Salt stress reveals differential physiological,biochemical and molecular responses in <Emphasis Type="Italic">T. monococcum</Emphasis> and <Emphasis Type="Italic">T. durum</Emphasis> wheat genotypes

Salt stress responses implicate a complex mechanism and differ from plant species to another. In this study, we analyzed the physiological, biochemical and molecular responses to salt stress of the diploid wheat (T. monococcum) and compared to the tetraploid wheat (T. durum). Our results showed that the diploid wheat cultivar (cv. Turkey) is relatively tolerant to different salt stress conditions than the tetraploid wheat cultivar (cv. Om Rabia3). This tolerance was manifested by significant germination, plant growth and uptake of water generating cell turgor and development. Moreover, total chlorophyll content was higher in the diploid wheat than that in the tetraploid wheat. The Na⁺ content in leaf blade of the cv. Om Rabia3 was significantly higher than that of the cv. Turkey, suggesting that the diploid cultivar accumulates less toxic sodium in the photosynthetic tissues. This mechanism could be explained by the recirculation of the toxic ions Na⁺ into the xylem sap by SOS1 protein, which coordinates with HKT-like proteins to reduce the accumulation of Na⁺ ions in leaf blade. Interestingly, the expression of the three genes SOS1, HKT and NHX was enhanced under salinity especially in leaf blade of the cv. Turkey. Moreover, this wheat cultivar induced the antioxidative enzymes CAT and SOD activity more efficiently than the other cultivar.     

Molecular Cloning and Functional Analysis of a Na<Superscript>+</Superscript>/H<Superscript>+</Superscript> Antiporter Gene <Emphasis Type="Italic">ThNHX1</Emphasis> from a Halophytic Plant <Emphasis Type="Italic">Thellungiella halophila</Emphasis>

Na⁺/H⁺ exchanger catalyzes the countertransport of Na⁺ and H⁺ across membranes. Using the rapid amplification of cDNA ends method, a Na⁺/H⁺ antiporter gene (ThNHX1) was isolated from a halophytic plant, salt cress (Thellungiella halophila). The deduced amino acid sequence contained 545 amino acid residues with a conserved amiloride-binding domain (⁸⁷LFFIYLLPPI⁹⁶) and shared more than 94% identity with that of AtNHX1 from Arabidopsis thaliana. The ThNHX1 mRNA level was upregulated by salt and other stresses (abscisic acid, polyethylene glycol, and high temperature). This gene partially complemented the Na⁺/Li⁺-sensitive phenotype of a yeast mutant that was deficient in the endosomal–vacuolar Na⁺/H⁺ antiporter ScNHX1. Overexpression of ThNHX1 in Arabidopsis increased salt tolerance of transgenic plants compared with the wild-type plants. In addition, the silencing of ThNHX1 gene in T. halophila caused the transgenic plants to be more salt and osmotic sensitive than wild-type plant. Together, these results suggest that ThNHX1 may function as a tonoplast Na⁺/H⁺ antiporter and play an important role in salt tolerance of T. halophila. Chunxia Wu, Xiuhua Gao, and Xiangqiang Kong contributed equally to this work.     

Increased tolerance to salt stress in the phosphate-accumulating <Emphasis Type="Italic">Arabidopsis</Emphasis> mutants <Emphasis Type="Italic">siz1</Emphasis> and <Emphasis Type="Italic">pho2</Emphasis>

High salinity is an environmental factor that inhibits plant growth and development, leading to large losses in crop yields. We report here that mutations in SIZ1 or PHO2, which cause more accumulation of phosphate compared with the wild type, enhance tolerance to salt stress. The siz1 and pho2 mutations reduce the uptake and accumulation of Na⁺. These mutations are also able to suppress the Na⁺ hypersensitivity of the sos3-1 mutant, and genetic analyses suggest that SIZ1 and SOS3 or PHO2 and SOS3 have an additive effect on the response to salt stress. Furthermore, the siz1 mutation cannot suppress the Li⁺ hypersensitivity of the sos3-1 mutant. These results indicate that the phosphate-accumulating mutants siz1 and pho2 reduce the uptake and accumulation of Na⁺, leading to enhanced salt tolerance, and that, genetically, SIZ1 and PHO2 are likely independent of SOS3-dependent salt signaling.     

