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Adverse environmental stresses are known to affect plant growth and development, thus reducing crop productivity. Abscisic acid-, stress -, and ripening-induced (ASR) proteins play important roles in protecting plants from adverse environmental conditions. However, the biological functions of ASR proteins in Brachypodium remain unclear. We characterized the Brachypodium BdASR2 gene and concluded that it plays a positive role in salt stress tolerance. Overexpressing BdASR2 plants showed enhanced salt tolerance and decreased ion leakage and MDA content as compared to WT plants under salt stress conditions. Moreover, the roots were longer in overexpressing BdASR2 plants than WT plants under salt stress. The overexpression of BdASR2 increased antioxidant enzyme activity and upregulated the expression of stress-related genes under salt stress conditions. Taken together, these results indicate that BdASR2 plays important roles in salt stress responses by controlling ROS homeostasis and regulating the expression of stress-related genes.

Background Flavonoids can protect plants against extreme temperatures and ROS due to their antioxidant activities. We found that deep-purple seed coat color was controlled by two gene interaction (12:3:1) from the cross between yellow and deep-purple seed coat colored inbreds. F2:3 seeds were grouped in 3 by seed coat color and germinated under chilling (4 °C) and non-acclimated conditions (18 °C) for a week, followed by normal conditions (18 °C) for three weeks and a subsequent chilling stress (4 °C) induction. We analyzed mean daily germination in each group. Additionally, to study the acclimation in relationship to the different seed coat colors on the germination ability and seedling performances under the cold temperatures, we measured the chlorophyll content, ROS scavenging activity, and expression levels of genes involved in ROS scavenging, flavonoid biosynthetic pathway, and cold response in seedlings. Results The results of seed color segregation between yellow and deep purple suggested a two-gene model. In the germination study, normal environmental conditions induced the germination of yellow-seed, while under chilling conditions, the germination ratio of deep purple-seed was higher than that of yellow-colored seeds. We also found that the darker seed coat colors were highly responsive to cold acclimation based on the ROS scavenging enzymes activity and gene expression of ROS scavenging enzymes, flavonoid biosynthetic pathway and cold responsive genes. Conclusions We suggest that deep purple colored seed might be in a state of innate pre-acquired stress response state under normal conditions to counteract stresses in a more effective way. Whereas, after the acclimation, another stress should enhance the cold genes expression response, which might result in a more efficient chilling stress response in deep purple seed seedlings. Low temperature has a large impact on the yield of crops. Thus, understanding the benefit of seed coat color response to chilling stress and the identification of limiting factors are useful for developing breeding strategies in order to improve the yield of wheat under chilling stress.

Abscisic acid-, stress-, and ripening-induced (ASR) proteins play an important role in protecting plants against adverse environmental conditions. Here, we identified 24 ASR genes in the wheat genome and analyzed their characteristics. Among these, five ASR genes highly induced by abscisic acid (ABA) and polyethylene glycol were cloned and further characterized. The TaASR genes were expressed in response to different abiotic stresses and ABA and were found to be localized in the nucleus and plasma membrane of transformed tobacco cells. Brachypodium distachyon transgenic plants overexpressing TaASR2D showed enhanced drought tolerance by regulating leaf transpiration. The expression levels of stress-related and ABA-responsive genes were higher in transgenic plants than in wild-type plants under drought stress conditions. Moreover, overexpression of TaASR2D increased the levels of both endogenous ABA and hydrogen peroxide in response to drought stress, and these plants showed hypersensitivity to exogenous ABA at the germination stage. Furthermore, plants overexpressing TaASR2D showed increased stomatal closure. Further analysis revealed that TaASR2D interacts with ABA biosynthesis and stress-related proteins in yeast and tobacco plants. Collectively, these findings indicate that TaASR2D plays an important role in the response of plants to drought stress by regulating the ABA biosynthesis pathway and redox homeostasis system.

