Dehydration Treatment Research Articles

Systemic Calcium Wave Propagation in Physcomitrella patens.

The adaptation to dehydration and rehydration cycles represents a key step in the evolution of photosynthetic organisms and requires the development of mechanisms by which to sense external stimuli and translate them into signaling components. In this study, we used genetically encoded fluorescent sensors to detect specific transient increases in the Ca2+ concentration in the moss Physcomitrella patens upon dehydration and rehydration treatment. Observation of the entire plant in a single time-series acquisition revealed that various cell types exhibited different sensitivities to osmotic stress and that Ca2+ waves originated from the basal part of the gametophore and were directionally propagated towards the top of the plant. Under similar conditions, the vascular plant Arabidopsis thaliana exhibited Ca2+ waves that propagated at a higher speed than those of P. patens. Our results suggest that systemic Ca2+ propagation occurs in plants even in the absence of vascular tissue, even though the rates can be different.

Transcriptome and Expression Profiling Analysis of Recalcitrant Tea (Camellia sinensis L.) Seeds Sensitive to Dehydration

The tea plant (Camellia sinensis (L.) O. Kuntze) is an economically important woody perennial nonalcoholic health beverage crop. Tea seeds are categorized as recalcitrant and are sensitive to dehydration treatment. However, the molecular basis of this phenomenon has not been investigated. Thus, we analyzed the genome-wide expression profiles of three dehydration stages using RNA-Seq and digital gene expression (DGE) technologies. We performed de novo assembly and obtained a total of 91,925 nonredundant unigenes, of which 58,472 were extensively annotated. By a hierarchical clustering of differentially expressed genes (DEGs), we found that 8929 DEGs were downregulated and 5875 DEGs were upregulated during dehydration treatment. A series of genes related to ABA biosynthesis and signal transduction, transcription factor, antioxidant enzyme, LEA protein, and proline metabolism that have been reported to function in dehydration process were found to be downregulated. Additionally, the expression profiles of 12 selected genes related to tea seed dehydration treatment were confirmed by qRT-PCR analysis. To our knowledge, this is the first genome-wide study elucidating the possible molecular mechanisms of sensitivity of recalcitrant tea seeds to dehydration. The results obtained in this study contribute to the preservation of tea seeds as genetic resources and can also be used to explore the mechanism of dehydration sensitivity of other recalcitrant seeds.

Water-Switchable Interfacial Bonding on Tooth Enamel Surface.

Tooth enamel is a distinctive nanocomposite with a highly organized hierarchical structure made of nanometer- and micrometer-scale building blocks. This structure has an excellent mechanical function that can last for decades thanks to an effective but underexploited interfacial chemical bonding between the building blocks. In this study, the nanomechanical system test (NST), scanning electron microscope (SEM), X-ray diffraction (XRD, including powder XRD or PXRD, small angle XRD or SAXRD, and grazing incidence small angle XRD or GISAXRD), and atomic force microscope (AFM) have been employed to analyze the water-mediated bonding on the enamel surface. Via the cycling between hydration, dehydration, and rehydration treatments, a reversible change in the interfacial distance (i.e., d-space in the XRD pattern) between hydroxyapatite (HAP) nanocrystallites have been found switchable between the embrittling and toughening on the enamel surface. From the hydrated to the dehydrated conditions, an energy dissipation to deform a unit volume (1 μm3) of biocomposite on the enamel surface and subsurface has decreased by 20%. This finding can help quantify and predict biomineral-surface properties in all humidity and develop new methods to protect tooth enamel of "dry-mouth" patients.

Dehydration Stress Contributes to the Enhancement of Plant Defense Response and Mite Performance on Barley.

