ABRE-binding Factor Research Articles

Abscisic Acid Flux Alterations Result in Differential Abscisic Acid Signaling Responses and Impact Assimilation Efficiency in Barley under Terminal Drought Stress

Abscisic acid (ABA) is a central player in plant responses to drought stress. How variable levels of ABA under short-term versus long-term drought stress impact assimilation and growth in crops is unclear. We addressed this through comparative analysis, using two elite breeding lines of barley (Hordeum vulgare) that show senescence or stay-green phenotype under terminal drought stress and by making use of transgenic barley lines that express Arabidopsis (Arabidopsis thaliana) 9-cis-epoxycarotenoid dioxygenase (AtNCED6) coding sequence or an RNA interference (RNAi) sequence of ABA 8'-hydroxylase under the control of a drought-inducible barley promoter. The high levels of ABA and its catabolites in the senescing breeding line under long-term stress were detrimental for assimilate productivity, whereas these levels were not perturbed in the stay-green type that performed better. In transgenic barley, drought-inducible AtNCED expression afforded temporal control in ABA levels such that the ABA levels rose sooner than in wild-type plants but also subsided, unlike as in the wild type , to near-basal levels upon prolonged stress treatment due to down-regulation of endogenous HvNCED genes. Suppressing of ABA catabolism with the RNA interference approach of ABA 8'-hydroxylase caused ABA flux during the entire period of stress. These transgenic plants performed better than the wild type under stress to maintain a favorable instantaneous water use efficiency and better assimilation. Gene expression analysis, protein structural modeling, and protein-protein interaction analyses of the members of the PYRABACTIN RESISTANCE1/PYRABACTIN RESISTANCE1-LIKE/REGULATORY COMPONENT OF ABA RECEPTORS, TYPE 2C PROTEIN PHOSPHATASE Sucrose non-fermenting1-related protein kinase2, and ABA-INSENSITIVE5/ABA-responsive element binding factor family identified specific members that could potentially impact ABA metabolism and stress adaptation in barley.

OsSLI1, a homeodomain containing transcription activator, involves abscisic acid related stress response in rice (Oryza sativa L.).

Homeodomain-leucine zipper type I (HD-Zip I) proteins are involved in the regulation of plant development and response to environmental stresses. In this study, OsSLI1 (Oryza sativa stress largely induced 1), encoding a member of the HD-Zip I subfamily, was isolated from rice. The expression of OsSLI1 was dramatically induced by multiple abiotic stresses and exogenous abscisic acid (ABA). In silico sequence analysis discovered several cis-acting elements including multiple ABREs (ABA-responsive element binding factors) in the upstream promoter region of OsSLI1. The OsSLI1-GFP fusion protein was localized in the nucleus of rice protoplast cells and the transcriptional activity of OsSLI1 was confirmed by the yeast hybrid system. Further, it was found that OsSLI1 expression was enhanced in an ABI5-Like1 (ABL1) deficiency rice mutant abl1 under stress conditions, suggesting that ABL1 probably negatively regulates OsSLI1 gene expression. Moreover, it was found that OsSLI1 was regulated in panicle development. Taken together, OsSLI1 may be a transcriptional activator regulating stress-responsive gene expression and panicle development in rice.

The tumor necrosis factor receptor‐associated factor (TRAF)‐like family protein SEVEN IN ABSENTIA 2 (SINA2) promotes drought tolerance in an ABA‐dependent manner in Arabidopsis

