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.     

Expression of Arabidopsis CBF1 regulated by an ABA/stress inducible promoter in transgenic tomato confers stress tolerance without affecting yield

Modern‐day plants are subjected to various biotic and abiotic stresses thereby limiting plant productivity and quality. It has previously been reported that the use of a strong constitutive 35S cauliflower mosaic virus (CaMV) promoter to drive the expression of Arabidopsis CBF1 in tomato improved tolerance to cold, drought and salt loading, at the expense of growth and yield under normal growth conditions. Hence in the present study, the suitability of expressing the Arabidopsis CBF1 driven by three copies of an ABA‐responsive complex (ABRC1) from the barley HAV22 gene in order to improve the agronomic performance of the transgenic tomato plants was investigated. Northern blot analysis indicated that CBF1 gene expression was induced by chilling, water‐deficit and salt treatment in the transgenic tomato plants. Under these tested stress conditions, transgenic tomato plants exhibited enhanced tolerance to chilling, water‐deficit, and salt stress in comparison with untransformed plants. Under normal growing conditions the ABRC1‐CBF1 tomato plants maintained normal growth and yield similar to the untransformed plants. The results demonstrate the promise of using ABRC1‐CBF1 tomato plants in highly stressed conditions which will in turn benefit agriculture.     

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.     

Tomato ABSCISIC ACID STRESS RIPENING (ASR) Gene Family Revisited

Tomato ABSCISIC ACID RIPENING 1 (ASR1) was the first cloned plant ASR gene. ASR orthologs were then cloned from a large number of monocot, dicot and gymnosperm plants, where they are mostly involved in response to abiotic (drought and salinity) stress and fruit ripening. The tomato genome encodes five ASR genes: ASR1, 2, 3 and 5 encode low-molecular-weight proteins (ca. 110 amino acid residues each), whereas ASR4 encodes a 297-residue polypeptide. Information on the expression of the tomato ASR gene family is scarce. We used quantitative RT-PCR to assay the expression of this gene family in plant development and in response to salt and osmotic stresses. ASR1 and ASR4 were the main expressed genes in all tested organs and conditions, whereas ASR2 and ASR3/5 expression was two to three orders of magnitude lower (with the exception of cotyledons). ASR1 is expressed in all plant tissues tested whereas ASR4 expression is limited to photosynthetic organs and stamens. Essentially, ASR1 accounted for most of ASR gene expression in roots, stems and fruits at all developmental stages, whereas ASR4 was the major gene expressed in cotyledons and young and fully developed leaves. Both ASR1 and ASR4 were expressed in flower organs, with ASR1 expression dominating in stamens and pistils, ASR4 in sepals and petals. Steady-state levels of ASR1 and ASR4 were upregulated in plant vegetative organs following exposure to salt stress, osmotic stress or the plant abiotic stress hormone abscisic acid (ABA). Tomato plants overexpressing ASR1 displayed enhanced survival rates under conditions of water stress, whereas ASR1-antisense plants displayed marginal hypersensitivity to water withholding.     

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.     

Expressing the yeast HAL1 gene in tomato increases fruit yield and enhances K+/Na+ selectivity under salt stress

The yeast HAL1 gene facilitates K⁺/Na⁺ selectivity and salt tolerance of cells. Ectopic expression of HAL1 in transgenic tomato (Lycopersicon esculentum Mill.) plants minimized the reduction in fruit production caused by salt stress. Maintenance of fruit production by transgenic plants was correlated with enhanced growth under salt stress of calli derived from the plants. The HAL1 transgene enhanced water and K⁺ contents in both leaf calli and leaves in the presence of salt, which indicates that HAL1 functions in plants using a similar mechanism to that in yeast, namely by facilitating K⁺/Na⁺ selectivity under salt stress.     

