A novel wheat α‐amylase inhibitor gene,TaHPS, significantly improves the salt and drought tolerance of transgenic Arabidopsis

On the basis of microarray analyses of the salt‐tolerant wheat mutant RH8706‐49, a previously unreported salt‐induced gene, designated as TaHPS [Triticum aestivum hypothetical (HPS)‐like protein], was cloned. Real‐time quantitative polymerase chain reaction analyses showed that expression of the gene was induced by abscisic acid, salt and drought. The encoded protein was found to be localized mainly in the plasma membranes. Transgenic Arabidopsis plants overexpressing TaHPS were more tolerant to salt and drought stresses than non‐transgenic wild‐type (WT) plants. Under salt stress, the root cells of the transgenic plants secreted more Na⁺ and guard cells took up more Ca²⁺ ions. Compared with wild‐type plants, TaHPS‐expressing transgenic plants showed significantly lower amylase activity and glucose and malic acid levels. Our results showed that the expression of TaHPS inhibited amylase activity, which subsequently led to a closure of stomatal apertures and thus improved plant tolerance to salt and drought.     

ABA Inducible Rice Protein Phosphatase 2C Confers ABA Insensitivity and Abiotic Stress Tolerance in Arabidopsis

Arabidopsis PP2C belonging to group A have been extensively worked out and known to negatively regulate ABA signaling. However, rice (Oryza sativa) orthologs of Arabidopsis group A PP2C are scarcely characterized functionally. We have identified a group A PP2C from rice (OsPP108), which is highly inducible under ABA, salt and drought stresses and localized predominantly in the nucleus. Genetic analysis revealed that Arabidopsis plants overexpressing OsPP108 are highly insensitive to ABA and tolerant to high salt and mannitol stresses during seed germination, root growth and overall seedling growth. At adult stage, OsPP108 overexpression leads to high tolerance to salt, mannitol and drought stresses with far better physiological parameters such as water loss, fresh weight, chlorophyll content and photosynthetic potential (Fv/Fm) in transgenic Arabidopsis plants. Expression profile of various stress marker genes in OsPP108 overexpressing plants revealed interplay of ABA dependent and independent pathway for abiotic stress tolerance. Overall, this study has identified a potential rice group A PP2C, which regulates ABA signaling negatively and abiotic stress signaling positively. Transgenic rice plants overexpressing this gene might provide an answer to the problem of low crop yield and productivity during adverse environmental conditions.     

The pepper late embryogenesis abundant protein CaLEA1 acts in regulating abscisic acid signaling,drought and salt stress response

As sessile organisms, plants are constantly challenged by environmental stresses, including drought and high salinity. Among the various abiotic stresses, osmotic stress is one of the most important factors for growth and significantly reduces crop productivity in agriculture. Here, we report a function of the CaLEA1 protein in the defense responses of plants to osmotic stress. Our analyses showed that the CaLEA1 gene was strongly induced in pepper leaves exposed to drought and increased salinity. Furthermore, we determined that the CaLEA1 protein has a late embryogenesis abundant (LEA)_3 homolog domain highly conserved among other known group 5 LEA proteins and is localized in the processing body. We generated CaLEA1‐silenced peppers and CaLEA1‐overexpressing (OX) transgenic Arabidopsis plants to evaluate their responses to dehydration and high salinity. Virus‐induced gene silencing of CaLEA1 in pepper plants conferred enhanced sensitivity to drought and salt stresses, which was accompanied by high levels of lipid peroxidation in dehydrated and NaCl‐treated leaves. CaLEA1‐OX plants exhibited enhanced sensitivity to abscisic acid (ABA) during seed germination and in the seedling stage; furthermore, these plants were more tolerant to drought and salt stress than the wild‐type plants because of enhanced stomatal closure and increased expression of stress‐responsive genes. Collectively, our data suggest that CaLEA1 positively regulates drought and salinity tolerance through ABA‐mediated cell signaling.     

Heterologous expression of Arabidopsis H+‐pyrophosphatase enhances salt tolerance in transgenic creeping bentgrass (Agrostis stolonifera L.)

