pH, abscisic acid and the integration of metabolism in plants under stressed and non-stressed conditions: cellular responses to stress and their implication for plant water relations

A paradigm for the response of plants to stress is presented which suggests that plants move towards a state of minimal metabolic activity as a stress intensifies and remain in that state until that stress is relieved. The paradigm is based on the proposition that cells that interface with the transpiration stream employ variations on the following theme to move towards that state. Tension on the apoplastic water opens a mechanosensitive Ca2+ channel, a response that is augmented by apoplastic ABA. The resulting elevated cytoplasmic Ca2+ deactivates a plasmalemma H+/ATPase and also activates a K(+)-H+ symport. The inflow of K+ and H+ depolarizes the membrane and renders the apoplast less acidic, the protons being removed to the vacuole and the K+ ions being re-exported via the K+ outward rectifying channel. The onset of darkness in guard and mesophyll cells deactivates the plasmalemma H+/ATPase and then the events outlined above ensue except that these cells do not appear to utilize either Ca2+ or ABA during these changes. In stressed cells it is proposed that elevated cytoplasmic Ca2+ activates the release of an ABA precursor from a stored form. ABA is then released in the apoplast after export of the precursor if the activity of the K(+)-H+ symport has brought the apoplastic pH close to 7.0. It is proposed that aquaporins in the xylem parenchyma and mesophyll cells are opened by elevated cytoplasmic Ca2+ when the water potential of the transpiration stream is high so that water can be stored in the 'xylem parenchyma reservoir'. The water in this reservoir is then used to increase the water potential in the transpiration stream when the water column is under tension and to help repair embolisms by a mechanism that resembles stomatal closure.     

Modifications of drug metabolism by disulfiram and diethyldithiocarbamate. II. D-Glucuronic acid pathway.

Hepatic enzymes connected with the formation and metabolism of free D-glucuronic acid were affected in rats after treatment with disulfiram or diethyldithiocarbamate (300 mg/kg, intragastrically, per day, 4 X). The activities of UDPglucose dehydrogenase, UDPglucuronic acid pyrophosphatase, UDPglucuronosyltransferase and L-gulonate dehydrogenase were enhanced, while those of glucose-6-phosphate dehydrogenase, beta-glucuronidase and D-glucuronolactone dehydrogenase were inhibited. These changes were more pronounced with disulfiram than diethyldithiocarbamate. Treatment with phenobarbital (80 mg/kg, i.p., per day, 4 X) enhanced UDP glucuronosyl-transferase, but brought about different effects on the other enzymes. Concurrent administration of phenobarbital with disulfiram or diethyldithiocarbamate led to potentiation or antagonism of the primary effects of each compound when given alone. The results suggest that activation of the D-glucuronic acid pathway may proceed in various ways, and that it is not necessarily followed by a simultaneous induction of the microsomal mixed-function oxygenase activity.     

Effect of ethylene perception inhibitor on growth, water relations, and abscisic acid content in wheat plants under water deficit

The effects of treatment with 1-methylcyclopropene (1-MCP, the inhibitor of ethylene receptors) of 7-day-old wheat (Triticum durum Desf., cv. Bezenchukskaya 139) seedlings on growth characteristics, water relations, and the content of phytohormones during three days after watering cessation were studied. In treated seedlings, a decrease in the water content in the substrate resulted in a decrease in stomatal conductance in the leaves by one day earlier than in untreated seedlings. This could be related to the more rapid and substantial accumulation of ABA in treated plants. There was no clear relationship between changes in the content of cytokinins and water relations under the influence of 1-MCP under drought conditions. The role of ethylene and ABA in the regulation of growth and water relations in plants suffering from water deficit is discussed.     

Stem pithiness in tomato plants: The effect of water stress and the role of abscisic acid

The pith parenchyma in tomato ( Lycopersicum esculentum ) stems was found to be disrupted in response to water stress (stem pithiness). The process of the degradation starts at the upper part of the stem and proceeds downwards as the stress is prolonged. The damage to the stem tissue was found to be irreversible upon rehydration of the stressed plants. Scanning electron microscopy revealed that the protoplast of the affected cells is disrupted first, followed by degradation of the cell wall.
Application of abscisic acid (ABA) induced pithiness in non-stressed plants and also enhanced the effect of a short period of dehydration. Kinetin, although causing severe wilting, did not induce pithiness. However, when given after a short period of water stress or within the period of stress, kinetin enhanced pithiness development.
In the course of the stress, ABA levels in the upper part of the stem and in the young leaves were higher than the levels found in the lower parts of the plant shoot. The increase in ABA levels was detected before any sign of pithiness.
It is suggested that ABA might be the triggering agent of the cellular degradation process initiated by water stress.     

