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Plant Response to Stress II Water Stress, Drought stress Development of water stress Adaptation to drought Signals and signalling pathways Basic processes affected by water stress Water stress • Water stress is induced when transpiration rate is higher than absorption rate • High transpiration rate • Low absorption rate • Gradient of water potential between substrate and shoot is prerequisite for water transport Importance for water deficiency: Ψw = Ψs + Ψp • - low air humidity, - high temperature, - high irradiance, - strong wind - low soil moisture - high concentration of salts, - low soil temperature • Plants can only absorb water if their water potential is more negative than that of the source (soil, etc.); Transport of water only from higher to lower water potential • When water potential in cells higher: osmotic adjustment. This is the net increase in accumulation of solutes within cells in response to stress; in cytoplasm or vacuole. • Transport of water for storage equilibrates small differences between transpiration rate and absorption rate Drought Stress/Water Stress/Osmotic Stress... Main effects of water stress 1. Reduction in cell and leaf expansion 2. Reduction in photosynthesis, due first to decreased stomatal conductance, then to inhibition of chloroplast metabolism. 3. Altered allocation - greater investment in nonphotosynthetic tissues such as roots & mycorrhizae Drought Stress/Water Stress/Osmotic Stress... Main effect of water stress : growth reduction Leaf expansion is very sensitive to water deficit Second effect: Reduction in photosynthesis, due first to decreased stomatal conductance, then to inhibition of chloroplast metabolism Drought Stress/Water Stress/Osmotic Stress... Effects of water deficiency on photosynthesis and transport of assimilates in millet. Phloem translocation seems to be less sensitive to water stress than photosynthesis. Drought Stress/Water Stress/Osmotic Stress... Physiological consequences of low water stress Responses to deal with stress The edt1 Mutant with Greatly Improved. Drought Tolerance . ©2008 by American Society of Plant Biologists Yu H et al. Plant Cell 2008;20:1134-1151 (A) and (B) Under short-day conditions, 4week-old wild type (A) and mutant (B) plants were similar, except that the mutant had more rosette leaves with shorter leaf petiole. Bars = 1 cm. (C) and (D) Under long-day conditions, the 4-week-old mutant showed more vigorous vegetative growth (D) and had more rosette leaves than the wild type (C). Bars = 1 cm. (E) to (G) Drought tolerance assay. The mutant and wild-type seeds were germinated in soil side by side at high density under the same greenhouse conditions. (E) Two-week-old seedlings immediately before drought stress. (F) Seedlings deprived of water for 7 d. (G) Seedlings deprived of water for 14 d. (H) Drought stress of the mutant and wild-type plant grown in the same pot. One mutant and one wild-type plant were grown in the same pot for 4 weeks under short-day conditions before drought stress was imposed. The photos were taken 2 weeks after watering was withheld. (I) Comparison of water loss rate of detached leaves from the mutant and wild type. Fourweek-old plants were derooted and allowed to dry in the air. Fresh weight was measured at the indicated times. Water loss was expressed as the percentage of the initial fresh weight. Values are mean ± SE (n = 20 plants The edt1 Mutant with Greatly Improved Drought Tolerance 3days 10 days 5 20 days (F) Number of lateral roots of the wild-type and the mutant (edt1) seedlings grown on medium as in (A). The number of lateral roots was counted at the indicated time points. Values are mean ± SE (n = 30 seedlings). (G) Root dry weight of wild-type and mutant (edt1) seedlings grown on medium as shown in (B). Values are mean ± SE (n = 30 seedlings, ** P < 0.01). (H) Root biomass of soil-grown wild-type and edt1 mutant plants as shown in (D). Fresh weight of roots was compared between the mutant and the wild-type plants. Values are mean ± SE (n = 10 seedlings, ** P < 0.01). ©2008 by American Society of Plant Biologists Improved Root Architecture of the edt1 Mutant. (A) The primary root of 3-d-old mutant seedlings (edt1) was longer than that of the wild type of the same age on Murashige and Skoog (MS) medium. Bar = 0.5 cm. (B) The primary root of 10-d-old mutant seedlings (edt1) was longer than that of the wild type of the same age on MS medium. Bar = 1 cm. (C) Twenty-day-old mutant weeks seedlings (edt1) have more lateral roots than wild-type seedlings of the same age. (D) Five-week-old mutant (edt1) and wild-type plants were grown in soil under short-day conditions. The plants were removed from the soil and carefully washed without damaging the taproots. Bar = 2 cm. (E) Primary root elongation of the wild-type and mutant seedlings (edt1) grown on medium as in (A). The length of the primary root was measured at the indicated time points. Values are mean ± SE (n = 30 seedlings). Yu H et al. Plant Cell 2008;20:1134-1151 The edt1 Mutant with Greatly Improved Drought Tolerance Reduced Stomatal Density and Increased Water Use Efficiency of the Mutant. WUE (mg photosynthate produced/g water transpired) Reduced Stomatal Density and Increased Water Use Efficiency of the Mutant. (A) Comparisons of adaxial epidermal imprint images of the wild type and the mutant (edt1) at the same x200 and x400 magnifications. (B) and (C) Comparisons of stomatal and cell density (B) and stomatal dimension (C) in the wild type and the edt1 mutant. Values are mean ± SE (n = 30 plants, * P < 0.05, **P < 0.01). (D) to (F) Comparisons of photosynthesis rate (D), transpiration rate (E), and WUE (F) in the wild type and the edt1 mutant. WUE (mg photosynthate produced/g water transpired) was measured as described in Methods. Three measurements were made for each plant, and five plants were used for each line. Values are mean ± SE (n = 30 plants, * P < 0.05, **P < 0.01). The edt1 Mutant with Greatly Improved Drought Tolerance The edt1 Mutant with Greatly Improved Drought Tolerance Quantification of ABA, Pro, and SOD Activity in the edt1 Mutant, 35S-HDG11, and Wild-Type Seedlings. (A) ABA contents. Ten-day-old seedlings of the wild type, edt1 mutant, and 35S-HDG11 transgenic lines were used for ABA quantification. ABA was determined by ELISA. Values are mean ± SE (n = 3 experiments, * P < 0.05, **P < 0.01). FW, fresh weight. (B) Pro contents. Ten-day-old seedlings of the wild type, edt1 mutant, and 35S-HDG11 transgenic lines were used for Pro quantification. Pro content was measured spectrophotometrically. Values are mean ± SE (n = 3 experiments, * P < 0.05, **P < 0.01). (C) Comparison of oxidative stress tolerance between the wild-type and edt1 mutant seedlings. The wild-type and mutant seeds were first germinated on MS medium and then transferred to MS medium containing 0 (Control) or 0.2 µM paraquat (Paraquat) and incubated under continuous light at 22°C for 1 week before the photographs were taken. (D) Survival rate of the wild type and mutant on medium containing 0 (Control) or 0.2 µM paraquat (Paraquat) over a period of 1 week. Survival rate (y axis) was defined as the percentage of the wild-type control. Values are mean ± SE (n = 50 plants, **P < 0.001). (E) Comparison of SOD activity between the edt1 mutant and the wild type. Ten-day-old mutant and wild-type seedlings were treated with 0 or 10% PEG 6000 in liquid MS medium for 6 h, and then SOD activity was assayed and presented as folds of the wild-type control. Values are mean ± SE (n = 3 exp., * P < 0.05, **P < 0.01). (F) Estimation of SOD mRNA level in the edt1 mutant and the wild type by RT-PCR. Total RNA was isolated from 1- and 2-weekold seedlings (1W and 2W) and leaves (L) and roots (R) of 4-week-old plants grown in soil. SOD transcript levels were estimated by RT-PCR for 30 cycles with specific primers for SOD. Tubulin (Tub) transcript levels serve as an equal loading standard. The experiment was repeated three times, and the typical result of an ethidium bromide–stained agarose gel is presented. (G) Estimation of