Disruption of <Emphasis Type="Italic">RCI2A</Emphasis> leads to over-accumulation of Na<Superscript>+</Superscript> and increased salt sensitivity in <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis> plants

For plant salt tolerance, it is important to regulate the uptake and accumulation of Na⁺ ions. The yeast pmp3 mutant which lacks PMP3 gene accumulates excess Na⁺ ions in the cell and shows increased Na⁺ sensitivity. Although the function of PMP3 is not fully understood, it is proposed that PMP3 contributes to the restriction of Na⁺ uptake and consequently salt tolerance in yeasts. In this paper, we have investigated whether the lack of RCI2A gene, homologous to PMP3 gene, causes a salt sensitive phenotype in Arabidopsis (Arabidopsis thaliana (L.) Heynh.) plants; and to thereby indicate the physiological role of RCI2A in higher plants. Two T-DNA insertional mutants of RCI2A were identified. Although the growth of rci2a mutants was comparable with that of wild type under normal conditions, high NaCl treatment caused increased accumulation of Na⁺ and more reduction of the growth of roots and shoots of rci2a mutants than that of wild type. Undifferentiated callus cultures regenerated from rci2a mutants also accumulated more Na⁺ than that from wild type under high NaCl treatment. Furthermore, when wild-type and rci2a plants were treated with NaCl, NaNO₃, Na₂SO₄, KCl, KNO₃, K₂SO₄ or LiCl, the rci2a mutants showed more reduction of shoot growth than wild type. Under treatments of tetramethylammonium chloride, CaCl₂, MgCl₂, mannitol or sorbitol, the growth reduction was comparable between wild-type and rci2a plants. These results suggested that RCI2A plays a role directly or indirectly for avoiding over-accumulation of excess Na⁺ and K⁺ ions in plants, and contributes to salt tolerance.     

Responses to nonaeration and/or salinity stress in hydroponically cultured <Emphasis Type="Italic">Populus nigra</Emphasis> and <Emphasis Type="Italic">Populus alba</Emphasis> cuttings

Growth, photosynthesis, and Na⁺, K⁺, Ca²⁺, and Mg²⁺ distributions were examined in two-year-old hydroponically cultured Populus nigra and Populus alba cuttings exposed to salt stress (0, 50, or 100 mM NaCl) for four or six weeks and to nonaeration stress for one or three weeks, followed by a three-week aeration period in 2/5 Hoagland solution. Salt stress with 100 mM NaCl totally inhibited height increase in P. nigra cuttings. Combined salinity and nonaeration inhibited height increase to a greater degree than either stress alone in both species. Simple salt stress did not affect diameter increase in P. alba, whereas combined high salinity (100 mM NaCl) and nonaeration inhibited diameter increase. Growth and biomass accumulation were more sensitive to salt stress in P. nigra cuttings than in P. alba, although P. alba showed a more rapid decrease in photosynthesis in response to nonaeration stress. Ion distributions in the leaves and roots differed between species. P. alba was superior to P. nigra in terms of Na⁺ exclusion capacity, such that most of the absorbed Na⁺ was confined to the root system, with little reaching the leaves. The distributions of K⁺, Ca²⁺, and Mg²⁺ in the leaves and roots of each species under the two stressors were also analyzed. The lower Na⁺/K⁺ ratio in leaves indicated that P. alba was more tolerant to salt stress than P. nigra.     