The content of abscisic acid and the activities of proteinases and trypsin inhibitory proteins, in the germinating seed of common beans under water stress conditions

Specific features of changes in the contents of free and bound abscisic acid (ABA) and the activities of neutral and alkaline proteinases and trypsin inhibitory proteins were determined in the embryonic axis and cotyledons of the common bean (Phaseolus vulgaris L.) after drying. The changes in ABA content, observed following the loss of 5% seed weight, were regarded as an adaptive reaction to stress, whereas the corresponding changes after the loss of 10% of the seed’s weight, was regarded as a result of a pathological disturbance of ABA metabolism. Both drying modes had a negative effect on the state of the proteinase-inhibitory system, as was apparent from the disruption of the regular inverse correlation between the activities of proteinases and serine proteinase inhibitory proteins. A comparison of the dynamics of these characteristics with the buildup of water stress demonstrates an inverse correlation between the content of free ABA and the activity of the proteinases studied. This suggests a potential inhibitory effect of this hormone on the function of the hydrolases in question in the germinating seed.     

Effect of water stress on metabolism of abscisic acid in Populus robusta× schnied and Euphorbia lathyrus L.

Abstract. Dihydrophaseic acid (DPA) has been identified in leaves from Euphorbia lathyrus L and Populus robusta x schnied. The formation of DPA from abscisic acid (ABA) was demonstrated using ¹⁴C-ABA. Measurements of ABA, DPA and phaseic acid (PA) concentrations were made in Euphorbia subjected to drought and waterlogging and in Populus subjected to rhythmic water stress. The results are consistent with the proposition that ABA concentrations are controlled by de novo biosynthesis and by metabolism via the PA /DPA pathway. The findings are discussed in relation to effects of the type of stress and its duration and to behaviour during stress relief.     

Jasmonoyl isoleucine accumulation is needed for abscisic acid build‐up in roots of Arabidopsis under water stress conditions

Phytohormones are central players in sensing and signalling numerous environmental conditions like drought. In this work, hormone profiling together with gene expression of key enzymes involved in abscisic acid (ABA) and jasmonate biosynthesis were studied in desiccating Arabidopsis roots. Jasmonic acid (JA) content transiently increased after stress imposition whereas progressive and concomitant ABA and Jasmonoyl Isoleucine (JA‐Ile) accumulations were detected. Molecular data suggest that, at least, part of the hormonal regulation takes place at the biosynthetic level. These observations also point to a possible involvement of jasmonates on ABA biosynthesis under stress. To test this hypothesis, mutants impaired in jasmonate biosynthesis (opr3, lox6 and jar1‐1) and in JA‐dependent signalling (coi1) were employed. Results showed that the early JA accumulation leading to JA‐Ile build up was necessary for an ABA increase in roots under two different water stress conditions. Signal transduction between water stress‐induced JA‐Ile accumulation and COI1 is necessary for a full induction of the ABA biosynthesis pathway and subsequent hormone accumulation in roots of Arabidopsis plants. The present work adds a level of interaction between jasmonates and ABA at the biosynthetic level.     

Effects of abscisic acid on the contents of polyamines and proline in common bean plants under salt stress