Increased salt tolerance with overexpression of cation/proton antiporter 1 genes: a meta‐analysis

Cation/proton antiporter 1 (CPA1) genes encode cellular Na⁺/H⁺ exchanger proteins, which act to adjust ionic balance. Overexpression of CPA1s can improve plant performance under salt stress. However, the diversified roles of the CPA1 family and the various parameters used in evaluating transgenic plants over‐expressing CPA1s make it challenging to assess the complex functions of CPA1s and their physiological mechanisms in salt tolerance. Using meta‐analysis, we determined how overexpression of CPA1s has influenced several plant characteristics involved in response and resilience to NaCl stress. We also evaluated experimental variables that favour or reduce CPA1 effects in transgenic plants. Viewed across studies, overexpression of CPA1s has increased the magnitude of 10 of the 19 plant characteristics examined, by 25% or more. Among the ten moderating variables, several had substantial impacts on the extent of CPA1 influence: type of culture media, donor and recipient type and genus, and gene family. Genes from monocotyledonous plants stimulated root K⁺, root K⁺/Na⁺, total chlorophyll, total dry weight and root length much more than genes from dicotyledonous species. Genes transformed to or from Arabidopsis have led to smaller CPA1‐induced increases in plant characteristics than genes transferred to or from other genera. Heterogeneous expression of CPA1s led to greater increases in leaf chlorophyll and root length than homologous expression. These findings should help guide future investigations into the function of CPA1s in plant salt tolerance and the use of genetic engineering for breeding of resistance.     

Overexpression of <Emphasis Type="Italic">SlERF1</Emphasis> tomato gene encoding an ERF-type transcription activator enhances salt tolerance

Ethylene-responsive factors (ERFs) play an important role in plant responses to stresses. For this study, we obtained sense- and antisense-SlERF1 transgenic tomato plants to analyze the function of the SlERF1 gene in tomato plants. Overexpression of SlERF1 in tomato plants enhanced salt tolerance during tomato seedling root development. In addition, tomato seedlings overexpressing SlERF1 showed the higher relative water content and lower MDA content and electrolyte leakage; they accumulated more free proline and soluble sugars, as compared with wild-type and antisense-SlERF1 transgenic tomato plants under salt stress. Moreover, SlERF1 activated the expression of stress-related genes, including LEA, P5CS, DREB3-1, and ltpg2 in tomato plants under salt stress. Thus, SlERF1 played a positive role in the salt tolerance of tomato plants.     

Overexpression of a wheat Na+/H+ antiporter gene (TaNHX2) enhances tolerance to salt stress in transgenic tomato plants (Solanum lycopersicum L.)

Soil salinity is a major environmental stress limiting plant productivity. Vacuole Na⁺/H⁺ antiporters play important roles for the survival of plants under salt stress conditions. We have developed salt stress tolerant transgenic tomato plants (Solanum lycopersicum cv. PED) by overexpression of the wheat Na⁺/H⁺ antiporter gene TaNHX2 using Agrobacterium tumefaciens strain LBA4404 harbouring a binary vector pBin438 that contains the TaNHX2 gene under the control of double CaMV 35S promoter and npt II as a selectable marker. PCR and Southern blot analysis confirmed that TaNHX2 gene has been integrated and expressed in the T₁ generation transgenic tomato plants. When TaNHX2 expressing plants were exposed to 100 or 150 mM NaCl, they were found to be more tolerant to salt stress compared to wild type plants. Biochemical analyses also showed that transgenic plants have substantial amount of relative water content and chlorophyll content under salt stress conditions compared to wild type plants. The relative water content in transgenic and wild type plants ranged from 68 to 75 % and 46–73 % and chlorophyll content fall in between 1.8 to 2.4 mg/g fw and 1.0 to 2.4 mg/g fw, respectively, in all stress conditions. In the present study, we observed a better germination rate of T₁ transgenic seeds under salt stress conditions compared with wild type plants. Our results indicated that TaNHX2-transgenic tomato plants coped better with salt stress than wild type plants.     