Under natural conditions, plants suffer different stresses simultaneously or in a sequential way. At present, the combined effect of biotic and abiotic stressors is one of the most important threats to crop production. Understanding how plants deal with the panoply of potential stresses affecting them is crucial to develop biotechnological tools to protect plants. As well as for drought stress, the economic importance of the spider mite on agriculture is expected to increase due to climate change. Barley is a host of the polyphagous spider mite Tetranychus urticae and drought produces important yield losses. To obtain insights on the combined effect of drought and mite stresses on the defensive response of this cereal, we have analyzed the transcriptomic responses of barley plants subjected to dehydration (water-deficit) treatment, spider mite attack, or to the combined dehydration-spider mite stress. The expression patterns of mite-induced responsive genes included many jasmonic acid responsive genes and were quickly induced. In contrast, genes related to dehydration tolerance were later up-regulated. Besides, a higher up-regulation of mite-induced defenses was showed by the combined dehydration and mite treatment than by the individual mite stress. On the other hand, the performance of the mite in dehydration stressed and well-watered plants was tested. Despite the stronger defensive response in plants that suffer dehydration and mite stresses, the spider mite demonstrates a better performance under dehydration condition than in well-watered plants. These results highlight the complexity of the regulatory events leading to the response to a combination of stresses and emphasize the difficulties to predict their consequences on crop production.

Genome-Wide Identification of Drought Response Genes in Soybean Seedlings and Development of Biomarkers for Early Diagnoses

Drought is a yield-limiting factor for soybean (Glycine max L. Merrill) production in North America. Understanding molecular mechanisms underlying early responses of the soybean plant to drought may help develop new techniques to manage the abiotic stress. The objectives of this research were to identify expressed genes responsive to drought stress at the seedling stage and to develop biomarkers for early diagnostic of genotypic difference in the stress tolerance. Using a GeneChip Soybean Genome Array and an improved algorithm, we identified 697 differentially expressed genes (DEGs), with 420 upregulated and 277 downregulated by a 6-h dehydration treatment. A majority of these DEGs encode transcription factors, protein kinases, hormone biosynthetic/signaling enzymes, or other regulatory proteins. MAPMAN and KEGG enrichment showed that the DEGs were mainly involved in the metabolic and hormone signaling pathways and identified the GmHDZ72/PYL/PP2C module as negative feedback of abscisic acid signaling pathway induced by dehydration stress. Ten DEGs were selected from various pathways and validated in a sample of 20 soybean cultivars varying in the level of drought tolerance. Five of the 10 validated DEGs, Glyma03g30040, Glyma11g05960, Glyma11g11430, Glyma12g22880, and Glyma16g02390, showed expression profiles strongly correlated with the plant height reduction after a 14-d drought treatment. These genes are the best candidate biomarkers to monitor plant early responses to drought stress before a symptom appears and to screen for drought-tolerant genotypes. This research provided a new set of transcriptomic data to develop gene regulatory networks underlying sensing drought signal and possible acclimatization in the early stage and enriched the genomics toolbox with a set of biomarkers for early diagnosis of drought damage and molecular breeding of drought tolerance in soybean.

The mining and evolutionary investigation of AP2/ERF genes in pear (Pyrus)

BackgroundIn plants, ERF genes participate in a variety of regulatory pathways, such as plant growth and biotic and/or abiotic stress responses. Although the genome of Chinese white pear (‘Dangshansuli’) has been released, knowledge regarding the ERF family in pear, such as gene functions, evolutionary history and expression patterns, remains limited.ResultsIn our study, a total of 155 members of ERF families were identified in pear (Pyrus bretschneideri). The Ka and Ks values suggested that whole-genome duplication (WGD) and dispersed duplication have effectively contributed to the expansion of the pear ERF family. Gene structure and phylogeny analysis divided the PbrERF family into 12 groups, and their gene functions were predicted by comparative analysis. qRT-PCR was carried out to verify the relative expression levels of 7 genes in group III using wild and cultivated pear fruits at three key developmental stages. Wild samples had higher expression of these genes than cultivated samples, especially at the enlarged fruit stage. The transcriptome data of pear seedlings subjected to dehydration treatment further revealed that 4 of the 7 genes responded to drought conditions.ConclusionThe AP2/ERF gene family is greatly expanded in pear. Comparative analysis revealed the probability of ERF genes performing functional roles in multiple pathways. Expression analysis at different stages of pear fruit development in wild and cultivated samples indicated that genes in group III might be involved in abiotic and/or biotic stresses. Further transcriptome data on seedlings subjected to drought treatment verified the potential role of ERF genes in stress response. These results will provide a valuable reference for understanding the function and evolution of the ERF family in higher plants.

