The CBL-interacting protein kinase OsCIPK1 phosphorylated by SAPK10 positively regulates responses to ABA and osmotic stress in rice

SubclassⅢsucrose nonfermenting1-related protein kinase 2s(SnRK2s)function in ABA and abiotic stress responses by unknown mechanisms.We found that osmotic stress/ABA-activated protein kinase 10(SAPK10),a member of rice SnRK2s,physically interacted with CBL-interacting protein kinase 1(OsCIPK1).OsCIPK1 expression was up-regulated by ABA and PEG treatment,and overexpression increased the ABA sensitivity of seed germination and root growth and plant osmotic stress tolerance.Osmotic stress or ABA-induced activation of OsCIPK1 is dependent on SAPK10.SAPK10 phosphorylates Thr-24 of OsCIPK1 in vitro,and this phosphorylation increases the activity of OsCIPK1 and positively regulates the function of OsCIPK1 in ABA responses and plant osmotic stress tolerance.This study suggests that OsCIPK1 is a direct phosphorylated substrate of SAPK10,and SAPK10-mediated phosphorylation of OsCIPK1 functions in ABA signaling and increases rice osmotic stress tolerance.

Overexpression of the transcription factor MdWRKY115 improves drought and osmotic stress tolerance by directly binding to the MdRD22 promoter in apple

Abiotic stress reduces plant yield and quality.WRKY transcription factors play key roles in abiotic stress responses in plants,but the molecular mechanisms by which WRKY transcription factors mediate responses to drought and osmotic stresses in apple(Malus×domestica Borkh.)remain unclear.Here,we functionally characterized the apple GroupⅢWRKY gene MdWRKY115.qRT-PCR analysis showed that MdWRKY115 expression was up-regulated by drought and osmotic stresses.GUS activity analysis revealed that the promoter activity of MdWRKY115 was enhanced under osmotic stress.Subcellular localization and transactivation assays indicated that MdWRKY115 was localized to the nucleus and had a transcriptional activity domain at the N-terminal region.Transgenic analysis revealed that the overexpression of MdWRKY115 in Arabidopsis plants and in apple callus markedly enhanced their tolerance to drought and osmotic stresses.DNA affinity purification sequencing showed that MdWRKY115 binds to the promoter of the stress-related gene MdRD22.This binding was further verified by an electrophoretic mobility shift assay.Collectively,these findings suggest that MdWRKY115 is an important regulator of osmotic and drought stress tolerance in apple.

JA-mediated MYC2/LOX/AOS feedback loop regulates osmotic stress response in tea plant

Osmotic stress caused by low-temperature,drought and salinity was a prevalent abiotic stress in plant that severely inhibited plant development and agricultural yield,particularly in tea plant.Jasmonic acid(JA)is an important phytohormone involving in plant stress.However,underlying molecular mechanisms of JA modulated osmotic stress response remains unclear.In this study,high concentration of mannitol induced JA accumulation and increase of peroxidase activity in tea plant.Integrated transcriptome mined a JA signaling master,MYC2 transcription factor is shown as a hub regulator that induced by mannitol,expression of which positively correlated with JA biosynthetic genes(LOX and AOS)and peroxidase genes(PER).CsMYC2 was determined as a nuclei-localized transcription activator,furthermore,ProteinDNA interaction analysis indicated that CsMYC2 was positive regulator that activated the transcription of CsLOX7,CsAOS2,CsPER1 and CsPER3via bound with their promoters,respectively.Suppression of CsMYC2 expression resulted in a reduced JA content and peroxidase activity and osmotic stress tolerance of tea plant.Overexpression of CsMYC2 in Arabidopsis improved JA content,peroxidase activity and plants tolerance against mannitol stress.Together,we proposed a positive feedback loop mediated by CsMYC2,CsLOX7 and CsAOS2 which constituted to increase the tolerance of osmotic stress through fine-tuning the accumulation of JA levels and increase of POD activity in tea plant.