SOD mRNA level by real-time RT-PCR. Using the same sample and primers as in (F), real-time RT-PCR was performed for 30 cycles. The relative transcript level was obtained as folds of the tubulin transcript level, which was used as the internal control. Values are mean ± SE (n = 3 experiments). The edt1 Mutant with Greatly Improved Drought Tolerance Identification of the T-DNA Tagged Locus and Activated Expression of the Tagged HDG11 Gene. (A) DNA gel blot analysis to estimate the copy number of T-DNA insertion. Total DNA was isolated from the edt1 mutant seedlings and digested with restriction endonuclease HindIII (H3) and EcoRI (R1), respectively. The wild type Columbia was used as control. Lane M is the 1-kb ladder molecular size marker. The DNA gel blot was probed with bar and washed at high stringency. The autoradiograph indicates that the mutant edt1 genome contains only one T-DNA insert. (B) Illustration of the chromosomal location of the T-DNA insertion and locus At1g73360 (not drawn to scale). The single T-DNA insertion site was identified through plasmid rescue. The relative location (inverted solid triangle) and orientation (small arrow) of the T-DNA are shown (top panel). The T-DNA right border with the four copies of 35S enhancers was inserted in the 5' UTR, at 50 bp upstream of the ATG codon of HDG11 (middle panel). The HDG11 gene encodes a predicted protein of 722 amino acids with two known functional domains: a homeodomain for DNA binding and a START domain presumably for ligand binding (bottom panel). (C) RT-PCR analysis of transcript levels of the neighboring genes. Transcript levels for At1g73350, At1g73360 (HDG11), and At1g73370 were compared in both roots (R) and leaves (L) in the wild type and the edt1 mutant. The experiment was repeated three times, and a typical result is presented. (D) Real-time RT-PCR analysis of transcript levels of the neighboring genes. Using the same samples and primers as in (C), real-time RT-PCR was performed for 30 cycles. The relative transcript level was obtained as folds of the tubulin transcript level, which was used as the internal control. Values are mean ± SE (n = 3 experiments). (E) RT-PCR analysis of the expression patterns of HDG11 in the mutant and the wild type. RNA was isolated from roots (R), rosette leaves (RL), cauline leaves (CL), inflorescence stem (ST), flower buds (FB), flowers (FL), young siliques (YSi), and mature siliques (MSi) of the wild type and the edt1 mutant plants, respectively. Tubulin was used as a loading control. The experiment was repeated three times, and a typical result is presented. (F) Real-time RT-PCR analysis of the expression patterns of HDG11 in the mutant and the wild type. Using the same samples and primers as in (E), real-time RT-PCR was performed for 30 cycles as described in Methods. The relative transcript level was obtained as folds of the tubulin transcript level as the internal control. Values are mean ± SE (n = 3 experiments). The edt1 Mutant with Greatly Improved Drought Tolerance Drought Stress/Water Stress/Osmotic Stress... Summary of Drought Stress Symptoms M acroscopic physiological dam age caused by dehydration. In general: can lead to many developmental changes including • Inhibition of elongation growth, cell division, changes in cell wall synthesis • delays in flowering • leaf abscission • stomatal closure, • Acceleration of ageing • Reduced tillering and root development. • curling and rolling of leaves during midday • Reduction of spikelets and florets during the development of the spike • grain shriveling (main stress). • during the development of the spike and during seed set: complete or partial sterility. Drought Stress/Water Stress/Osmotic Stress... Summary of Drought Stress Symptoms Biochem ical dam age caused by dehydration. • Production of stress proteins • Accumulation of osmotically active compounds (proline, glycinebetaine, sugars, sugar alcohols) • ROS production and development of antioxidative systems • Inhibition of photosynthesis, • Changes in enzyme activities (decrease in activity