment. Here, we report that in yeast, Sln1 osmosensor histidine kinase monitors changes in turgor pressures. Reductions in turgor caused by either hyperosmotic stress, nystatin, or removal of cell wall activate MAPK Hog1 specifically through the SLN1 branch, but not through the SHO1 branch of the high osmolarity glycerol pathway. The integrity of the periplasmic region of Sln1 was essential for its sensor function. We found that activity of the plant histidine kinase cytokinin response 1 (Cre1) is also regulated by changes in turgor pressure, in a manner identical to that of Sln1, in the presence of cytokinin. We propose that Sln1 and Cre1 are turgor sensors, and that similar turgor-sensing mechanisms might regulate hyperosmotic stress responses both in yeast and plants.

Biophysical limitation of leaf cell elongation in source-reduced barley.

The biophysical basis of reduced leaf elongation rate in source-reduced barley ( Hordeum vulgare L. cv Golf) was studied. Reduction in source strength was achieved by removing the blade of leaves 1 and 2 at the time leaf 3 had emerged 3.0-6.7 cm from the encircling sheath. Third leaves of source-reduced plants elongated at 10-36% lower velocities than those of control plants. Removal of source leaves had no significant effect on maximum relative elemental growth rates (REGRs) and the length of the elongation zone (42-46 mm) but caused a shift of high REGR towards the basal portion of the elongation zone. Cell turgor was similar between treatments in the zone of maximal REGR (16-24 mm from base), but was significantly lower in source-reduced plants in the distal part of the elongation zone, where REGR was also lower. Throughout the elongation zone, osmolality and growth-associated water potential gradients were significantly smaller in source-reduced plants; bulk concentrations of sugars (hexoses, sucrose) were also lower. However, even in control plants, sugars contributed little to bulk osmotic pressure (6-11%). The most likely biophysical limitation to leaf (cell) elongation in source-reduced barley was a reduction in turgor in the distal half of the elongation zone. It is proposed that in the proximal half, increase in average tissue hydraulic conductance enabled source-reduced plants to maintain turgor and REGR at control level, while spending less energy on solute transport.

THE RELATION OF MIDDAY STEM WATER POTENTIAL TO THE GROWTH AND PHYSIOLOGY OF FRUIT TREES UNDER WATER LIMITED CONDITIONS

Reductions in rates of both vegetative and fruit growth, and reductions in other physiological parameters such as leaf conductance and photosynthesis, are all well known responses of plants to water limited conditions. One simple hypothesis to explain the overall integration of these responses at the whole plant level is that a reduction in the availability of soil water to the roots will ultimately lead to a reduction in the water potential of the plant as a whole, and hence that a corresponding reduction in the turgor pressure of cells will occur. A reduction in cell turgor may lead, directly or indirectly, to many of the well known plant physiological responses to water limited conditions. A key observation that does not support this simple hypothesis is that under some water limited conditions, reductions in growth and stomatal conductance can occur in the absence of any detectable reduction in the water potential of fully exposed, transpiring leaves. This observation however, does not take into account the possibility that individual leaf stomatal responses may mask reductions in whole plant water potential. By using midday measurements of water potential made on non-transpiring leaves (midday stem water potential) we have found a clear relation between water potential and a number of physiological responses, such as fruit growth and leaf photosynthesis, across a range of fruit tree species growing under field conditions. In almond, overall tree size after two years of growth was linearly related to the average midday stem water potential measured during the same period. A similar linear relation was observed between fruit size and the seasonal average midday stem water potential measured in pear trees. In prune trees, despite an overall linear relation between water potential and light-saturated photosynthesis at the individual leaf level, evidence was found that whole canopy photosynthesis may be relatively unaffected by moderate levels of water stress (midday stem water potential around -1.5 MPa). We have found that moderate water stress is associated with improvements in fruit quality and economic return in prunes, and are currently using midday stem water potential to guide the management of RDI (regulated deficit irrigation) in this crop.

Abscisic acid-induced stomatal closure mediated by cyclic ADP-ribose.

Abscisic acid (ABA) is a plant hormone involved in the response of plants to reduced water availability. Reduction of guard cell turgor by ABA diminishes the aperture of the stomatal pore and thereby contributes to the ability of the plant to conserve water during periods of drought. Previous work has demonstrated that cytosolic Ca2+ is involved in the signal transduction pathway that mediates the reduction in guard cell turgor elicited by ABA. Here we report that ABA uses a Ca2+-mobilization pathway that involves cyclic adenosine 5'-diphosphoribose (cADPR). Microinjection of cADPR into guard cells caused reductions in turgor that were preceded by increases in the concentration of free Ca2+ in the cytosol. Patch clamp measurements of isolated guard cell vacuoles revealed the presence of a cADPR-elicited Ca2+-selective current that was inhibited at cytosolic Ca2+ >/= 600 nM. Furthermore, microinjection of the cADPR antagonist 8-NH2-cADPR caused a reduction in the rate of turgor loss in response to ABA in 54% of cells tested, and nicotinamide, an antagonist of cADPR production, elicited a dose-dependent block of ABA-induced stomatal closure. Our data provide definitive evidence for a physiological role for cADPR and illustrate one mechanism of stimulus-specific Ca2+ mobilization in higher plants. Taken together with other recent data [Wu, Y., Kuzma, J., Marechal, E., Graeff, R., Lee, H. C., Foster, R. & Chua, N.-H. (1997) Science 278, 2126-2130], these results establish cADPR as a key player in ABA signal transduction pathways in plants.

Expression of the Kdp ATPase Is Consistent with Regulation by Turgor Pressure

The kdpFABC operon of Escherichia coli encodes the four protein subunits of the Kdp K+ transport system. Kdp is expressed when growth is limited by the availability of K+. Expression of Kdp is dependent on the products of the adjacent kdpDE operon, which encodes a pair of two-component regulators. Studies with kdp-lac fusions led to the suggestion that change in turgor pressure acts as the signal to express Kdp (L. A. Laimins, D. B. Rhoads, and W. Epstein, Proc. Natl. Acad. Sci. USA 78:464-468, 1981). More recently, effects of compatible solutes, among others, have been interpreted as inconsistent with the turgor model (H. Asha and J. Gowrishankar, J. Bacteriol. 175:4528-4537, 1993). We re-examined the effects of compatible solutes and of medium pH on expression of Kdp in studies in which growth rate was also measured. In all cases, Kdp expression correlated with the K+ concentration when growth began to slow. Making the reasonable but currently untestable assumptions that the reduction in growth rate by K+ limitation is due to a reduction in turgor and that addition of betaine does not increase turgor, we concluded that all of the data on Kdp expression are consistent with control by turgor pressure.