Adaptive strategy of Nitraria sibirica to transient salt,alkali and osmotic stresses via the alteration of Na+/K+fluxes around root tips

Nitraria sibirica Pall.is an important shrub with a strong salt-alkali tolerance,but the mechanism underlying this tolerance remains obscure.In this study,N.sibirica,with salt-sensitive Vigna radiata(Linn.)Wilczek as the control,was subjected to transient salt stress(100 mM NaCl),alkali stress(50 mM Na_(2)CO_(3)),and osmotic stress(175 mM mannitol).The ionic fluxes of Na^(+)and K^(+)in the root apical region were measured.Results show that,under salt and alkali stress,N.sibirica roots exhibited higher capacities to limit Na+influx and reduce K+efflux,thereby resulting in lower Na^(+)/K^(+)ratios compared with V.radiata roots.Alkali stress induced stronger Na^(+)influx and K+efflux in the root salt stress treatment;Na^(+)influx was mainly observed in the root cap,while K^(+)efflux was mainly observed in the elongation zone.While under osmotic stress,N.sibirica roots showed stronger Na+efflux and weaker K+efflux than V.radiata roots.Na+efflux was mainly observed in the root elongation zone,while K+efflux was in the root cap.These results reveal the ionic strategy of N.sibirica in response to transient salt,alkali,and osmotic stresses through the regulation of Na+/K+flux homeostasis.

Relationship between ATPase activity and conjugated polyamines in mito-chondrial membrane from wheat seedling roots under osmotic stress

The effects of osmotic stress on the ATPase activity, the contents of —SH group and conjugated polyamines in mitochondrial membrane from wheat seedling [Triticum aestivum L. cv. Yumai No.18(drought-tolerant) and cv. Yumai No.9(drought-sensitive)] roots were investigated. The results showed that ATPase activity and —SH group content decreased with polyethylene glycol(PEG) 6000(-0.55 MPa) treatment for 7 d, in concert with the decrease of the ratio of noncovalently conjugated spermidine(NCC-Spd)/noncovalently conjugated putrescine(NCC-Put) and increase of the covalently conjugated putrescine(CC-Put). Osmotic stress injury to Yangmai No.9 seedlings was alleviated greatly with 1 mmol/L exogenous spermidine(Spd), in concert with marked increases of the ratio of NCC-Spd/NCC-Put, —SH group contents and ATPase activity in mitochondrial membrane. Under osmotic stress, the concomitant treatment of Yumai No.18 seedlings with methylglyoxyl bis(guanylhydrazone) (MGBG), an inhibitor of S-adenosyl methionine decarboxylase(SAMDC), and phenanthrolin (o-Phen), an inhibitor of transglutaminase(TGase), caused a significant decrease of the ratio of NCC-Spd / NCC-Put, CC-Put contents, respectively, in concert with the marked decreases of ATPase activity, —SH group content and its tolerance to osmotic stress. All the results above suggested that osmotic stress tolerance of wheat seedlings was associated with the ATPase activity, the contents of —SH group, NCC-Spd and CC-Put in mitochondrial membrane.

Interactive effect of shade and PEG-induced osmotic stress on physiological responses of soybean seedlings

Intensively farmed crops used to experience numerous environmental stresses.Among these,shade and drought significantly influence the morpho-physiological and biochemical attributes of plants.However,the interactive effect of shade and drought on the growth and development of soybean under dense cropping systems has not been reported yet.This study investigated the interactive effect of PEG-induced osmotic stress and shade on soybean seedlings.The soybean cultivar viz.,C-103 was subjected to PEG-induced osmotic stress from polyethylene glycol 6000(PEG-6000)under shading and non-shading conditions.PEG-induced osmotic stress significantly reduced the relative water contents,morphological parameters,carbohydrates and chlorophyll contents under both light environments.A significant increase was observed in osmoprotectants,reactive oxygen species and antioxidant enzymes in soybean seedlings.Henceforth,the findings revealed that,seedlings grown under non-shading conditions produced more malondialdehyde and hydrogen peroxide contents as compared to the shade-treated plants when subjected to PEG-induced osmotic stress.Likewise,the shaded plants accumulated more sugars and proline than non-shaded ones under drought stress.Moreover,it was found that nonshaded grown plants were more sensitive to PEG-induced osmotic stress than those exposed to shading conditions,which suggested that shade could boost the protective mechanisms against osmotic stress or at least would not exaggerate the adverse effects of PEG-induced osmotic stress in soybean seedlings.