The Arabidopsis vacuolar H⁺‐pyrophosphatase (AVP1), when over‐expressed in transgenic (TG) plants, regulates root and shoot development via facilitation of auxin flux, and enhances plant resistance to salt and drought stresses. Here, we report that TG perennial creeping bentgrass plants over‐expressing AVP1 exhibited improved resistance to salinity than wild‐type (WT) controls. Compared to WT plants, TGs grew well in the presence of 100 mm NaCl, and exhibited higher tolerance and faster recovery from damages from exposure to 200 and 300 mm NaCl. The improved performance of the TG plants was associated with higher relative water content (RWC), higher Na⁺ uptake and lower solute leakage in leaf tissues, and with higher concentrations of Na⁺, K⁺, Cl^‐ and total phosphorus in root tissues. Under salt stress, proline content was increased in both WT and TG plants, but more significantly in TGs. Moreover, TG plants exhibited greater biomass production than WT controls under both normal and elevated salinity conditions. When subjected to salt stress, fresh (FW) and dry weights (DW) of both leaves and roots decreased more significantly in WT than in TG plants. Our results demonstrated the great potential of genetic manipulation of vacuolar H⁺‐pyrophosphatase expression in TG perennial species for improvement of plant abiotic stress resistance.     

Trehalose Biosynthesis in Response to Abiotic Stresses

Trehalose is a non‐reducing disaccharide that is present in diverse organisms ranging from bacteria and fungi to invertebrates, in which it serves as an energy source, osmolyte or protein/membrane protectant. The occurrence of trehalose and trehalose biosynthesis pathway in plants has been discovered recently. Multiple studies have revealed regulatory roles of trehalose‐6‐phosphate, a precursor of trehalose, in sugar metabolism, growth and development in plants. Trehalose levels are generally quite low in plants but may alter in response to environmental stresses. Transgenic plants overexpressing microbial trehalose biosynthesis genes have been shown to contain increased levels of trehalose and display drought, salt and cold tolerance. In‐silico expression profiling of all Arabidopsis trehalose‐6‐phosphate synthases (TPSs) and trehalose‐6‐phosphate phosphatases (TPPs) revealed that certain classes of TPS and TPP genes are differentially regulated in response to a variety of abiotic stresses. These studies point to the importance of trehalose biosynthesis in stress responses.     

Expression of an Arabidopsis vacuolar H+‐pyrophosphatase gene (AVP1) in cotton improves drought‐ and salt tolerance and increases fibre yield in the field conditions

The Arabidopsis gene AVP1 encodes a vacuolar pyrophosphatase that functions as a proton pump on the vacuolar membrane. Overexpression of AVP1 in Arabidopsis, tomato and rice enhances plant performance under salt and drought stress conditions, because up‐regulation of the type I H⁺‐PPase from Arabidopsis may result in a higher proton electrochemical gradient, which facilitates enhanced sequestering of ions and sugars into the vacuole, reducing water potential and resulting in increased drought‐ and salt tolerance when compared to wild‐type plants. Furthermore, overexpression of AVP1 stimulates auxin transport in the root system and leads to larger root systems, which helps transgenic plants absorb water more efficiently under drought conditions. Using the same approach, AVP1‐expressing cotton plants were created and tested for their performance under high‐salt and reduced irrigation conditions. The AVP1‐expressing cotton plants showed more vigorous growth than wild‐type plants in the presence of 200 mm NaCl under hydroponic growth conditions. The soil‐grown AVP1‐expressing cotton plants also displayed significantly improved tolerance to both drought and salt stresses in greenhouse conditions. Furthermore, the fibre yield of AVP1‐expressing cotton plants is at least 20% higher than that of wild‐type plants under dry‐land conditions in the field. This research indicates that AVP1 has the potential to be used for improving crop’s drought‐ and salt tolerance in areas where water and salinity are limiting factors for agricultural productivity.     