Cinética de la disolución de ulexita calcinada en soluciones de sulfato de amonio

La ulexita es uno de los minerales de boro, que incluye una cantidad considerable de agua de hidratación. Se puede usar como materia prima en la producción de compuestos de boro. Una parte del agua existente en la composición química de la ulexita puede eliminarse de la matriz sólida aplicando tratamiento de deshidratación que permite obtener una estructura porosa, que aumenta la velocidad de reacción. En el presente trabajo, se estudió el efecto de la temperatura de deshidratación sobre la cinética de disolución de ulexita en soluciones de sulfato de amonio, en un reactor discontinuo estudiando los parámetros de concentración de solución, relación sólido/líquido, velocidad de agitación y la temperatura de reacción. Se determinó que la velocidad de disolución del material calcinado aumentaba al aumentar la concentración de la solución y la temperatura de reacción y con la disminución de la relación de sólido/líquido. La velocidad de disolución más alta se alcanzó con la muestra calcinada a 150 °C. Se encontró que la velocidad de disolución se ajusta al modelo pseudohomogéneo de primer orden. La energía de activación del proceso de disolución se estimó en 42 kJ·mol-1.

Preliminary comparison of atmospheric CO2 enhancement to photosynthesis of Pyropia yezoensis (Bangiales, Rhodophyta) leafy thalli and filamentous thalli

SUMMARYIntertidal macroalgae are submerged in seawater at high tide and exposed to air at low tide. When they are exposed to the air, CO2 is the main inorganic carbon source. In this study, the photosynthetic performances of PSI and PSII were measured in different generations of Pyropia yezoensis (leafy thalli and filamentous thalli) that had been exposed to air containing different CO2 concentrations. Changes in photosynthesis during dehydration and salt treatment under the different CO2 concentrations were also analyzed. The results showed that in leafy thalli, the effective photochemical quantum yield of PSII (YII) was enhanced as CO2 increased, which suggested that CO2 assimilation was enhanced and that they can utilize CO2 in the air directly, even when they are subjected to moderate stress. These findings could explain why, in P. yezoensis aquaculture, moderate exposure to air does not lead to a decrease in crop yield. However, in filamentous thalli, there were no significant differences in YII at the CO2 concentrations tested. The expression of genes involved in the Calvin cycle in leafy thalli was higher than that in filamentous thalli. CO2 uptake and biomass of P. yezoensis leafy thalli is larger than filamentous thalli, which may be due to its different carbon utilization mechanism and the adaptation of intertidal environment in the evolutionary process.

PeCHYR1, a ubiquitin E3 ligase from Populus euphratica, enhances drought tolerance via ABA-induced stomatal closure by ROS production in Populus.

SummaryDrought, a primary abiotic stress, seriously affects plant growth and productivity. Stomata play a vital role in regulating gas exchange and drought adaptation. However, limited knowledge exists of the molecular mechanisms underlying stomatal movement in trees. Here, PeCHYR1, a ubiquitin E3 ligase, was isolated from Populus euphratica, a model of stress adaptation in forest trees. PeCHYR1 was preferentially expressed in young leaves and was significantly induced by ABA (abscisic acid) and dehydration treatments. To study the potential biological functions of PeCHYR1, transgenic poplar 84K (Populus alba × Populus glandulosa) plants overexpressing PeCHYR1 were generated. PeCHYR1 overexpression significantly enhanced H2O2 production and reduced stomatal aperture. Transgenic lines exhibited increased sensitivity to exogenous ABA and greater drought tolerance than that of WT (wild‐type) controls. Moreover, up‐regulation of PeCHYR1 promoted stomatal closure and decreased transpiration, resulting in strongly elevated WUE (water use efficiency). When exposed to drought stress, transgenic poplar maintained higher photosynthetic activity and biomass accumulation. Taken together, these results suggest that PeCHYR1 plays a crucial role in enhancing drought tolerance via ABA‐induced stomatal closure caused by hydrogen peroxide (H2O2) production in transgenic poplar plants.