Tumor necrosis factor receptor-associated factor (TRAF) proteins play crucial roles in plant development and response to abiotic stress. Here, we present genetic evidence that SEVEN IN ABSENTIA 2 (SINA2), a TRAF-like family protein, is involved in abscisic acid (ABA)-related drought stress signaling in Arabidopsis. Gene expression, protein subcellular localization, protein-protein interaction, and a transient transcription dual-luciferase assay were performed. The drought tolerance of SINA2 loss-of-function mutants and SINA2-overexpressing plants was investigated. In Arabidopsis, SINA2 was significantly induced by ABA and drought treatment. The SINA2-YFP fusion protein was predominately localized in the nuclei and cytoplasm. Loss of function of SINA2 (sina2) reduced drought tolerance, whereas overexpression of SINA2 increased stomatal closure, decreased water loss, and therefore improved drought resistance in transgenic plants. Upon ABA treatment, expression of some key ABA- and stress-responsive genes decreased in the sina2 mutant, but increased in SINA2-overexpressing plants. Furthermore, SINA2 was induced in the ABA-deficient mutant by ABA, but not by drought stress. Thus, the drought response of SINA2 was ABA-dependent. ProSINA2::LUC expression in Arabidopsis protoplasts further revealed that ABA-responsive element (ABRE) binding (AREB) protein 1 (AREB1) AREB2 and ABRE-binding factor 3 (ABF3) might regulate SINA2 expression at the transcriptional level. Our results indicate that SINA2 functions as a positive molecular link between drought tolerance and ABA signaling in Arabidopsis.

Heterologous expression of Arabidopsis ABF4 gene in potato enhances tuberization through ABA-GA crosstalk regulation

Potato (Solanum tuberosum L.) tuberization is regulated by many signals, such as abscisic acid (ABA), sucrose and gibberellic acid (GA). ABA and sucrose are positive modulators, while GA is an inhibitor of the process. ABF (ABRE-binding factor) proteins are transcription factors involved in ABA and stress signaling. Previously, we reported that S. tuberosum StABF1 could mediate the ABA effects on tuberization. The aim of the present study was to evaluate the potential use of ABF genes to enhance tuberization and to determine the molecular mechanism involved. For this purpose, transgenic potato plants expressing the Arabidopsis ABF4 or ABF2 genes were generated, and their tuberization capacity and response to tuberization-related signals were analyzed in vitro. The results indicate that both ABF4 and ABF2 proteins positively regulate potato tuber induction; however, only ABF4 expression significantly increases the number and weight of the tubers obtained, without stunting growth. ABF4 and ABF2 transgenic plants exhibit ABA hypersensitivity during tuberization, accompanied by a GA-deficient phenotype. ABF4 expression triggers a significant rise in ABA levels in stolons under tuber-inducing conditions as compared with wild-type plants and a transcriptional deregulation of GA metabolism genes. Our results demonstrate that Arabidopsis ABF4 functions in potato ABA-GA signaling crosstalk during tuberization by regulating the expression of ABA- and GA-metabolism genes. ABF4 gene might be a potential tool to increase tuber production, since its heterologous expression in potato enhances tuber induction without affecting plant growth.

A WRKY gene from Tamarix hispida, ThWRKY4, mediates abiotic stress responses by modulating reactive oxygen species and expression of stress-responsive genes

WRKY transcription factors are involved in various biological processes, such as development, metabolism and responses to stress. However, their exact roles in abiotic stress tolerance are largely unknown. Here, we demonstrated a working model for the function of a WRKY gene (ThWRKY4) from Tamarix hispida in the stress response. ThWRKY4 is highly induced by abscisic acid (ABA), salt and drought in the early period of stress (stress for 3, 6, or 9 h), which can be regulated by ABF (ABRE binding factors) and Dof (DNA binding with one finger), and also can be crossregulated by other WRKYs and autoregulated as well. Overexpression of ThWRKY4 conferred tolerance to salt, oxidative and ABA treatment in transgenic plants. ThWRKY4 can improve the tolerance to salt and ABA treatment by improving activities of superoxide dismutase and peroxidase, decreasing levels of O2 (-) and H2O2, reducing electrolyte leakage, keeping the loss of chlorophyll, and protecting cells from death. Microarray analyses showed that overexpression of ThWRKY4 in Arabidopsis leads to 165 and 100 genes significantly up- and downregulated, respectively. Promoter scanning analysis revealed that ThWRKY4 regulates the gene expression via binding to W-box motifs present in their promoter regions. This study shows that ThWRKY4 functions as a transcription factor to positively modulate abiotic stress tolerances, and is involved in modulating reactive oxygen species.