of Rubisco, PEPC, nitratereductase) • higher root carbon allocation, transport of assimilates, respiration • Changes in biosynthesis and catabolism of phytohormones, especially ABA • Changes in absorption and transport of ions • cuticular wax increase: reduces cuticular transpiration, increases reflection Water stress affects almost all processes in plants Drought Stress/Water Stress/Osmotic Stress... Tentative scheme of photosynthetic control under drought Under drought, stomatal closure (2) in proportion to the degree of the stress progressively limitating ΔpH in the chloroplasts. (3) CO2 assimilation is reduced and CO2:O2 ratio drops thereby increasing photorespiration Less ATP and/or Mehler reaction (4). Since these consumption processes consume relatively less ATP than does photosynthesis, they should lead to a certain increase of trans-thylakoid ΔpH (5). Impaired ATPase and/or ETR may also interfere with Photorespiration the build up of of trans-thylakoid ΔpH. (6) The xanthophyll deepoxidation that follows increased ΔpH should lead to increased NPQ (7). Thermal dissipation in the antenna becomes progressively more important and Fs is consecutively lowered (7). The relationship between Fs and stomata conductivity provides a method for remote sensing stress. Adaptations of plants to water stress 1) drought avoidance – whole growth cycle in a wet season, leaf fall under drought: plants complete their life cycles during the “wet season, before the onset of drought; 2) drought tolerance (resistance) ability to function while dehydrated • drought tolerance at low cell water potential – survive with minimum metabolism: seeds, pollen grains, resurrection plants • drought tolerance at higher cell water potential. adaptation reactions against drought stress - regulation of water loss (stomata, cuticle, trichomes, leaf movements, leaf shape, leaf area, C3→CAM), Drought Stress/Water Stress/Osmotic Stress... Adaptation reactions against water stress Leaf structure: water deficit increase wax deposition on the leaf surface a) The production of a thicker cuticle that reduces water loss from the epidermis (cuticle transpiration) b) A thicker cuticle also decreases CO2 permeability, but leaf photosynthesis remains unaffected because the epidermal cells underneath the cuticle are nonphotosynthetic Drought Stress/Water Stress/Osmotic Stress... Adaptation reactions against water stress Rooting characteristics: water deficit enhances root extension into deeper, moist soil a) Inhibition of leaf expansion by water deficit reduces the consumption of carbon and energy, and a greater proportion of the plant’s assimilates can be distributed to the root system, where they can support further root growth b) The enhanced water uptake resulting from root growth is less pronounced in reproductive plants than in vegetative plants Adaptations of plants to water stress 1) drought avoidance – whole growth cycle in a wet season, leaf fall under drought: plants complete their life cycles during the “wet season, before the onset of drought; 2) drought tolerance (resistance) ability to function while dehydrated • drought tolerance at low cell water potential – survive with minimum metabolism: seeds, pollen grains, resurrection plants • drought tolerance at high cell water potential. adaptation reactions against drought stress - regulation of water loss (stomata, cuticle, trichomes, leaf movements, leaf shape, leaf area, C3→CAM), - regulation of absorption (amount and morphology of roots, osmotic adjustment), - efficient water transport with low water potential, - water storages (stems, trunks, fruits), - production of protection compounds (e.g. carotenoids, osmolytes, stress proteins, antioxidants) Osmotic adjustment helps plants to cope with water stress Water uptake from the soil happens when soil A decrease in Ψ S helps to potential is higher than plant water potential maintain turgor, Ψ P, and contributes also to a lower water potential decreases. Osmotic adjustment is a net increase in solute content per cell. Many solutes contribute