A rice bZIP transcription factor, OsbZIP16, regulates abiotic stress tolerance when over-expressed in Arabidopsis

Plants exposed to adverse environmental conditions are invariably compromised in their growth and development. The bZIP class of transcription factors (TF) form a large group among stress signalling components that regulate plant responses towards stress. We identified bZIP TF encoding genes that are expressed differentially in indica rice under stress and here we functionally characterize one such gene, OsbZIP16. Although, OsbZIP16 forms a clade with its orthologous monocot protein sequences, we find in our study that it can impart tolerance to abiotic stress in Arabidopsis. OsbZIP16 is expressed strongly upon dehydration, salt and ABA treatment in Oryza sativa cv. IR64 seedlings. It localizes in the cell nucleus and the gene product is capable of transcriptional activation, thus providing evidence for its capability as a functional TF. Upon overexpression in Arabidopsis, OsbZIP16ox plants show wild type morphology, however, these plants showed tolerance when subjected to drought stress at vegetative stage and set healthy seeds on recovery. The OsbZIP16ox seedlings showed reduced sensitivity to mannitol, ABA and sodium chloride during germination and also reduced ROS accumulation upon H2O2 exposure. Thus, OsbZIP16 regulates abiotic stress responses and is also a good candidate gene that can be utilised to impart tolerance in plants under water deficit conditions.

Protein changes in the shoot-tips of vanilla (Vanilla planifolia) in response to osmoprotective treatments

Cryogenic storage of vanilla shoot-tips represents the safest biotechnological strategy for the long-term conservation of the vanilla germplasm, but successful cryopreservation depends on its tolerance to both dehydration stress imposed by cryoprotective treatments and thermal stress produced by immersion in liquid nitrogen. In this work, we evaluated the impact of various osmoprotective treatments on protein expression patterns in vanilla (Vanilla planifolia) shoot-tips subjected to successive dehydration steps prior to cryopreservation. Two-dimensional electrophoretic protein profiles of shoot-tips dissected from in vitro grown plants and preconditioned on semisolid media with 0.3 M sucrose for one day, and shoot-tips preconditioned, loaded with a solution of 0.4 M sucrose and 2 M glycerol, and subsequently exposed to plant vitrification solution 3 (50% (w/v) sucrose and 50% (w/v) glycerol), were compared with non-treated dissected shoot-tips. We observed an increase in the expression level of six protein spots (fold change exceeding 1.5) and a decrease (fold change not exceeding 0.6) of ten protein spots after preconditionig treatment, whereas the profiles after preconditioning, loading and exposure to vitrification solution showed an increase in the expression level of 21 protein spots and a decrease in the expression level of 13. Most proteins identified were down-regulated and belonged to groups of biosynthesis, folding, and protein degradation. Many others were related to energetic metabolism, defense, and cell structure. These preliminary results contribute to knowledge of the proteome of this species and partially clarify its sensitivity to osmotic dehydration treatments.

OsERF71 confers drought tolerance via modulating ABA signaling and proline biosynthesis

Plants have evolved multiple protective strategies to adapt to adverse environmental conditions. Upland rice (UR) has evolved as a “drought-resistant type”. However, little is known about genes or mechanisms in UR that underlying drought tolerance at the molecular level. Here we report isolation and functional characterization of the ERF gene, OsERF71, from the UR variety, IRAT109. The expression of OsERF71 was induced by abscisic acid (ABA) and various abiotic stresses preferentially in IRAT109 under ABA, dehydration, and polyethyleneglycol (PEG) treatments. OsERF71 was verified as a nuclear-localized protein and had transcriptional activity in yeast cells. Overexpression of the OsERF71 in Nipponbare demonstrated a significant increase in tolerance to drought stress and a reduced rate of water loss. In contrast, OsERF71 interference lines were sensitive to drought stress and exhibited a higher rate of water loss. OsERF71-overexpressing lines also showed enhanced tolerance to high salinity. Moreover, OsERF71 regulated the expression of several ABA- responsive and proline biosynthesis genes under drought stress, resulting in enhanced sensitivity to exogenous ABA treatment and proline accumulation. Accordingly, we suggest that OsERF71 plays a positive role in drought stress tolerance by increasing the expression of genes associated with ABA signaling and proline biosynthesis under stress.