ABA signaling in stress-response and seed development

KEY MESSAGE : We review the recent progress on ABA signaling, especially ABA signaling for ABA-dependent gene expression, including the AREB/ABF regulon, SnRK2 protein kinase, 2C-type protein phosphatases and ABA receptors. Drought negatively impacts plant growth and the productivity of crops. Drought causes osmotic stress to organisms, and the osmotic stress causes dehydration in plant cells. Abscisic acid (ABA) is produced under osmotic stress conditions, and it plays an important role in the stress response and tolerance of plants. ABA regulates many genes under osmotic stress conditions. It also regulates gene expression during seed development and germination. The ABA-responsive element (ABRE) is the major cis-element for ABA-responsive gene expression. ABRE-binding protein (AREB)/ABRE-binding factor (ABF) transcription factors (TFs) regulate ABRE-dependent gene expression. Other TFs are also involved in ABA-responsive gene expression. SNF1-related protein kinases 2 are the key regulators of ABA signaling including the AREB/ABF regulon. Recently, ABA receptors and group A 2C-type protein phosphatases were shown to govern the ABA signaling pathway. Moreover, recent studies have suggested that there are interactions between the major ABA signaling pathway and other signaling factors in stress-response and seed development. The control of the expression of ABA signaling factors may improve tolerance to environmental stresses.

The inhibitory effect of ABA on floral transition is mediated by ABI5 in Arabidopsis

Seed germination and flowering initiation are both transitions responding to similar seasonal cues. This study shows that ABSCISIC ACID-INSENSITIVE MUTANT 5 (ABI5), a bZIP transcription factor, which plays an important role in the abscisic acid (ABA)-arrested seed germination, is robustly associated with the floral transition in Arabidopsis. Under long-day conditions, overexpression of ABI5 could delay floral transition through upregulating FLOWERING LOCUS C (FLC) expression. In contrast, ectopically overexpressing FLC in an abi5 mutant reversed the earlier flowering phenotype. Further analysis indicated that transactivation of FLC could be promoted by ABI5 and/or other abscisic acid-responsive element (ABRE)-binding factors (ABFs). The expression of FLC that was promoted by ABI5 and/or other ABFs could be blocked in a triple SNF1-related protein kinase (SnRK) mutant, snrk2.2/2.3/2.6, despite the presence of ABA. In sharp contrast, when SnRK2.6 was coexpressed, the reduction of transactivity of FLC was reverted in mesophyll protoplasts of snrk2.2/2.3/2.6. Additional results from analysing transgenic plants carrying mutations of phosphoamino acids (ABI5 S42AS145AT201A), which are conserved in ABI5, suggested that SnRK2-mediated ABI5 and/or ABF phosphorylation may be crucial for promoting FLC expression. The transgenic plants ABI5 S42AS145AT201A were insensitive to ABA in seed germination, in addition to having an earlier flowering phenotype. Direct binding of ABI5 to the ABRE/G-box promoter elements existing in FLC was demonstrated by chromatin immunoprecipitation. Mutations at the ABRE/G-box regions in FLC promoter sequences abolished the ABI5-promoted transactivation of FLC. In summary, these results may decipher the inhibitory effect of ABA on floral transition in Arabidopsis.