Changes induced by osmotic stress in the morphology, biochemistry, physiology, anatomy and stomatal parameters of almond species(Prunus L. spp.) grown in vitro

We investigated the influence of different levels of osmotic stress on growth and development in selected wild almond species （eight Prunus spp.） grown in vitro. The study, while endorsing the efficacy of in vitro screening of auxiliary buds of wild almond for osmotic stress tolerance, showed species variability in its response to osmotic stress. Osmotic stress reduced growth and development of all the species. How-ever, the putative tolerant Prunus spp. showed better performance than the putative susceptible genotypes. On average there was an 80% de-crease in shoot dry weight at -1.2 MPa. Reduction in shoot weight was more common in osmotic stress-susceptible species in the section labeled‘Euamygdalus’. The tolerant Prunus species produced smaller changes in biochemical responses than the sensitive cultivars for malondialdehyde content, catalase activity, relative permeability of protoplast membranes, and net photosynthetic rate. The tolerant species maintained cell integrity better than drought sensitive species. Wild almond species in the section labeled ‘Spartioides’ （Prunus arabica （Olivier） Neikle, Prunus glauca＆amp;nbsp;（Browicz） A.E. Murray, Prunus scoparia Spach） and ‘Lycioides’ （Pru-nus lycioides Spach, Prunus reuteri Bossi. et Bushe） were best adapted to osmotic stress. Increase in chlorophyll concentration and leaf thickness under high osmotic stress can be considered as preliminary selection parameters for osmotic stress tolerance in Prunus spp. The study con-firmed the efficacy of the in vitro method for screening of large number of genotypes for osmotic stress tolerance in wild almond species.

Loss of Responsiveness to Osmotic Stress in Maize Root:the Effect of Water Channel Blocker HgCl_2

In order to investigate the effect of water channel blocker HgCl2 on the hydraulic resistance in roots of maize seedlings, a xylem pressure probe was used to monitor the changes in root xylem pressure in response to NaCl- or mannitol-induced osmotic stresses before and after the application of HgCl2. When the maize roots were subjected to 500 umol L-1 HgCl2 in root bathing solution, not only a considerable decline in xylem pressure （increase in xylem tension） was observed, but the loss of responsiveness of the plant to both salt- and mannitol-induced osmotic stresses in terms of xylem pressure change was seen as well when the transpiration rate of the plant was not significantly changed. The results are similar but different from the reversed osmosis by the Fenton reaction in the internodes of Chara coralline, showing that the mechanisms of water transport across cell membrane in plant roots are far more complicated than expected.

Relationship between H_(2)O_(2) accumulation and NO synthesis during osmotic stress:promoted somatic embryogenesis of Fraxinus mandshurica

Osmotic stress promotes somatic embryogenesis of Fraxinus mandshurica,which leads to accumulation of reactive oxygen species(ROS).The single pieces of cotyledons of F.mandshurica were used as explants to induce somatic embryogenesis in osmotic-stress medium.Furthermore,the hydrogen peroxide H_(2)O_(2) content of explanted cells was varied by adding exogenous H_(2)O_(2) or catalase solution to assess the effects of the exogenous H_(2)O_(2)on somatic embryogenesis,intracellular H_(2)O_(2)accumulation,and the relationship between signaling mediated by ROS or reactive nitrogen species.The results revealed that exogenous H_(2)O_(2)(100?300μmol L^(–1))increased the number of somatic embryos.On 60th day of exogenous H_(2)O_(2)(200μmol L^(–1))treatment,the number of somatic embryos of explants treated,which was 136.54%,was higher than the control.Moreover,exogenous H_(2)O_(2)(100μmol L^(–1))significantly increased the intracellular H_(2)O_(2)content and enhanced the activities of superoxidase dismutase and peroxidase.Finally,exogenous H_(2)O_(2)(100μmol L^(–1))activated the intracellular non-enzymatic pathway for nitric oxide(NO)synthesis.The somatic embryogenesis in broadleaf trees increases with the change of endogenic ROS content,and depends on the upregulation of antioxidant enzymes.Both H_(2)O_(2)and NO,as signaling molecules,were found to be involved in the process of somatic embryogenesis in broadleaf trees.In the process of exogenous H_(2)O_(2)promoting somatic embryogenesis,NO synthesis depended on non-enzymatic reactions.These results provide a scientific basis for resolving the mechanism by which ROS levels are regulated during somatic embryogenesis of broadleaf trees and establish a reasonable and efficient technology system for regulating somatic embryogenesis of trees.