Acclimation-induced metabolic reprogramming contributes to rapid desiccation tolerance acquisition in Boea hygrometrica

The acquisition of rapid desiccation tolerance for the resurrection plant Boea hygrometrica requires a cycle of drought acclimation. Here, a gas chromatography-mass spectrometry based global untargeted metabolite profiling analysis was conducted on leaves of B. hygrometrica during dehydration and rehydration. Metabolic profiles between non-acclimated and drought acclimated plants were clearly distinguished in principal-component and hierarchical cluster analysis, indicating that drought acclimation response required the process in metabolic reprogramming. Accumulation of numerous metabolites involved in carbon and energy metabolism as well as amino acid and fatty acid biosynthesis was observed to be significantly changed upon drought acclimation and dehydration treatments. Using partial least-squares discriminant analysis, 25 known metabolites and 51 compounds with unknown structures were identified as putative biomarkers contributed to the discrimination between drought acclimated and non-acclimated plants. Remarkably, as revealed by transcriptome data analysis, some up-accumulated metabolites in acclimated plants, such as maltose, glutaric acid, L-tryptophan and α-tocopherol, which are in good correlation with the increased desiccation tolerance, and were precisely controlled by the transcript changes in multigene family members of related enzymes. Furthermore, genes positively correlated with the up-accumulated biomarkers in transcription abundances were enriched in multiple biological processes, such as “ubiquitin-dependent protein catabolism” and “abscisic acid-activated signaling pathway”. Taken together, our observations indicate that the accumulation of important metabolites correlates with the transcriptional activity of biosynthetic related enzymes and putative regulators involved in ubiquitination and ABA signal transduction during a short period of drought acclimation might contribute to the acquisition of rapid desiccation tolerance in B. hygrometrica.

Angiotensin AT1A receptors expressed in vasopressin-producing cells of the supraoptic nucleus contribute to osmotic control of vasopressin.

Angiotensin II (ANG) stimulates the release of arginine vasopressin (AVP) from the neurohypophysis through activation of the AT1 receptor within the brain, although it remains unclear whether AT1 receptors expressed on AVP-expressing neurons directly mediate this control. We explored the hypothesis that ANG acts through AT1A receptors expressed directly on AVP-producing cells to regulate AVP secretion. In situ hybridization and transgenic mice demonstrated localization of AVP and AT1A mRNA in the supraoptic nucleus (SON) and the paraventricular nucleus (PVN), but coexpression of both AVP and AT1A mRNA was only observed in the SON. Mice harboring a conditional allele for the gene encoding the AT1A receptor (AT1Aflox) were then crossed with AVP-Cre mice to generate mice that lack AT1A in all cells that express the AVP gene (AT1AAVP-KO). AT1AAVP-KO mice exhibited spontaneously increased plasma and serum osmolality but no changes in fluid or salt-intake behaviors, hematocrit, or total body water. AT1AAVP-KO mice exhibited reduced AVP secretion (estimated by measurement of copeptin) in response to osmotic stimuli such as acute hypertonic saline loading and in response to chronic intracerebroventricular ANG infusion. However, the effects of these receptors on AVP release were masked by complex stimuli such as overnight dehydration and DOCA-salt treatment, which simultaneously induce osmotic, volemic, and pressor stresses. Collectively, these data support the expression of AT1A in AVP-producing cells of the SON but not the PVN, and a role for AT1A receptors in these cells in the osmotic regulation of AVP secretion.

Isolation and characterization of MbWRKY2 gene involved in enhanced drought tolerance in transgenic tobacco

ABSTRACTPlant-specific WRKY transcription factors are widely involved in abiotic stress responses. In this study, a WRKY gene was isolated from Malus baccata (L.) Borkh and designated it as MbWRKY2. Subcellular localization showed that MbWRKY2 was localized in the nucleus. The expression levels of MbWRKY2 were up-regulated by abiotic stresses in M. baccata. When MbWRKY2 was introduced into tobacco, it improved drought stress tolerance in transgenic plants. The transgenic tobaccos had the higher contents of chlorophyl, proline, relative water content, AsA, and GSH, increased activities of CAT, APX, SOD, and POD, and decreased levels of MDA, H2O2, and electrolyte leakage than wild-type, especially when dealt with dehydration treatment. Moreover, the MbWRKY2-OE plants enhanced the expression of oxidative stress response and stress-related genes involved in osmotic adjustment and membrane protection. These results suggest that MbWRKY2 gene plays a positive regulatory role in drought stress response.