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.     

Phelipanche aegyptiaca parasitism impairs salinity tolerance in young leaves of tomato

The parasite Phelipanche aegyptiaca infests tomato, a crop plant that is commonly cultivated in semi‐arid environments, where tomato may be subject to salt stress. Since the relationship between the two stresses —salinity and parasitism – has been poorly investigated in tomato, the effects of P. aegyptiaca parasitism on tomato growing under moderate salinity were examined. Tomatoes were grown with regular or saline water irrigation (3 and 45 mM Cl^?, respectively) in soils infested with P. aegyptiaca . The infested plants accumulated higher levels of sodium and chloride ions in the roots, shoots and leaves (old and young) under both salinity levels vs. non‐infected plants. There was a positive linear correlation between P. aegyptiaca biomass and salt accumulation in young tomato leaves, and a negative linear correlation between parasite biomass and the osmotic potential of young tomato leaves. Concentrations of the osmoprotectants proline, myoinositol and sucrose were reduced in infected tomato plants, which impaired the host's osmotic adjustment ability. The sensitivity of P. aegyptiaca to salt stress was manifested as a decrease in biomass. In conclusion, P. aegyptiaca parasitism reduced the salt tolerance of tomato plants by promoting the accumulation of salts from the rhizosphere and impairing the host's osmotic adjustment ability.     

Overexpression of the PtSOS2 gene improves tolerance to salt stress in transgenic poplar plants

In higher plants, the salt overly sensitive (SOS) signalling pathway plays a crucial role in maintaining ion homoeostasis and conferring salt tolerance under salinity condition. Previously, we functionally characterized the conserved SOS pathway in the woody plant Populus trichocarpa. In this study, we demonstrate that overexpression of the constitutively active form of PtSOS2 (PtSOS2TD), one of the key components of this pathway, significantly increased salt tolerance in aspen hybrid clone Shanxin Yang (Populus davidiana × Populus bolleana). Compared to the wild‐type control, transgenic plants constitutively expressing PtSOS2TD exhibited more vigorous growth and produced greater biomass in the presence of high concentrations of NaCl. The improved salt tolerance was associated with a decreased Na⁺ accumulation in the leaves of transgenic plants. Further analyses revealed that plasma membrane Na⁺/H⁺ exchange activity and Na⁺ efflux in transgenic plants were significantly higher than those in the wild‐type plants. Moreover, transgenic plants showed improved capacity in scavenging reactive oxygen species (ROS) generated by salt stress. Taken together, our results suggest that PtSOS2 could serve as an ideal target gene to genetically engineer salt‐tolerant trees.     

Growth of mycorrhizal tomato and mineral acquisition under salt stress

 High salt levels in soil and water can limit agricultural production and land development in arid and semiarid regions. Arbuscular mycorrhizal fungi (AMF) have been shown to decrease plant yield losses in saline soils. The objective of this study was to examine the growth and mineral acquisition responses of greenhouse-grown tomato to colonization by the AMF Glomus mosseae [(Nicol. And Gerd.) Gerd. and Trappe] under varied levels of salt. NaCl was added to soil in the irrigation water to give an EC_e of 1.4 (control), 4.7 (medium) and 7.4 dS m^–1 (high salt stress). Plants were grown in a sterilized, low P (silty clay) soil-sand mix. Mycorrhizal colonization was higher in the control than in saline soil conditions. Shoot and root dry matter yields and leaf area were higher in mycorrhizal than in nonmycorrhizal plants. Total accumulation of P, Zn, Cu, and Fe was higher in mycorrhizal than in nonmycorrhizal plants under both control and medium salt stress conditions. Shoot Na concentrations were lower in mycorrhizal than in nonmycorrhizal plants grown under saline soil conditions. The improved growth and nutrient acquisition in tomato demonstrate the potential of AMF colonization for protecting plants against salt stress in arid and semiarid areas. Accepted: 21 February 2000