The effects of ABA treatment on the contents of polyamines (PAs) and proline (Pro) in the glycophyte Phaseolus vulgaris L. during plant adaptation to salt stress were studied. Two-week-old common bean seedlings grown in the phytotron chamber on the Jonson nutrient medium were subjected to salinity for 6 days by one-time NaCl addition to medium up to final concentrations of 50 and 100 mM. During first three days of salinity, the root system was daily treated with ABA (1, 5, 10, or 50 μM) for 30 min. Salt stress (100 mM NaCl) elevated the level of endogenous ABA, increased the content of Pro 14-fold, reduced sharply the content of free PAs (putrescine, spermidine, spermine, and cadaverine), and the accumulation of 1,3-diaminopropan, a product of oxidation of high-molecular PAs. Common bean plant treatment with 1 μM ABA weakened the adverse effects of salt stress (100 mM NaCl), which was manifested in the maintenance of plant growth, stimulation of chlorophyll (a and b) and carotenoid accumulation, a stabilization of water and Na⁺ balance. Seedling treatment with ABA suppressed NaCl-induced Pro and intracellular ABA accumulation and restored the levels of putrescine and spermidine. The content of spermine in the leaves of plants subjected to salt stress and treated with ABA was approximately threefold higher than in control plants, whereas the content of cadaverine increased under similar conditions more than fivefold. Simultaneously, the contents of 1,3-diaminopropan and malondialdehyde as well as activity of superoxide dismutase were reduced, which indicates a weakening of oxidative stress, one of the possible causes of defensive ABA effects against salt stress. In addition, the suppression by exogenous ABA of Pro accumulation and stimulation of PA content under salt stress confirm indirectly our hypothesis that ABA is involved in the coordinated regulation of two biosynthetic pathways, Pro and PA formation, which use a common precursor, glutamate, and play an important protective role during stress in plants.     

Energy metabolism in largemouth bass (Micropterus floridanus salmoides) from stressed and non-stressed environments: adaptations in the secondary stress response

White muscle proteins, carbohydrates, lipids, adenylates, phosphagen and the AEC, (adenylate energy charge) were measured in bass inhabiting stressful and non-stressful environments. Within an environment, in June stressed bass had higher carbohydrates, while non-stressed bass had lower ATP, total adenylates and AEC. Comparisons between environments revealed: non-stressed bass had higher ATP/ADP and AEC's in May, CrP/ATP in June, and protein in July; while stressed bass had higher carbohydrate in June and lipid in July. Other metabolites varied insignificantly. AEC's of both groups were within the optimal range indicating physiological compensation (adaptation) of energy metabolism (secondary stress response) occurred in bass inhabiting the stressful environment.     

Diurnal variations in abscisic acid content and stomatal response to applied abscisic acid in leaves of irrigated and non-irrigated Arbutus unedo plants under naturally fluctuating environmental conditions

Summary Endogenous abscisic acid content (ABA) of Arbutus unedo leaves growing under natural conditions in a macchia near Sobreda, Portugal, was very high (0.25 to 2.3 g g¹ fresh weight). Highest concentrations were found during the very early morning hours and at midday. During the late morning hours and in the late afternoon ABA concentrations decreased to between one-third and one-fourth of peak values. The samples for ABA content were obtained from both irrigated ( between-10 and-25 bar) and non-irrigated plants experiencing natural water stress during the dry season ( of-50 bar). During the course of the measurement day, stomatal conductance was relatively constant and conductance of watered plants was 50 to 100% greater than that of unwatered plants. No clear correlations between ABA content and stomatal conductance and/or xylem water potential were observed. Despite large differences in water potential and differences in degree of stomatal opening, absolute concentrations of ABA were not found to differ.Small quantities (8–14 pmoles cm² leaf area) of ABA were applied to leaves of irrigated and non-irrigated Arbutus unedo plants by injection into the petiole. These extremely small ABA doses resulted in transient reductions in stomatal conductance. The effectiveness with which injected ABA closed stomata was highest during the morning and decreased substantially at midday. Increased sensitivity to injected ABA may again occur in the late afternoon but recent measurements suggest that this may depend on long-term drought experience of the plants. The characteristics of the response to injected ABA were similar in irrigated and non-irrigated plants although irrigated plants responded in general more strongly.     

Accumulation of sugars in the xylem apoplast observed under water stress conditions is controlled by xylem pH

Severe water stress constrains, or even stops, water transport in the xylem due to embolism formation. Previously, the xylem of poplar trees was shown to respond to embolism formation by accumulating carbohydrates in the xylem apoplast and dropping xylem sap pH. We hypothesize that these two processes may be functionally linked as lower pH activates acidic invertases degrading sucrose and inducing accumulation of monosaccharides in xylem apoplast. Using a novel in vivo method to measure xylem apoplast pH, we show that pH drops from ~6.2 to ~5.6 in stems of severely stressed plants and rises following recovery of stem water status. We also show that in a lower pH environment, sugars are continuously accumulating in the xylem apoplast. Apoplastic carbohydrate accumulation was reduced significantly in the presence of a proton pump blocker (orthovanadate). These observations suggest that a balance in sugar concentrations exists between the xylem apoplast and symplast that can be controlled by xylem pH and sugar concentration. We conclude that lower pH is related to loss of xylem transport function, eventually resulting in accumulation of sugars that primes stems for recovery from embolism when water stress is relieved.     