Comparative transcriptome analysis reveals that photosynthesis contributes to drought tolerance of Nostoc flagelliforme (Nostocales, Cyanobacteria)

:Nostoc flagelliforme, a photosynthetic cyanobacterium, is well adapted to drought. However, exploration of factors behind this important feature at a molecular level is limited because of inadequate genome data. This study used transcript sequence data of N. flagelliforme and examined its gene expression in response to drought stress on the basis of RNA-Seq. More than 1.4 G clean reads were generated for each sample using the Illumina sequencing platform. A total of 392 and 659 differentially expressed genes were identified, of which 369 and 502 were upregulated and 23 and 157 downregulated after 6- and 48-h dehydration treatments of N. flagelliforme, respectively. Functional analyses demonstrated that the genes involved in photosynthesis as well as those involved in starch and sucrose metabolism were closely related to response to drought stress. The photosynthetic products, sucrose and trehalose, played important roles as osmotic regulators, stress protectants and reactive oxygen species scavengers in response to drought stress in N. flagelliforme. Quantitative real-time reverse-transcriptase polymerase chain reaction analysis, physiological assays and cell ultrastructure observation further confirmed the results from the RNA-Seq. These findings provide novel information about the molecular mechanisms of drought tolerance in N. flagelliforme.

Morphogenetic response of shoot tips to cryopreservation by encapsulation-dehydration in a solid mutant and periclinal chimeras of Chrysanthemum\xa0\xd7\xa0grandiflorum /Ramat./Kitam.

Cryopreservation is widely applied to many economically important species excluding chimera plants which are problematic for long-term conservation. Their storage problems can be circumvented only by cryopreserving meristems. This study looked at the morphogenetic response of shoot tips of periclinal chimera chrysanthemum ‘Lady Orange’ and ‘Lady Salmon’, as well as the solid mutant ‘Richmond’, that were cryopreserved by encapsulation-dehydration technique. By applying 10 µM ABA in the preculture medium followed by 4-day-long dehydration treatment, the explant survival reached up to 67%. Besides the stimulation of typical single shoot recovery, cryopreservation led to direct or indirect multiple shoot formation, shoot malformation, as well as inhibited their spontaneous rooting. Microscopic analysis revealed three types of structural damages of shoot tips which can correspond with their morphogenetic response in recovery culture. No influence of cryostorage on the acclimatisation efficiency of the recovered chrysanthemums was observed.

Exploring the role of Inositol 1,3,4-trisphosphate 5/6 kinase-2 (GmITPK2) as a dehydration and salinity stress regulator in Glycine max (L.) Merr. through heterologous expression in E. coli

Phytic acid (PA) is implicative in a spectrum of biochemical and physiological processes involved in plant stress response. Inositol 1,3,4, Tris phosphate 5/6 kinase (ITPK), a polyphosphate kinase that converts Inositol 1,3,4 trisphosphate to Inositol 1,3,4,5/6 tetra phosphate, averting the inositol phosphate pool towards PA biosynthesis, is a key regulator that exists in four different isoforms in soybean. In the present study, in-silico analysis of the promoter region of ITPKs was done and among the four isoforms, promoter region of GmITPK2 showed the presence of two MYB binding elements for drought inducibility and one for ABA response. Expression profiling through qRT-PCR under drought and salinity stress showed higher expression of GmITPK2 isoform compared to the other members of the family. The study revealed GmITPK2 as an early dehydration responsive gene which is also induced by dehydration and exogenous treatment with ABA. To evaluate the osmo-protective role of GmITPK2, attempts were made to assess the bacterial growth on Luria Broth media containing 200 mM NaCl, 16% PEG and 100 μM ABA, individually. The transformed E. coli BL21 (DE3) cells harbouring the GmITPK2 gene depicted better growth on the media compared to the bacterial cells containing the vector alone. Similarly, the growth of the transformed cells in the liquid media containing 200 mM NaCl, 16% PEG and 100 μM ABA showed higher absorbance at 600 nm compared to control, at different time intervals. The GmITPK2 recombinant E. coli cells showing tolerance to drought and salinity thus demonstrated the functional redundancy of the gene across taxa. The purity and specificity of the recombinant protein was assessed and confirmed through PAGE showing a band of ∼35 kDa on western blotting using Anti- Penta His- HRP conjugate antibody. To the best of our knowledge, the present study is the first report exemplifying the role of GmITPK2 isoform in drought and salinity tolerance in soybean.