Effects of short-term osmotic stress on leaf hydraulic conductivity and ZmPIPs mRNA accumulation in maize seedlings

Plants maintain water balance by varying hydraulic properties, and plasma membrane intrinsic proteins（PIPs） may be involved in this process. Leaf xylem and root hydraulic conductivity and the m RNA contents of four highly expressed Zm PIP genes（Zm PIP1;1, Zm PIP1;2, Zm PIP2;2, and Zm PIP2;5） in maize（Zea mays） seedlings were investigated. Under well-watered conditions, leaf hydraulic conductivity（K_（leaf）） varied diurnally and was correlated with whole-plant hydraulic conductivity. Similar diurnal rhythms of leaf transpiration rate（E）, K_（leaf） and root hydraulic conductivity（K_（root）） in well-watered plants are important for maintaining whole-plant water balance. After 2 h of osmotic stress treatment induced by 10% polyethylene glycol 6000, the K_（root） of stressed plants decreased but K_（leaf） increased, compared with well-watered plants. The m RNA contents of four Zm PIPs were significantly up-regulated in the leaves of stressed plants, especially for Zm PIP1;2. Meanwhile, Zm PIP2;5 was significantly down-regulated in the roots of stressed plants. After 4 h of osmotic stress treatment, the E and leaf xylem water potentials of stressed plants unexpectedly increased. The increase in K_（leaf） and a partial recovery of K_（root） may have contributed to this process. The m RNA content of Zm PIP1;2 but not of the other three genes was up-regulated in roots at this time. In summary, the m RNA contents of these four Zm PIPs associated with K_（leaf） and K_（root） change in maize seedlings during short-term osmotic stress, especially for Zm PIP1;2 and Zm PIP2;5, which may help to further reveal the hydraulic resistance adjustment role of Zm PIPs.

GmGPDH12,a mitochondrial FAD-GPDH from soybean, increases salt and osmotic stress resistance by modulating redox state and respiration

In plants,glycerol-3-phosphate dehydrogenase(GPDH)catalyzes the interconversion of glycerol-3-phosphate(G3P)and dihydroxyacetone phosphate(DHAP)coupled to the reduction/oxidation of the nicotinamide adenine dinucleotide(NADH)pool,and plays a central role in glycerolipid metabolism and stress response.Previous studies have focused mainly on the NAD+-dependent GPDH isoforms,neglecting the role of flavin adenine dinucleotide(FAD)-dependent GPDHs.We isolated and characterized three mitochondrialtargeted FAD-GPDHs in soybean,of which one isoform(GmGPDH12)showed a significant transcriptional response to NaCl and mannitol treatments,suggesting the existence of a major FAD-GPDH isoform acting in soybean responses to salt and osmotic stress.An enzyme kinetic assay showed that the purified GmGPDH12 protein possessed the capacity to oxidize G3P to DHAP in the presence of FAD.Overexpression and RNA interference of GmGPDH12 in soybean hairy roots resulted in elevated tolerance and sensitivity to salt and osmotic stress,respectively.G3P contents were significantly lower in GmGPDH12-overexpressing hair roots and higher in knockdown hair roots,indicating that GmGPDH12 was essential for G3P catabolism.A significant perturbation in redox status of NADH,ascorbic acid(ASA)and glutathione(GSH)pools was observed in GmGPDH12-knockdown plants under stress conditions.The impaired redox balance was manifested by higher reactive oxygen species generation and consequent cell damage or death;however,overexpressing plants showed the opposite results for these traits.GmGPDH12 overexpression contributed to maintaining constant respiration rates under salt or osmotic stress by regulating mRNA levels of key mitochondrial respiratory enzymes.This study provides new evidence for the roles of mitochondria-localized GmGPDH12 in conferring resistance to salt or osmotic stress by maintaining cellular redox homeostasis,protecting cells and respiration from oxidative injury.