In wheat (Triticum aestivum L.), the floret development stage is an important step in determining grain yield per spike; however, the molecular mechanisms underlying floret development remain unclear. In this study, we elucidated the role of TaF-box2, a member of the F-box-containing E3 ubiquitin protein ligases, which is involved in floret development and anthesis of wheat. TaF-box2 was transiently expressed in the plasma membrane and cytoplasm of both tobacco and wheat. We also found that the SCF

Abscisic acid-, stress-, ripening-induced (ASR) proteins are some of the most important small proteins involved in plant responses to abiotic stresses and hormone signals. Recently, BdASR1 was revealed to be upregulated in response to abiotic stresses and hormone treatments and regulate expression of stress-related genes and drought tolerance in tobacco plants. However, the biological and molecular functions of BdASR1 remain to be elucidated. Here, we isolated and characterized BdASR1-interacting protein using the yeast two-hybrid assay. The expression of the interaction protein, BdERF96, increased under drought and oxidative stress corresponding to the expression of BdASR1. Subcellular localization of BdERF96 was detected in the plasma membrane and nucleus. The interaction of BdASR1 and BdERF96 at the plasma membrane and nucleus was demonstrated using bimolecular fluorescence complementation analysis. The findings imply that BdERF96 in association with BdASR1 could play a role in the positive response to drought and oxidative stresses.

Plants accumulate key metabolites as a response of biotic/abiotic stress conditions. In seed coats, anthocyanins, carotenoids, and chlorophylls can be found. They have been associated as important antioxidants that affect germination. In wheat, anthocyanins can impart the seed coat color which have been recognized as health-promoting nutrients. Transcription factors act as master regulators of cellular processes. Transcription complexes such as MYB-bHLH-WD40 (MBW) regulate the expression of multiple target genes in various plant species. In this study, the spatiotemporal accumulation of seed coat pigments in different developmental stages (10, 20, 30, and 40 days after pollination) was analyzed using cryo-cuts. Moreover, the accumulation of phenolic, anthocyanin, and chlorophyll contents was quantified, and the expression of flavonoid biosynthetic genes was evaluated. Finally, transcriptome analysis was performed to analyze putative MYB genes related to seed coat color, followed by further characterization of putative genes. TaTCL2, an MYB gene, was cloned and sequenced. It was determined that TaTCL2 contains a SANT domain, which is often present in proteins participating in the response to anthocyanin accumulation. Moreover, TaTCL2 transcript levels were shown to be influenced by anthocyanin accumulation during grain development. Interaction network analysis showed interactions with GL2 (HD-ZIP IV), EGL3 (bHLH), and TTG1 (WD40). The findings of this study elucidate the mechanisms underlying color formation in Triticum aestivum L. seed coats.

The effects of drought acclimation on plant growth and water relations, membrane status, photosynthetic activity, proline content, membrane lipid peroxidation, enzyme antioxidant activity, and drought-responsive gene expression patterns were investigated under progressive drought conditions with drought acclimation (DA) and non-acclimation (NA) treatments in four wheat genotypes. Initial water stressapplied at 10 days after seedling transfer induced acclimation to subsequent water stress (S2), following re-watering. A reversible decline in plant growth and water relations, membrane stability, and photosynthetic activity resulted in increased lipid peroxidation, reactive oxygen species, membrane injury, enzyme antioxidant activities, and H2O2 and osmolyte accumulation in DA plants. TaWRKY2, TaNAC1, TaMYB2, TabHLH1, and TabZIP1 levels were upregulated to a greater extent in NA than in DA plants, during S2. The results suggest wheat plants employ physiological, biochemical and molecular mechanism under drought improving their ability to survive subsequent water stress in the later period of growth and development.