to osmotic adjustment. K+, sugars, organic acids, amino acids Osmotic adjustment may occur over a period of days. Costs of osmotic adjustment: synthesis of organic solutes, maintenance of solute gradients, and energy, which can not be used for other functions Ψw = Ψs + Ψp Osmotic and elastic adjustment Osmotic adjustment • Will be induced by decrease in soil water potential, air humidity, etc. By 1. ion uptake 2. production and accumulation of osmoprotectants: osmotically active substances 2.1 sugars (glucose, trehalose, saccharose), sugar alcohols (mannitol, sorbitol, glycerol), Accumulation of mono- and di-saccharides by inhibition of starch synthesis from new photosynthates, degradation of starch, inhibiton of respiration 3. polyamines, amino acids (proline), betaines (glycinebetaine) Synthesis in chloroplasts by oxidation of choline in two steps Sugars serve not only as osmotica but also in signalling pathways, or gene expression regulation • Membrane protection, source of C or N, defence against ROS • Dehydrines – ripening of seeds or pollen grains, in plant vegetative parts during stresses, induced also by abscisic acid (ABA) Elastic adjustment • expansin, endoglucanase, transglycosylase, peroxidase • At different plant species are different adaptations, amount of osmoticum is not always in correlation with water stress tolerance Drought Stress/Water Stress/Osmotic Stress... Adaptation against the stress stress Adaptation reactionsreactions against water stress/osmotic Adaptation of cytosolic water potential Accumulation of compatible solutes: solutes must accumulate in the cytoplasm to maintain water potential equilibrium within the cell Function of the stress-induced cytosolic osmolytes for lowering the Ψcyt of the roots: 1. Maintain the turgor 2. Driving force for water up-take 3. Stabilisation of membrane and proteins 4. Scavenger of ROS and organic radicals Drought Stress/Water Stress/Osmotic Stress... Gesamtmenge an gelösten Stoffen und einzelnen Substanzen In Wurzel (A/C) und Blatt (B/D) unter verschiedenen Stressbedingungen (Wasserpotential) nach 30 min und 24 Std. Ogawa and Yamauchi 2006 Drought Stress/Water Stress/Osmotic Stress... Adaptation reactions against water stress Stomatal control: stomatal close during water deficit in response to ABA Changes in water potential, stomatal resistance , and ABA content in maize in response to water stress. As the soil dried out, the water potential of the leaf decreased, and the ABA content and stomatal resistance increased a) During water deficit, some of the ABA stored in the chloroplasts is released to the apoplast of the mesophyll cells b) ABA is synthesized at a higher rate, and more ABA accumulate in the leaf apoplast Function of abscisic acid Drought response: Stomata closure Acidic xylem sap favors uptake of the undissociated form of ABA (ABAH) by the mesophyll cells During water stress, the slightly alkaline xylem sap favors the dissociation of ABAH to ABA- Because ABA- does not easily pass through membranes, under conditions of water stress, more ABA reaches guard cells … stomatal closure mediated by ABA cADPR – cyclic ADP-ribose ROS – reactive oxygen species R – Receptor IP3 – inositol triphosphate NO – Nitric oxide PA – Phosphopatidic acid PLC – phospholipase D S1P – Spingosine-1Phosphate PLC – phospholipase C 29 Drought Stress/Water Stress/Osmotic Stress... Generation and scavenging of ROS and lipid peroxides Drought Stress/Water Stress/Osmotic Stress... Generation and scavenging of ROS and lipid peroxides Generation and scavenging of superoxide radical and hydrogen peroxide, and hydroxyl radicalinduced lipid peroxidation and glutathione peroxidase-mediated lipid (fatty acid) stabilization. APX, Ascorbate peroxidase; ASC, Ascorbate; DHA, Dehydroascorbate; DHAR, Dehydroascorbate reductase; Fd, Ferredoxin; GR, Glutathione reductase; GSH, Red glutathione; GSSG, Oxiglutathione; HO, Hydroxyl radical; LH, Lipid; L LOO-1; LOOH, Unstable lipid radicals and hydroperoxides; LOH, Stable lipid (fatty acid); MDHA, Monodehydro-ascorbate; MDHAR, Mono