Overexpression of Arabidopsis ABF3 gene confers enhanced tolerance to drought and heat stress in creeping bentgrass

The Arabidopsis, abscisic acid responsive element-binding factor 3, ABF3 is known to play an important role in stress responses via regulating the expression of stress-responsive genes. In this study, we introduced pCAMBIA3301 vector harboring the ABF3 gene into creeping bentgrass (Agrostis stolonifera) through Agrobacterium-mediated transformation in order to develop a stress-tolerant variety of turfgrass. After transformation, putative transgenic plants were selected using the herbicide resistance assay. Genomic integration of the transgene was confirmed by genomic PCR and Southern blot analysis, and gene expression was validated by northern blot analysis. Under drought-stressed condition, the transgenic plants overexpressing ABF3 displayed significantly enhanced drought tolerance with higher water content and slower water loss rate than the control plants. Furthermore, the stomata of the ABF3 transgenic plants closed more than those of wild-type creeping bentgrass plants, under both non-stressed and ABA treatment conditions. In addition, the transgenic plants showed enhanced tolerance to heat stress. These results suggest that the overexpression of the ABF3 gene in creeping bentgrass might enhance survival in water-limiting and high temperature environments through increased stomatal closure and reduced water losses.

Isolation and characterization of two ABRE-binding proteins: EABF and EABF1 from the oil palm

Abscisic acid (ABA) is an important phytohormone involved in the abiotic stress resistance in plants. The ABA-responsive element (ABRE) binding factors play significant roles in the plant development and response to abiotic stresses, but none so far have been isolated and characterized from the oil palm. Two ABA-responsive cDNA clones, named EABF and EABF1, were isolated from the oil palm fruits using yeast one-hybrid system. The EABF had a conserved AP2/EREBP DNA-binding domain (DNA-BD) and a potential nuclear localization sequence (NLS). No previously known DNA-BD was identified from the EABF1 sequence. The EABF and EABF1 proteins were classified as DREB/CBF and bZIP family members based on the multiple sequence alignment and phylogenetic analysis. Both proteins showed ABRE-binding and transcriptional activation properties in yeast. Furthermore, both proteins were able to trans-activate the down-stream expression of the LacZ reporter gene in yeast. An electrophoretic mobility shift assay revealed that in addition to the ABRE sequence, both proteins could bind to the DRE sequence as well. Transcriptional analysis revealed that the expression of EABF was induced in response to the ABA in the oil palm fruits and leaves, but not in roots, while the EABF1 was constitutively induced in all tissues. The expressions of both genes were strongly induced in fruits in response to the ABA, ethylene, methyl jasmonate, drought, cold and high-salinity treatments, indicating that the EABF and EABF1 might act as connectors among different stress signal transduction pathways. Our results indicate that the EABF and EABF1 are novel stress-responsive transcription factors, which are involved in the abiotic stress response and ABA signaling in the oil palm and could be used for production of stress-tolerant transgenic crops.

Delta subclass HD-Zip proteins and a B-3 AP2/ERF transcription factor interact with promoter elements required for expression of the Arabidopsis cytochrome c oxidase 5b-1 gene

We have identified transcription factors that interact with a promoter region involved in expression of the Arabidopsis thaliana COX5b-1 gene, which encodes an isoform of the cytochrome c oxidase zinc binding subunit. Elements with the core sequence ATCATT, involved in induction by sugars, are recognized both in vitro and in one-hybrid assays in yeast by HD-Zip proteins from the delta subclass and, though less efficiently, by the trihelix transcription factor GT-3b. DistalB-like elements (CCACTTG), required for induction by abscisic acid (ABA), interact with ESE1, a member of the B-3 subgroup of AP2/ERF transcription factors. The HD-Zip protein Athb-21 and ESE1 are able to interact in yeast two-hybrid assays with the ABA responsive element binding factor AREB2/ABF4, which binds to a G-box absolutely required for expression of the COX5b-1 gene. Overexpression of the identified transcription factors in plants produces an increase in COX5b-1 transcript levels. Moreover, these factors are able to induce the expression of a reporter gene located in plants under the control of the relevant COX5b-1 promoter regions required for expression. Analysis of promoter regions of COX5b genes from different plant species suggests that the identified transcription factors were recruited for the regulation of COX5b gene expression at different stages during the evolution of dicot plants.