BdASR4 expression was up-regulated during abiotic stress and hormone treatments. Plants over-expressing BdASR4 improved drought tolerant. BdASR4 may regulate antioxidant activities and transcript levels of stress-related and abscisic acid-responsive genes. Abiotic stress conditions negatively affect plant growth and developmental processes, causing a reduction in crop productivity. The abscisic acid-, stress-, ripening-induced (ASR) proteins play important roles in the protection of plants from abiotic stress. Brachypodium distachyon L. is a well-studied monocot model plant. However, ASR proteins of Brachypodium have not been widely studied. In this study, five ASR genes of Brachypodium plant were cloned and characterized. The BdASR genes were expressed in response to various abiotic stresses and hormones. In particular, BdASR4 was shown to encode a protein containing a nuclear localization signal in its C-terminal region, which enabled protein localization in the nucleus. To further examine functions of BdASR4, transgenic Brachypodium plants harboring BdASR4 were generated. Over-expression of BdASR4 was associated with strong drought tolerance, and plants over-expressing BdASR4 preserved more water and displayed higher antioxidant enzyme activities than did the wild-type plants. The transcript levels of stress-responsive genes, reactive oxygen species scavenger-associated genes, and abscisic acid-responsive genes tended to be higher in transgenic plants than in WT plants. Moreover, plants over-expressing BdASR4 were hypersensitive to exogenous abscisic acid at the germination stage. Taken together, these findings suggest multiple roles for BdASR4 in the plant response to drought stress by regulating antioxidant enzymes and the transcription of stress- and abscisic acid-responsive genes.

Durum wheat (Triticum turgidum ssp. durum) is one of the main species of cultivated wheat. In arid and semi-arid areas, salinity stress reduces durum wheat productivity. This study used 26 durum wheat accessions from semi-arid regions in Tunisia to analyze plant tolerance to salt stress. Salt stress was experimentally applied by regularly submerging pots in NaCl solution. The salt tolerance trait index (STTI) and salt tolerance index (STI) of various growth parameters were used as criteria to select for salt tolerance. Analysis of genetic relationships was carried out to determine the genetic distance between durum wheat accessions. Based on simple sequence repeats analysis, a molecular marker for salt stress resistance in durum wheat was developed. Salt-treated plants had reduced morphological traits compared to control plants. Most STTIs in all genotypes were below 100 %. Based on STI, 8 accessions were found to be salt-resistant, 16 were salt-moderate, two were salt-susceptible. Analysis of the genetic relationships among 28 Tunisian durum wheat accessions revealed that landraces of the same nominal type are closely related. Of the 94 SSR primers investigated, three were selected and used to design sequence characterized amplified region (SCAR) primers. One SCAR primer pair, KUCMB_Xgwm403_2, produced a 207 bp band that was present in salt-resistant durum wheat lines but absent in salt-susceptible lines. The results suggest that KUCMB_Xgwm403_2 could be a potential genetic tag for salt-tolerant durum wheats.

Gibberellic acid (GA) is involved in the regulation of plant growth and development. We defined GA-stimulated transcript (GAST) gene family and characterized its four members (TaGAST1, 2, 3, and 4) in wheat spikes. Triticum aestivum whole spikes were collected at ten developmental stages and dehulled spikelets were obtained at various days after flowering. Expression of TaGAST1, 2, 3, and 4 was analyzed using RT-PCR at inflorescence development stages, in different tissues, and after phytohormones application. To identify proteins interacting with TaGAST1, yeast two-hybridization was performed and BiFC analysis was used for verification. TaGAST1 was expressed at the inflorescence stage and only expressed in seedlings under abscisic acid (ABA) treatment after phytohormone treatment. TaGAST2 and TaGAST3 showed moderate expression in the spike, vigorous transcript accumulation in the seedling, and up-regulation by exogenous GA in early germination stages. TaGAST4 was predominantly expressed in the seedling. Wheat cyclophilin A-1 (TaCypA1), identified as a TaGAST1-interacting protein, showed opposite expression pattern in the developing spike to TaGAST1. TaCypA1 transcript was slightly up-regulated by GA, slightly down-regulated by paclobutrazol, and was maintained after ABA treatment. The interaction of TaGAST1 with TaCypA1 is targeted to the plasma membrane. TaGAST1 was specifically expressed in the wheat spike and was stimulated by exogenous GA treatment. TaGAST2 and TaGAST3 expression in germinating seeds and seedlings was higher than that in the spike stage. TaGAST4 was not expressed in all developmental stages. TaGAST1 and TaCypA1 might be expressed antagonistically during wheat spike development.