Selection of Maize Genotypes with Tolerance to Osmotic Stress Associated with Salinity

Thirteen different inbred lines in relation to the type of grain and life cycles were characterized by testing for osmotic stress associated with salinity. The identification of tolerant genotypes would be an effective strategy to overcome the saline stress. Osmotic stress reduces immediately the expansion of the roots and young leaves which determine a reduction in the size of the plant. A completely randomized design was adopted to test seedlings under controlled conditions of light and temperature. Two treatments were used: 0 mM NaCl (as control) and 100 mM NaCl. After 15 days of complete salinization, the seedlings were harvested and several morphological traits were measured. The morphological traits of growth were leaf growth (Ar1, Ar2, Ar3 and Ar4), dry masses of shoot and root (SDM and RDM, respectively). Also, traits associated with water economy were registered: leaf water loss (LWL) and relative water content (RWC). The morphological traits were expressed in relative terms, while the traits associated with the economy of water were expressed in absolute terms. Uni and multivariate techniques were applied to identify genotypes with divergent behaviors to osmotic stress tolerance. Also, a Tolerance Index was employed to identify superior genotypes. Four clusters were obtained after applying a Cluster Analysis and Principal Component Analysis (PCA). The genotypes were compared to each other with a test of DMS. The results obtained with different statistical techniques converged. Some variables presented a differential weight classification of genotypes. The morphological traits like RDM, SDM, Ar3, Ar4 and Ar5 were the most discriminating. Tolerance Index allowed to classify genotypes, thus SC2 and AD3 lines were that reached highest value of the index and therefore would be tolerant lines, while AF3 and LP3 had a low index and were seen as sensible.

Osmotic Stress Resistance of Transgenic Tobacco Expressed Soybean Lysine-rich Protein Gene

PM2 gene （accession number： M80664） with high lysine content from soybean （Glycine max） was found in GenBank by changing three BLASTp parameters. Amino acid composition analysis of PM2 showed that Lys content was on the high level of 18.22%. Protein encoded by PM＇2 also belonged to the family of late embryogenesis abundant （LEA） proteins, which was considered that it had a strong relation with the abiotic stress resistance. In this experiment, PM2 gene was obtained from dry soybean seeds by RT-PCR, plant expression vector pEMTPM2 was constructed, and then transformed into tobacco by using agrobacterium-mediated method. Eight salt and drought tolerant lines were obtained from 31 differentiated lines. Real-time PCR showed that PM2 gene overexpressed in all four PCR positive lines with the osmotic stress resistance. These results confirmed that the overexpression of PM2 gene enhanced the osmotic stress resistance of transgenic tobacco.

Nrf2 Pathway Involvement in the Beneficial Skin Effects of Moderate Ionic Osmotic Stress—The Case of The Dead Sea Water

Objectives: Exposing skin to moderate ionic osmotic stress (MIOS) triggers several biochemical responses. The objective of this work is to reveal the mechanism triggered by MIOS on the skin surface. Furthermore, this work aims to study the involvement of the Nrf2 (nuclear factor erythroid-2-related factor 2) pathway, activated by MIOS, and its beneficial effect in protecting skin against stress via the stimulation of phase II enzymes. Methods: HaCaT cells and human skin organ culture were exposed to Dead Sea Water (DSW) as MIOS inducers and the induction of internal ROS elevation, Nrf2 translocation, mRNA gene expressions of the phase II enzymes, heme-oxygenase 1 (HO1), and Catalase (CAT) were determined. Results: Skin exposure to MIOS increases Nrf2 translocation to the nucleus, leading to increased levels of ROS, HO1, and CAT. Furthermore, exposing skin to MIOS promotes protection against UVB-related risks. This is demonstrated by attenuation of the expression of biomarkers, related to UVB-induced damage, Caspase-3, IL-8, and IL-1β. Conclusions: Skin exposure to MIOS leads to the activation of Nrf2 skin defense pathway and, therefore, could present beneficial advantages to human skin health, as demonstrated on human skin models. The beneficial effects of MIOS, induced by DSW are significantly superior to eq. NaCl brine, suggests that MIOS protection of skin against stress is partially related to specific mineral combinations.