Constitutive Activation of Transcription Factor OsbZIP46 Improves Drought Tolerance in Rice

OsbZIP46 is one member of the third subfamily of bZIP transcription factors in rice (Oryza sativa). It has high sequence similarity to ABA-responsive element binding factor (ABF/AREB) transcription factors ABI5 and OsbZIP23, two transcriptional activators positively regulating stress tolerance in Arabidopsis (Arabidopsis thaliana) and rice, respectively. Expression of OsbZIP46 was strongly induced by drought, heat, hydrogen peroxide, and abscisic acid (ABA) treatment; however, it was not induced by salt and cold stresses. Overexpression of the native OsbZIP46 gene increased ABA sensitivity but had no positive effect on drought resistance. The activation domain of OsbZIP46 was defined by a series of deletions, and a region (domain D) was identified as having a negative effect on the activation. We produced a constitutive active form of OsbZIP46 (OsbZIP46CA1) with a deletion of domain D. Overexpression of OsbZIP46CA1 in rice significantly increased tolerance to drought and osmotic stresses. Gene chip analysis of the two overexpressors (native OsbZIP46 and the constitutive active form OsbZIP46CA1) revealed that a large number of stress-related genes, many of them predicted to be downstream genes of ABF/AREBs, were activated in the OsbZIP46CA1 overexpressor but not (even down-regulated) in the OsbZIP46 overexpressor. OsbZIP46 can interact with homologs of SnRK2 protein kinases that phosphorylate ABFs in Arabidopsis. These results suggest that OsbZIP46 is a positive regulator of ABA signaling and drought stress tolerance of rice depending on its activation. The stress-related genes activated by OsbZIP46CA1 are largely different from those activated by the other rice ABF/AREB homologs (such as OsbZIP23), further implying the value of OsbZIP46CA1 in genetic engineering of drought tolerance.

An ABRE Promoter Sequence is Involved in Osmotic Stress-Responsive Expression of the DREB2A Gene, Which Encodes a Transcription Factor Regulating Drought-Inducible Genes in Arabidopsis

In plants, osmotic stress-responsive transcriptional regulation depends mainly on two major classes of cis-acting elements found in the promoter regions of stress-inducible genes: ABA-responsive elements (ABREs) and dehydration-responsive elements (DREs). ABRE has been shown to perceive ABA-mediated osmotic stress signals, whereas DRE is known to be involved in an ABA-independent pathway. Previously, we reported that the transcription factor DRE-BINDING PROTEIN 2A (DREB2A) regulates DRE-mediated transcription of target genes under osmotic stress conditions in Arabidopsis (Arabidopsis thaliana). However, the transcriptional regulation of DREB2A itself remains largely uncharacterized. To elucidate the transcriptional mechanism associated with the DREB2A gene under osmotic stress conditions, we generated a series of truncated and base-substituted variants of the DREB2A promoter and evaluated their transcriptional activities individually. We found that both ABRE and coupling element 3 (CE3)-like sequences located approximately -100 bp from the transcriptional initiation site are necessary for the dehydration-responsive expression of DREB2A. Coupling our transient expression analyses with yeast one-hybrid and chromatin immunoprecipitation (ChIP) assays indicated that the ABRE-BINDING PROTEIN 1 (AREB1), AREB2 and ABRE-BINDING FACTOR 3 (ABF3) bZIP transcription factors can bind to and activate the DREB2A promoter in an ABRE-dependent manner. Exogenous ABA application induced only a modest accumulation of the DREB2A transcript when compared with the osmotic stress treatment. However, the osmotic stress-induced DREB2A expression was found to be markedly impaired in several ABA-deficient and ABA-insensitive mutants. These results suggest that in addition to an ABA-independent pathway, the ABA-dependent pathway plays a positive role in the osmotic stress-responsive expression of DREB2A.