A rapid method for analysis of abscisic acid (ABA) in crude extracts of water stressed Arabidopsis thaliana plants by liquid chromatography--mass spectrometry in tandem mode.

We describe a quick, simple method for the extraction and quantification of the phytohormone (+)-abscisic acid (ABA) in samples of plants subjected to different water deficit treatments. The method includes an extraction with acetone/water/acetic acid (80:19:1, v/v), evaporation of the extracts and finally injection into the liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS-MS) system in multiple reaction monitoring (MRM) mode. The objective of this work has been to show the applicability of the method to quantify the endogenous content of ABA in Arabidopsis thaliana (Columbia, Col-0) leaves at three different degrees of water stress. Control plants, had almost constant low levels of ABA (2-3 ng g-1, f.w.) throughout the 3 weeks of the experiment. Nevertheless, stressed plants increase the ABA content between the first and the second week (from 10 to 21 ng g-1, f.w.). The results suggest that this method is useful for quantifying ABA from plant material and that it avoids tedious and time-consuming extraction, purification and/or derivatization processes.     

Abscisic acid in lichens: variation, water relations and metabolism

For the first time the well known drought stress hormone abscisic acid, which is involved in regulating processes increasing desiccation tolerance in many plant systems, was analysed in lichens. ABA was detected in all 26 species investigated. In contrast to higher plants and liverworts, the ABA content increased after hydration of air dry lichen thalli and decreased in desiccating lichen material. Experiments with Baeomyces rufus (Huds.) Rebent indicated that the mycobiont might be the major site of ABA biosynthesis. After incubation of hydrated lichen thalli with radioactive ABA for up to 72 h no metabolism to phaseic acid and dihydrophaseic acid could be detected. Fluctuations of internal ABA might be a result of ABA release to the external medium.     

Quantification of free and conjugated abscisic acid in five genotypes of barley (Hordeum vulgare L.) under water stress conditions

The goal of the present study was to obtain new insights into the mechanisms underlying drought stress adaptations in barley plants. For this purpose we evaluated changes in endogenous abscisic acid (ABA) and its glucose ester conjugate (ABAGE), as well as changes in proline content, water relations and growth parameters of five barley genotypes with different drought resistance characteristics. Different responses among the five genotypes studied (Ardahoui, Pakistan, Rihane, Manel ad Roho) led to changes in their pattern of growth and development under drought conditions. Water stress induced a reduction in relative water content, as well as an increase in proline content and endogenous ABA concentrations in all tested genotypes. The lack of water led to a 2-fold increase in proline content for var. Rihane and to a 5-fold increase in endogenous ABA for cv. Ardhaoui. Also, increases in endogenous ABAGE in all genotypes except for cv. Ardhaoui were observed. Our results show that changes in ABA and ABAGE correlated with variations in proline content and growth parameters of these genotypes which present different mechanisms to cope with water stress. We also suggest that new regulatory mechanisms implying ABA mobilization can be of great importance in adaptation of barley to drought.     

Study of the ionome and uptake fluxes in cherry tomato plants under moderate water stress conditions

Nutritional imbalance under water-deficit conditions depresses plant growth by affecting nutrient uptake, transport, and distribution. The present work analyses the variations in the foliar concentrations of macro- and micronutrients as well as the transport of these nutrients in five cherry tomato cultivars under well-watered and moderately water-stressed conditions with the aim of establishing whether the ionome of the plants is related to the degree of sensitivity or tolerance to this type of stress. The results show a general reduction in growth together with a lower concentrations and uptake both of macro- as well as micronutrients in all the cultivars studied, except for cv. Zarina, which showed better growth and increased in concentrations and uptake nitrogen, phosphorus, magnesium, potassium, and chloride with respect to control plants. In conclusion, in this work, our results suggest that a better understanding of the role of the mineral elements in plant resistance to drought could improve fertilization in arid and semi-arid regions in order to increase the tolerance of plants grown under these conditions.