dehydro-ascorbate reductase; NE, Non-enzymatic reaction; PHGPX, Phospholipid-hydroperoxide glutathione peroxidase; SOD, Superoxide dismutase. Water stress signalling • Signal of water stress - decreased cell water content, - decreased water potential and its components osmotic and pressure potentials, - increased concentration of solutes, - decreased cell volume, - change in membrane tension, - changes in structure of macromolecules due to changes in their hydration, - changes in interaction between cell wall and plasmalemma • Water stress receptors are not sufficiently known, probably different for different signals • For root-shoot communication hydraulic and chemical signals are used • Direct and indirect effects of water stress Drought Stress/Salt Stress/Osmotic Stress... Stress signalling Signalling pathway leading to stress induced changes in gene expression (Xiong et al. 2002) A common framework model for the signal transduction of abiotic stress in higher plants. Drought Stress/Salt Stress/Osmotic Stress... The simple signalling model Transcriptional network of abiotic stress responses Saibo, N. J. M. et al. Ann Bot 2009 103:609-623; doi:10.1093/aob/mcn227 Copyright restrictions may apply. DREB1A gene regulates a lot of target genes to enhance tolerance to environmental stresses Environmental Stress Drought and High salinity Signal Perception Signal Perception DREB1A Tolerance gene expression Tolerance gene expression Overexpression of DREB1A Tolerance gene expression Tolerance gene expression DREB1A Tolerance gene expression Tolerance gene expression Expression of more than 40 genes Enhanced expression of more than 40 genes Environmental stress tolerance Enhanced stress tolerance Strategies towards increased stress tolerance Combinations of promoters and transcription factors ① Constitutive promoter (35S) + DREB1A Constitutive promoter DREB1A gene The constitutive promoter leads to generate DREB1A products all the time ② Stress-inducible promoter (RD29A) + DREB1A Stress-inducible promoter DREB1A gene The stress-inducible promoter leads to generate DREB1A products only under stress conditions Drought Stress Dehydration tolerance of transgenic plants Liu et al., 1998, Plant Cell, 10, 1391-1406 Drought Tolerance of the 35S:DREB1Ab and 35S:DREB1Ac Transgenic Plants. Control, 3-week-old plants growing under normal conditions; drought stress, water withheld from plants for 2 weeks. Percentages of surviving plants and numbers of surviving plants per total number of tested plants are indicated under the photographs. wt, wild type. Drought Stress/Cold Stress/Osmotic Stress... stress tolerance of transgenic plants Arabidopsis ::DREB1 2 Tage -6 °C 2 Wochen ohne Wasser 2 Stunden 600 mM NaCl Drought Stress/Water Stress/Osmotic Stress... stress tolerance of transgenic plants Tolerance and growth properties in dependence of the promoter Drought Stress/Water Stress/Osmotic Stress... dehydration tolerance of transgenic plants Drought Stress/Water Stress/Osmotic Stress... stress tolerance of transgenic plants Drought Stress/Water Stress/Osmotic Stress... stress tolerance of transgenic plants Drought-stress tolerance of transgenic Arabidopsis plants expressing constitutively active DREB2A (DREB2A CA). Wild-type plants could not survive without watering for two weeks. In contrast, three lines of DREB2A CA transgenic plants could survive despite severe desiccation. Plants marked with * and ** had significantly higher survival rates than the wild type (x2 test, P < 0.05 and P < 0.01, respectively). Drought Stress/Water Stress/Osmotic Stress... Expression of ABFs (bZIP proteins; ABRE binding factors) is induced by ABA and various stress treatments ABA-responsive cis-elements 1. (C/T)ACGTGGC 2. GGACACGTGGC 3. ACGCGTGTCCTC 4. GCCGCGTGGC 5. GGACGCGTGGC Analysis of ABF expression. ABA and stress inducibility of ABF expression were examined by RNA gel blot analysis or RT- PCR. A, inducibility of ABF expression. 