Characterization of potential ABA receptors in Vitis vinifera

Molecular control mechanisms for abiotic stress tolerance are based on the activation and regulation of specific stress-related genes. The phytohormone abscisic acid (ABA) is a key endogenous messenger in a plant's response to such stresses. A novel ABA binding mechanism which plays a key role in plant cell signaling cascades has recently been uncovered. In the absence of ABA, a type 2C protein phosphatase (PP2C) interacts and inhibits the kinase SnRK2. Binding of ABA to the PYR/PYLs receptors enables interaction between the ABA receptor and the PP2C protein, and abrogates the SnRK2 inactivation. The active SnRK2 is then free to activate the ABA-responsive element Binding Factors which target ABA-dependent gene expression. We used the grape as a model to study the ABA perception mechanism in fruit trees. The grape ABA signaling cascade consists of at least seven ABA receptors and six PP2Cs. We used a yeast two-hybrid system to examine physical interaction in vitro between the grape ABA receptors and their interacting partners, and found that twenty-two receptor-PP2C interactions can occur. Moreover, quantifying these affinities by the use of the LacZ reporter enables us to show that VvPP2C4 and VvPP2C9 are the major binding partners of the ABA receptor. We also tested in vivo the root and leaf gene expression of the various ABA receptors and PP2Cs in the presence of exogenic ABA and under different abiotic stresses such as high salt concentration, cold and drought, and found that many of these genes are regulated by such abiotic environmental factors. Our results indicate organ specificity in the ABA receptor genes and stress specificity in the VvPP2Cs. We suggest that VvPP2C4 is the major PP2C involved in ABA perception in leaves and roots, and VvRCAR6 and VvRCAR5 respectively, are the major receptors involved in ABA perception in these organs. Identification, characterization and manipulation of the central players in the ABA signaling cascades in fruit trees is likely to prove essential for improving their performance in the future.

Phosphorylation-mediated regulation of a rice ABA responsive element binding factor

OREB1 is a rice ABRE binding factor characterized by the presence of multiple highly-conserved phosphorylation domains (C1, C2, C3, and C4) and two kinase recognition motifs, RXXS/T and S/TXXE/D, within different functional domains. An in vitro kinase assay showed that OREB1 is phosphorylated not only by the SnRK2 kinase, but also by other Ser/Thr protein kinases, such as CaMKII, CKII, and SnRK3. Furthermore, the N-terminal phosphorylation domain C1 was found to be differentially phosphorylated by the SnRK2/SnRK3 kinase and by hyperosmotic/cold stress, suggesting that the C1 domain may function in decoding different signals. The phosphorylation-mediated regulation of OREB1 activity was investigated through mutation of the SnRK2 recognition motif RXXS/T within each phosphorylation module. OREB1 contains a crucial nine-amino acid transactivation domain located near the phosphorylation module C1. Deletion of the C1 domain increased OREB1 activity, whereas mutation of Ser 44, Ser 45, and Ser 48 of the C1 domain to aspartates decreased OREB1 activity. In the C2 domain, a double mutation of Ser 118 and Ser 120 to alanines suppressed OREB1 activity. These findings strongly suggest that selective phosphorylation of the C1 or C2 modules may positively or negatively regulate OREB1 transactivation. In addition, mutation of Ser 385 of the C4 domain to alanines completely abolished the interaction between OREB1 and a rice 14-3-3 protein, GF14d, suggesting that SnRK2-mediated phosphorylation may regulate this interaction. These results indicate that phosphorylation domains of OREB1 are not functionally redundant and regulate at least three different functions, including transactivation activity, DNA binding, and protein interactions. The multisite phosphorylation of OREB1 is likely a key for the fine control of its activity and signal integration in the complex stress signaling network of plant cells.