OsCBSCBSPB4 is a Two Cystathionine-β-Synthase Domain-containing Protein from Rice that Functions in Abiotic Stress Tolerance.

Cystathionine β-synthase (CBS) domains have been identified in a wide range of proteins of unrelated functions such as, metabolic enzymes, kinases and channels, and usually occur as tandem re-peats, often in combination with other domains. In plants, CBS Domain-Containing Proteins (CDCPs) form a multi-gene family and only a few are so far been reported to have a role in development via regu-lation of thioredoxin system as well as in abiotic and biotic stress response. However, the function of majority of CDCPs still remains to be elucidated in plants. Here, we report the cloning, characterization and functional validation of a CBS domain containing protein, OsCBSCBSPB4 from rice, which pos-sesses two CBS domains and one PB1 domain. We show that OsCBSCBSPB4 encodes a nucleo-cytoplasmic protein whose expression is induced in response to various abiotic stress conditions in salt-sensitive IR64 and salt-tolerant Pokkali rice cultivars. Further, heterologous expression of OsCBSCB-SPB4 in E. coli and tobacco confers marked tolerance against various abiotic stresses. Transgenic tobac-co seedlings over-expressing OsCBSCBSPB4 were found to exhibit better growth in terms of delayed leaf senescence, profuse root growth and increased biomass in contrast to the wild-type seedlings when subjected to salinity, dehydration, oxidative and extreme temperature treatments. Yeast-two hybrid stud-ies revealed that OsCBSCBSPB4 interacts with various proteins. Of these, some are known to be in-volved in abiotic stress tolerance. Our results suggest that OsCBSCBSPB4 is involved in abiotic stress response and is a potential candidate for raising multiple abiotic stress tolerant plants.

Ectopic Expression of Pumpkin NAC Transcription Factor CmNAC1 Improves Multiple Abiotic Stress Tolerance in Arabidopsis.

Drought, cold and salinity are the major environmental stresses that limit agricultural productivity. NAC transcription factors regulate the stress response in plants. Pumpkin (Cucurbita moschata) is an important cucurbit vegetable crop and it has strong resistance to abiotic stress; however, the biological functions of stress-related NAC genes in this crop are largely unknown. This study reports the function of CmNAC1, a stress-responsive pumpkin NAC domain protein. The CmNAC1-GFP fusion protein was transiently expressed in tobacco leaves for subcellular localization analysis, and we found that CmNAC1 is localized in the nucleus. Transactivation assay in yeast cells revealed that CmNAC1 functions as a transcription activator, and its transactivation domain is located in the C-terminus. CmNAC1 was ubiquitously expressed in different organs, and its transcript was induced by salinity, cold, dehydration, H2O2, and abscisic acid (ABA) treatment. Furthermore, the ectopic expression (EE) of CmNAC1 in Arabidopsis led to ABA hypersensitivity and enhanced tolerance to salinity, drought and cold stress. In addition, five ABA-responsive elements were enriched in CmNAC1 promoter. The CmNAC1-EE plants exhibited different root architecture, leaf morphology, and significantly high concentration of ABA compared with WT Arabidopsis under normal conditions. Our results indicated that CmNAC1 is a critical factor in ABA signaling pathways and it can be utilized in transgenic breeding to improve the abiotic stress tolerance of crops.