A rice really interesting new gene H2‐type E3 ligase,OsSIRH2‐14, enhances salinity tolerance via ubiquitin/26S proteasome‐mediated degradation of salt‐related proteins

Salinity is a deleterious abiotic stress factor that affects growth, productivity, and physiology of crop plants. Strategies for improving salinity tolerance in plants are critical for crop breeding programmes. Here, we characterized the rice (Oryza sativa) really interesting new gene (RING) H2‐type E3 ligase, OsSIRH2‐14 (previously named OsRFPH2‐14), which plays a positive role in salinity tolerance by regulating salt‐related proteins including an HKT‐type Na⁺ transporter (OsHKT2;1). OsSIRH2‐14 expression was induced in root and shoot tissues treated with NaCl. The OsSIRH2‐14‐EYFP fusion protein was predominately expressed in the cytoplasm, Golgi, and plasma membrane of rice protoplasts. In vitro pull‐down assays and bimolecular fluorescence complementation assays revealed that OsSIRH2‐14 interacts with salt‐related proteins, including OsHKT2;1. OsSIRH2‐14 E3 ligase regulates OsHKT2;1 via the 26S proteasome system under high NaCl concentrations but not under normal conditions. Compared with wild type plants, OsSIRH2‐14‐overexpressing rice plants showed significantly enhanced salinity tolerance and reduced Na⁺ accumulation in the aerial shoot and root tissues. These results suggest that the OsSIRH2‐14 RING E3 ligase positively regulates the salinity stress response by modulating the stability of salt‐related proteins.     

RhEXPA4, a rose expansin gene, modulates leaf growth and confers drought and salt tolerance to Arabidopsis

Drought and high salinity are major environmental conditions limiting plant growth and development. Expansin is a cell-wall-loosening protein known to disrupt hydrogen bonds between xyloglucan and cellulose microfibrils. The expression of expansin increases in plants under various abiotic stresses, and plays an important role in adaptation to these stresses. We aimed to investigate the role of the RhEXPA4, a rose expansin gene, in response to abiotic stresses through its overexpression analysis in Arabidopsis. In transgenic Arabidopsis harboring the Pro _RhEXPA4 ::GUS construct, RhEXPA4 promoter activity was induced by abscisic acid (ABA), drought and salt, particularly in zones of active growth. Transgenic lines with higher RhEXPA4 level developed compact phenotypes with shorter stems, curly leaves and compact inflorescences, while the lines with relatively lower RhEXPA4 expression showed normal phenotypes, similar to the wild type (WT). The germination percentage of transgenic Arabidopsis seeds was higher than that of WT seeds under salt stress and ABA treatments. Transgenic plants showed enhanced tolerance to drought and salt stresses: they displayed higher survival rates after drought, and exhibited more lateral roots and higher content of leaf chlorophyll a under salt stress. Moreover, high-level RhEXPA4 overexpressors have multiple modifications in leaf blade epidermal structure, such as smaller, compact cells, fewer stomata and midvein vascular patterning in leaves, which provides them with more tolerance to abiotic stresses compared to mild overexpressors and the WT. Collectively, our results suggest that RhEXPA4, a cell-wall-loosening protein, confers tolerance to abiotic stresses through modifying cell expansion and plant development in Arabidopsis.     

Overexpression of AtNHX5 improves tolerance to both salt and water stress in rice (Oryza sativa L.)

Overexpression of NHX genes has been previously shown to improve salt tolerance of transgenic plants. In this study, transgenic rice plants overexpressing AtNHX5 showed not only high salt tolerance, but also high drought tolerance. Measurements of ion levels indicated that Na⁺ and K⁺ contents were all higher in AtNHX5 overexpressing shoots than in wild type (WT) shoots in high saline conditions. After exposure to water deficiency and salt stress, the WT plants all died, while the AtNHX5 overexpressing rice plants had a higher survival rate, dry weight, leaf water content, and leaf chlorophyll contents, accumulated more proline, and had less membrane damage than the WT plants. In addition, seeds of both transgenic and WT plants germinated on 1/2 MS medium supplemented with 250 mM mannitol, but overexpression of AtNHX5 improved the shoot growth of the seedlings. Taken together, the results indicate that AtNHX5 gene could enhance the tolerance of rice plants to multiple environmental stresses by promoting the accumulation of more effective osmolytes (ions or proline) to counter the osmotic stress caused by abiotic factors.     