How plants sense and respond to osmotic stress

Drought is one of the most serious abiotic stresses to land plants.Plants sense and respond to drought stress to survive under water deficiency.Scientists have studied how plants sense drought stress,or osmotic stress caused by drought,ever since Charles Darwin,and gradually obtained clues about osmotic stress sensing and signaling in plants.Osmotic stress is a physical stimulus that triggers many physiological changes at the cellular level,including changes in turgor,cell wall stiffness and integrity,membrane tension,and cell fluid volume,and plants may sense some of these stimuli and trigger downstream responses.In this review,we emphasized water potential and movements in organisms,compared putative signal inputs in cell wall-containing and cell wall-free organisms,prospected how plants sense changes in turgor,membrane tension,and cell fluid volume under osmotic stress according to advances in plants,animals,yeasts,and bacteria,summarized multilevel biochemical and physiological signal outputs,such as plasma membrane nanodomain formation,membrane water permeability,root hydrotropism,root halotropism,Casparian strip and suberin lamellae,and finally proposed a hypothesis that osmotic stress responses are likely to be a cocktail of signaling mediated by multiple osmosensors.We also discussed the core scientific questions,provided perspective about the future directions in this field,and highlighted the importance of robust and smart root systems and efficient source-sink allocations for generating future high-yield stress-resistant crops and plants.

Role of Arabidopsis UV RESISTANCE LOCUS 8 in Plant Growth Reduction under Osmotic Stress and Low Levels of UV-B

In high-light environments, plants are exposed to different types of stresses, such as an excess of UV-B, but also drought stress which triggers a common morphogenic adaptive response resulting in a general reduction of plant growth. Here, we report that the Arabidopsis thaliana UVRESISTANCE LOCUS 8 （UVR8） gene, a known regulator of the UV-B morphogenic response, was able to complement a Saccharomyces cerevisiae osmo-sensitive mutant and its expression was induced after osmotic or salt stress in Arabidopsis plants. Under low levels of UV-B, plants overexpressing UVR8 are dwarfed with a reduced root development and accumulate more flavonoids compared to control plants. The growth defects are mainly due to the inhibition of cell expansion. The growth inhibition triggered by UVR8 overexpression in plants under low levels of UV-B was exacerbated by mannitol-induced osmotic stress, but it was not significantly affected by ionic stress. In contrast, uvr8-6 mutant plants do not differ from wild-type plants under standard conditions, but they show an increased shoot growth under high-salt stress. Our data suggest that UVR8-mediated accumulation of flavonoid and possibly changes in auxin homeostasis are the underlying mechanism of the observed growth phenotypes and that UVR8 might have an important role for integrating plant growth and stress signals.

Effects of lanthanum on the growth and essential oil components of lavender under osmotic stress

The effects of lanthanum on growth, soluble sugar content, essential oil contents and quality were studied in lavender plant（Lavandula Angustifolia Mill, Variety 701） under osmotic stress by means of PEG-6000 solution. The results show that osmotic stress could reduce the growth rate of lavender, and increase the content of SSC（soluble sugar concentration） to some extent. Compared with control group,the contents of essential oil in flowers and leaves respectively increase by 45.6% and 48.3% in the osmotic stress group induced by 15% PEG-6000. However, the presence of lanthanum can make the contents of essential oil in the flowers and leaves of stressed lavender plants enhance by 19.4% and 18.6%, respectively. Under osmotic stress, the relative contents of four kinds of lavender essential oil compositions of camphor, linalool, linalyl acetate and lavandulol acetate in the lavender flowers were successively 22.63%,0.61%,25.46%,7.03%,and 20.17%,0.62%,20.72%,10.80%,respectively in the presence and absence of lanthanum. The contents of all four main components of lavender essential oils meet the requirements of the national standard under osmotic stress in the presence of lanthanum. However, in the absence of lanthanum, the contents of linalyl acetate and lavandulol acetate does not meet the requirements of the national standard under osmotic stress. Moreover, the contents of five components of borneol, camphor,eudesmol, caryophyllene oxide and bicyclic sesquiphellandrene from the essential oil of lavender leaves are 12.89%, 3.84%, 8.76%, 11.30% and 8.10%, respectively. The total content of above five components accounts for 44.89% of the essential oil of the lavender leaves. Particularly, the borneol content in leaf essential oil is up to 12.89%. It is 125.2 times the amount of borneol in the flower essential oil. In conclusion, the suitable concentration of lanthanum can improve the adaptability of lavender plants, and heighten the content and quality of lavender essential oil to some extent under osmotic stress.