Overexpression of PtrABF gene, a bZIP transcription factor isolated from Poncirus trifoliata, enhances dehydration and drought tolerance in tobacco via scavenging ROS and modulating expression of stress-responsive genes

BackgroundDrought is one of the major abiotic stresses affecting plant growth, development and crop productivity. ABA responsive element binding factor (ABF) plays an important role in stress responses via regulating the expression of stress-responsive genes.ResultsIn this study, a gene coding for ABF (PtrABF) was isolated from Poncirus trifoliata (L.) Raf. PtrABF had a complete open reading frame of 1347 bp, encoding a 448 amino acid peptide, and shared high sequence identities with ABFs from other plants. PtrABF was subcellularly targeted to the nucleus, exhibited transactivation activity in yeast cell and could bind to ABRE, supporting its role as a transcription factor. Expression levels of PtrABF were induced by treatments with dehydration, low temperature and ABA. Ectopic expression of PtrABF under the control of a CaMV 35S promoter in transgenic tobacco plants enhanced tolerance to both dehydration and drought. Under dehydration and drought conditions, the transgenic plants accumulated lower levels of reactive oxygen species compared with wild type, accompanied by higher activities and expression levels of three antioxidant enzymes. In addition, steady-state mRNA levels of nine stress-responsive genes coding for either functional or regulatory proteins were induced to higher levels in the transgenic lines with or without drought stress.ConclusionsPtrABF is a bZIP transcription factor and functions in positive modulation of drought stress tolerance. It may be an important candidate gene for molecular breeding of drought- tolerant plants.

The transcription factor SlAREB1 confers drought, salt stress tolerance and regulates biotic and abiotic stress‐related genes in tomato

Members of the abscisic acid-responsive element binding protein (AREB)/abscisic acid-responsive element binding factor (ABF) subfamily of basic leucine zipper (bZIP) transcription factors have been implicated in abscisic acid (ABA) and abiotic stress responses in plants. Here we describe two members identified in cultivated tomato (Solanum lycopersicum), named SlAREB1 and SlAREB2. Expression of SlAREB1 and SlAREB2 is induced by drought and salinity in both leaves and root tissues, although that of SlAREB1 was more affected. In stress assays, SlAREB1-overexpressing transgenic tomato plants showed increased tolerance to salt and water stress compared to wild-type and SlAREB1-down-regulating transgenic plants, as assessed by physiological parameters such as relative water content (RWC), chlorophyll fluorescence and damage by lipoperoxidation. In order to identify SlAREB1 target genes responsible for the enhanced tolerance, microarray and cDNA-amplified fragment length polymorphism (AFLP) analyses were performed. Genes encoding oxidative stress-related proteins, lipid transfer proteins (LTPs), transcription regulators and late embryogenesis abundant proteins were found among the up-regulated genes in SlAREB1-overexpressing lines, especially in aerial tissue. Notably, several genes encoding defence proteins associated with responses to biotic stress (e.g. pathogenesis-related proteins, protease inhibitors, and catabolic enzymes) were also up-regulated by SlAREB1 overexpression, suggesting that this bZIP transcription factor is involved in ABA signals that participate in abiotic stress and possibly in response to pathogens.

The ABRE-binding bZIP transcription factor OsABF2 is a positive regulator of abiotic stress and ABA signaling in rice

Abscisic acid (ABA) is an important phytohormone involved in abiotic stress tolerance in plants. The group A bZIP transcription factors play important roles in the ABA signaling pathway in Arabidopsis but little is known about their functions in rice. In our current study, we have isolated and characterized a group A bZIP transcription factor in rice, OsABF2 ( Oryza sativa ABA-responsive element binding factor 2). It was found to be expressed in various tissues in rice and induced by different types of abiotic stress treatments such as drought, salinity, cold, oxidative stress, and ABA. Subcellular localization analysis in maize protoplasts using a GFP fusion vector indicated that OsABF2 is a nuclear protein. In yeast experiments, OsABF2 was shown to bind to ABA-responsive elements (ABREs) and its N-terminal region found to be necessary to transactivate a downstream reporter gene. A homozygous T-DNA insertional mutant of OsABF2 is more sensitive to salinity, drought, and oxidative stress compared with wild type plants. In addition, this Osabf2 mutant showed a significantly decreased sensitivity to high levels of ABA at germination and post-germination. Collectively, our present results indicate that OsABF2 functions as a transcriptional regulator that modulates the expression of abiotic stress-responsive genes through an ABA-dependent pathway.