HbCIPK2, a novel CBL-interacting protein kinase from halophyte Hordeum brevisubulatum, confers salt and osmotic stress tolerance

Protein kinases play an important role in regulating the response to abiotic stress in plant. CIPKs are plant‐specific signal transducers, and some members have been identified. However, the precise functions of novel CIPKs still remain unknown. Here we report that HbCIPK2 is a positive regulator of salt and osmotic stress tolerance. HbCIPK2 was screened out of the differentially expressed fragments from halophyte Hordeum brevisubulatum by cDNA‐AFLP technique, and was a single‐copy gene without intron. Expression of HbCIPK2 was increased by salt, drought and ABA treatment. HbCIPK2 is mainly localized to the plasma membrane and nucleus. Ectopic expression of 35S:HbCIPK2 not only rescued the salt hypersensitivity in Arabidopsis mutant sos2‐1, but also enhanced salt tolerance in Arabidopsis wild type, and exhibited tolerance to osmotic stress during germination. The HbCIPK2 contributed to the ability to prevent K⁺ loss in root and to accumulate less Na⁺ in shoot resulting in K⁺/Na⁺ homeostasis and protection of root cell from death, which is consistent with the gene expression profile of HbCIPK2‐overexpressing lines. These findings imply possible novel HbCIPK2‐mediated salt signalling pathways or networks in H. brevisubulatum.     

A tomato glutaredoxin gene SlGRX1 regulates plant responses to oxidative, drought and salt stresses

Glutaredoxins (Grxs) are ubiquitous small heat-stable disulfide oxidoreductases that play a crucial role in plant development and response to oxidative stress. Here, a novel cDNA fragment (SlGRX1) from tomato encoding a protein containing the consensus Grx family domain with a CGFS active site was isolated and characterized. Southern blot analysis indicated that SlGRX1 gene had a single copy in tomato genome. Quantitative real-time RT-PCR analysis revealed that SlGRX1 was expressed ubiquitously in tomato including leaf, root, stem and flower, and its expression could be induced by oxidative, drought, and salt stresses. Virus-induced gene silencing mediated silencing of SlGRX1 in tomato led to increased sensitivity to oxidative and salt stresses with decreased relative chlorophyll content, and reduced tolerance to drought stress with decreased relative water content. In contrast, over-expression of SlGRX1 in Arabidopsis plants significantly increased resistance of plants to oxidative, drought, and salt stresses. Furthermore, expression levels of oxidative, drought and salt stress related genes Apx2, Apx6, and RD22 were up-regulated in SlGRX1-overexpressed Arabidopsis plants when analyzed by quantitative real-time PCR. Our results suggest that the Grx gene SlGRX1 plays an important role in regulating abiotic tolerance against oxidative, drought, and salt stresses.     

Ectopic expression of ADP ribosylation factor 1 (SaARF1) from smooth cordgrass (Spartina alterniflora Loisel) confers drought and salt tolerance in transgenic rice and Arabidopsis

Salinity and drought are two very important abiotic stressors that negatively impact the growth and yield of all sensitive crop plants. Genes from halophytes have been shown to be useful to engineer crop plants that can survive under adverse soil and water conditions. The present report establishes, for the first time, the physiological role of a class one ADP ribosylation factor gene (SaARF1) from the halophyte Spartina alterniflora (smooth cordgrass) in imparting salinity and drought stress tolerance when expressed in both monocot (rice) and dicot (Arabidopsis) systems. The Arabidopsis and rice plants overexpressing ARF1 are many-fold more tolerant to salt and drought than wild-type (WT) plants. The transgenics exhibited improved growth and productivity relative to WT through tissue tolerance by maintaining higher relative water content and membrane stability, and higher photosynthetic yield by retaining higher chlorophyll concentration and fluorescence under stress conditions compared to WT. These findings indicated that genes from halophyte resources can be useful to engineer and improve salt and drought stress tolerance in both monocot and dicot plants.