Osmotic Stress Modulates the Balance between Exocytosis and Clathrin-Mediated Endocytosis in Arabidopsis thaliana

The sessile life style of plants creates the need to deal with an often adverse environment, in which water availability can change on a daily basis, challenging the cellular physiology and integrity. Changes in os- motic conditions disrupt the equilibrium of the plasma membrane： hypoosmotic conditions increase and hyperosmotic environment decrease the cell volume. Here, we show that short-term extracellular osmotic treatments are closely followed by a shift in the balance between endocytosis and exocytosis in root mer- istem cells. Acute hyperosmotic treatments （ionic and nonionic） enhance clathrin-mediated endocytosis simultaneously attenuating exocytosis, whereas hypoosmotic treatments have the opposite effects. In addition to clathrin recruitment to the plasma membrane, components of early endocytic trafficking are essential during hyperosmotic stress responses. Consequently, growth of seedlings defective in elements of clathrin or early endocytic machinery is more sensitive to hyperosmotic treatments. We also found that the endocytotic response to a change of osmotic status in the environment is dominant over the presum- ably evolutionary more recent regulatory effect of plant hormones, such as auxin. These results imply that osmotic perturbation influences the balance between endocytosis and exocytosis acting through clathrin- mediated endocytosis. We propose that tension on the plasma membrane determines the addition or removal of membranes at the cell surface, thus preserving cell integrity.

Effect of Osmotic Stress in Early Stages of Ontogenesis on Root Respiration, Growth, Sugar Content,and Cell Injury in Maize Seedlings Differing in Drought Sensitivity

Cultivars of maize （Zea mays L.） with different sensitivity to drought were exposed to 0.3 mol/L sorbitol （-1.4 MPa water potential） for 24 h. Exposure to water deficiency significantly reduced the growth of both shoots （coleoptile and hypocotyl） and roots. Shoot growth was inhibited more than the growth of roots. Osmotic stress enhanced accumulation of soluble sugars. Electrolyte leakage, a cell injury index, was slightly increased after 0.3 mol/L sorbitoh Respiration was measured in the presence and absence of 2,6-dlchloro-phenol indophenoh 2,6-Dichloro-phenol indophenol did not influence respiration rates, because statistically equal results were observed under both conditions. Total respiration （VT） decreased after osmoticum treatment. There were no significant differences in the VT among the cultlvars analysed. The decrease In VT was caused by a decline In the activities and capacities of both cytochrome （Vcyt, Vcyt） and alternative pathway （Valt, Valt） of respiration. A high residual respiration （Vres） was observed, up to 27% of total uninhibited respiration. The result of uncoupler use clearly indicated that coupling was maintained after 24 h of osmotic stress. The recovery of the respiration rate was comparable with that of non-stressed control rates. According to these observations, no possible mltochondrial damage is expected. Water deficiency did not induce a stimulation of the alternative oxidase, so we assume that the stimulation of the alternative pathway is not related to drought stress resistance; rather, the function of the alternative pathway is to balance carbon metabolism and electron transport in a response to a changing environment.

Integrin-like Protein Is Involved in the Osmotic Stressinduced Abscisic Acid Biosynthesis in Arabidopsis thaliana

We studied the perception of plant cells to osmotic stress that leads to the accumulation of abscisic acid （ABA） in stressed Arabidopsis thaliana L. cells. A significant difference was found between protoplasts and cells in terms of their responses to osmotic stress and ABA biosynthesis, implying that cell wall and/or cell wall-plasma membrane interaction are essential in identifying osmotic stress. Western blotting and immunofluorescence localization experiments, using polyclonal antibody against human integrin β1, revealed the existence of a protein similar to the integrin protein of animals in the suspension-cultured cells located in the plasma membrane fraction. Treatment with a synthetic pentapeptide, Gly-Arg-Gly-Asp-Ser （GRGDS）, which contains an RGD domain and interacts specifically with integrin protein and thus blocks the cell wall-plasma membrane interaction, significantly inhibited osmotic stress-induced ABA biosynthesis in cells, but not in protoplasts. These results demonstrate that cell wall and/or cell wall-plasma membrane interaction mediated by integrin-Iike proteins played important roles in osmotic stress-induced ABA biosynthesis in Arabidopsis thaliana.