25 µg of total RNAs isolated from untreated plants or plants treated with ABA, high salt, cold, or drought were transferred to a membrane and probed with specific probes. B, time course of ABA induction. RT- PCR reactions were performed using 0.5 mg of total RNAs from plants treated with 100 mM ABA for 0 min, 30 min, 1 h, 2 h, 4 h, 8 h 12 h, 16 h, and 24 h. actin, a control reaction performed with an actin gene of A. thaliana. Choi et al., 2000, JBC, 275, 1723-1730 Drought Stress/Water Stress/Osmotic Stress... Both ABF3- and ABF4 (bZIP proteins; ABRE binding factors) -transgenic plants survived the drought conditions better than did the wild-type plants 16% 33% Kang et al., 2002, Plant Cell, 14, 343-357 Drought Tolerance of 35S-ABF3 and 35S-ABF4 Plants. (A) Transgenic and wildtype plants were grown on soil in the same container for 2 weeks, withheld from water for 11 days, and then rewatered. The photographs were taken 3 days after the rewatering. (B) Plants at similar developmental stages (2-week-old wild-type plants and 3-week old ABF4 plants) were withheld from water for 12 days and then rewatered. The photographs were taken 3 days after the rewatering. Drought Stress/Water Stress/Osmotic Stress... Both ABF3- and ABF4-transgenic plants survived the drought conditions better than did the wild-type plants 20% 85% Drought Tolerance of 35S-ABF3 and 35S-ABF4 Plants. Stomatal aperture of ABF transgenic plants. Stomatal guard cells were observed in the middle of the watering period. Kang et al., 2002, Plant Cell, 14, 343-357 Drought Stress/Water Stress/Osmotic Stress... Both ABF3- and ABF4-transgenic plants survived the drought conditions better than did the wild-type plants Drought Tolerance of 35S-ABF3 and 35S-ABF4 Plants. (C) and (D) Leaves of similar developmental stages were excised and weighed at various times after the detachment. Kang et al., 2002, Plant Cell, 14, 343-357 Changes in gene expression induced by drought, salinity or cold (Seki et al. 2002) Classification of the drought, cold or high-salinity stress inducible genes identified on the basis of microarray analyses. In total, 277 drought-inducible, 53 cold-inducible and 194 high-salinity stress-inducible genes were identified by cDNA microarray analysis. The drought, cold or high-salinity stress-inducible genes identified were grouped into the following seven groups: (1) highly cold-stress-inducible; (2) highly drought-stress-inducible; (3) highly high-salinitystress inducible; (4) drought, cold and high-salinity stress-inducible; (5) genes that were highly induced by drought and high-salinity stress; (6) genes that were highly induced by drought and cold stress; (7) genes that were highly induced by cold and highsalinity stress. The number of genes whose expression ratio is more than fivefold for each stress treatment and less than fivefold for the other stress treatments is indicated. Numbers in parentheses represent the number of genes whose expression ratio is more than fivefold for each stress treatment and less than threefold for the other stress treatments. Water stress and protein synthesis 1) Inhibition of synthesis of some proteins 2) Stimulation of synthesis of other proteins 3) Synthesis of specific stress proteins • A) proteins taking part in signal transduction and gene expression, e.g. transcription factors (MYC, MYB), protein kinases (MAPK), enzymes of phospholipid metabolism (phospholipase C, D) • B) proteins participating in stress tolerance, e.g. membrane proteins, proteins of water and ion channels, protection factors (chaperones, LEA proteins), syntases of osmoprotectants, stress proteins localized in chloroplasts, specific inhibitors of proteolytic activity, antioxidants, antioxidative enzymes, proteins taking part in recovery after stress Functions of water-stress-inducible gene products Regulatory proteins Functional proteins Water channel protein Transcription factors (MYB, MYC, DREB, AREB) Detoxification enzymes Protection factors of Macromolecules (LEA protein) Key enzymes for Osmolyte biosynthesis (proline, sugar) Protein kinases (CDPK, MAPK) Water stress Phospholipid metabolism (PLC, PIP5K) ABA biosynthetic enzyme Detoxication enzyme