TaABF1, ABA response element binding factor 1, is related to seed dormancy and ABA sensitivity in wheat ( Triticum aestivum L.) seeds

A seed-specific bZIP type transcription factor involved in abscisic acid (ABA) signal transduction of developing wheat ( Triticum aestivum L.) seeds is TaABF1, ABA response element binding factor 1. The expression of TaABF1 was examined in wheat cultivars possessing different levels of seed dormancy and ABA sensitivity at 40–50 days after pollination. Zenkoujikomugi, Norin61, and OW104 were dormant cultivars showing high sensitivity to ABA. Chihokukomugi, RL4137, and Chinese Spring, all non-dormant cultivars, show low sensitivity to ABA. Tamaizumi, AUS1408, and Kitakei-1354 represent intermediate levels of seed dormancy and sensitivity to ABA. The relation between seed dormancy and ABA sensitivity is high ( r = 0.89) in these cultivars. Relative amounts of transcripts of TaABF1 in embryos revealed high coefficients of correlation to seed dormancy ( r = 0.78) and ABA sensitivity ( r = 0.86). These results suggest that TaABF1 plays a role in the regulation of seed dormancy and ABA sensitivity in wheat.

The bZIP transcription factor OsABF1 is an ABA responsive element binding factor that enhances abiotic stress signaling in rice

A number of basic leucine zipper (bZIP) transcription factors are known to function in stress signaling in plants but few have thus far been functionally characterized in rice. In our current study in rice, we have newly isolated and characterized the OsABF1 (Oryza sativa ABA responsive element binding factor 1) gene that encodes a bZIP transcription factor. Its expression in seedling shoots and roots was found to be induced by various abiotic stress treatments such as anoxia, salinity, drought, oxidative stress, cold and abscisic acid (ABA). Subcellular localization analysis in maize protoplasts using GFP fusion vectors indicated that OsABF1 is a nuclear protein. In a yeast experiment, OsABF1 was shown to bind to ABA responsive elements (ABREs) and its N-terminal region was necessary to transactivate the downstream reporter gene. The homozygous T-DNA insertional mutants Osabf1-1 and Osabf1-2 were more sensitive in response to drought and salinity treatments than wild type plants. Furthermore, the upregulated expression of some ABA/stress-regulated genes in response to ABA treatment was suppressed in these Osabf1 mutants. Our current results thus suggest that OsABF1 is involved in abiotic stress responses and ABA signaling in rice.

Arabidopsis mutant deficient in 3 abscisic acid-activated protein kinases reveals critical roles in growth, reproduction, and stress

Abscisic acid (ABA) is an important phytohormone regulating seed dormancy, germination, seedling growth, and plant transpiration. We report here an Arabidopsis triple mutant that is disrupted in 3 SNF1-related protein kinase subfamily 2 (SnRK2s) and nearly completely insensitive to ABA. These SnRK2s, SnRK2.2, SnRK2.3, and SnRK2.6 (also known as OST1), are activated by ABA and can phosphorylate the ABA-responsive element binding factor family of b-ZIP transcription factors, which are important for the activation of ABA-responsive genes. Although stomatal regulation of snrk2.6 and seed germination and seedling growth of the snrk2.2/2.3 double mutant are insensitive to ABA, ABA responses are still present in these mutants, and the growth and reproduction of these mutants are not very different from those of the WT. In contrast, the snrk2.2/2.3/2.6 triple mutant grows poorly and produces few seeds. The triple mutant plants lose water extremely fast when ambient humidity is not high. Even on 50 muM ABA, the triple mutant can germinate and grow, whereas the most insensitive known mutants cannot develop on 10 muM ABA. In-gel kinase assays showed that all ABA-activated protein kinase activities are eliminated in the triple mutant. Also, the expression of ABA-induced genes examined is completely blocked in the triple mutant. These results demonstrate that the protein kinases SnRK2.2, SnRK2.3, and SnRK2.6 have redundant functions, and suggest that ABA signaling is critical for plant growth and reproduction.