Involvement of Long-Distance Na<Superscript>+</Superscript> Transport in Maintaining Water Potential Gradient in the Medium-Root-Leaf System of a Halophyte <Emphasis Type="Italic">Suaeda altissima</Emphasis>

The aim of this study was to determine the range of NaCl concentrations in the nutrient solution that allow Suaeda altissima (L.) Pall., a salt-accumulating halophyte, to maintain the upward gradient of water potential in the “medium-root-leaf” system. We evaluated the contribution of Na⁺ ions in the formation of water potential gradient and demonstrated that Na⁺ loading into the xylem is involved in this process. Plants were grown in water culture at NaCl concentrations ranging from zero to 1 M. The water potential of leaf and root cells was measured with the method of isopiestic thermocouple psychrometry. When NaCl concentration in the growth medium was raised in the range of 0–500 mM (the medium water potential was lowered accordingly), the root and leaf cells of S. altissima decreased their water potential, thus promoting the maintenance of the upward water potential gradient in the medium-root-leaf system. Growing S. altissima at NaCl concentrations f 750 mM and 1 M disordered water homeostasis and abolished the upward gradient of water potential between roots and leaves. At NaCl concentrations of 0–250 mM, the detached roots of S. altissima were capable of producing the xylem exudate. The concentration of Na⁺ in the exudate was 1.3 to 1.6 times higher than in the nutrient medium; the exudate pH was acidic and was lowered from 5.5 to 4.5 with the rise in the salt concentration. The results indicate that the long-distance Na⁺ transport and, especially, the mechanism of Na⁺ loading into the xylem play a substantial role in the formation of water potential gradient in S. altissima. The accumulation of Na⁺ in the xylem and acidic pH values of the xylem sap suggest that Na⁺ loading into the xylem is carried out by the Na⁺/H⁺ antiporter of the plasma membrane in parenchymal cells of the root stele.__________Translated from Fiziologiya Rastenii, Vol. 52, No. 4, 2005, pp. 549–557.Original Russian Text Copyright © 2005 by Balnokin, Kotov, Myasoedov, Khailova, Kurkova, Lun’kov, Kotova.     

Cloning of two SOS1 transporters from the seagrass <Emphasis Type="Italic">Cymodocea nodosa</Emphasis>. SOS1 transporters from <Emphasis Type="Italic">Cymodocea</Emphasis> and <Emphasis Type="Italic">Arabidopsis</Emphasis> mediate potassium uptake in bacteria

Two cDNAs isolated from Cymodocea nodosa, CnSOS1A, and CnSOS1B encode proteins with high-sequence similarities to SOS1 plant transporters. CnSOS1A expressed in a yeast Na⁺-efflux mutant under the control of a constitutive expression promoter mimicked AtSOS1 from Arabidopsis; the wild type cDNA did not improve the growth of the recipient strain in the presence of Na⁺, but a cDNA mutant that expresses a truncated protein suppressed the defect of the yeast mutant. In similar experiments, CnSOS1B was not effective. Conditional expression, under the control of an arabinose responsive promoter, of the CnSOS1A and CnSOS1B cDNAs in an Escherichia coli mutant defective in Na⁺ efflux was toxic, and functional analyses were inconclusive. The same constructs transformed into an E. coli K⁺-uptake mutant revealed that CnSOS1A was also toxic, but that it slightly suppressed defective growth at low K⁺. Truncation in the C-terminal hydrophilic tail of CnSOS1A relieved the toxicity and proved that CnSOS1A was an excellent low-affinity K⁺ and Rb⁺ transporter. CnSOS1B mediated a transient, extremely rapid K⁺ or Rb⁺ influx. Similar tests with AtSOS1 revealed that it was not toxic and that the whole protein exhibited excellent K⁺ and Rb⁺ uptake characteristics in bacteria.     

Production of Exo-polysaccharide by <Emphasis Type="Italic">Rhizobium</Emphasis> sp.

Two fold increase in the yield of glucose and maltose containing exo-polysaccharide (EPS) by Rhizobium sp. was observed during its growth in modified YEMB. EPS production, plant growth promotion activity and root colonization of Rhizobium sp. studies showed enhanced EPS synthesis, more seed germination and over all improvement in plant growth over control and R. meliloti treatment. Groundnut seeds bacterized with Rhizobium sp. resulted in 69.75% more root length, 49.51% more shoot height, 13.75% more number of branches and 13.60% more number of pods over the control and R. meliloti treatment. Bacterization of wheat seeds increased the dry matter yield of roots (1.7-fold), and roots adhering soil (RAS) (1.5) and shoot mass (1.9-fold). Rhizobium sp. inoculation also increased the population density of EPS-producing bacteria on the rhizoplane. Roots of plants inoculated with Rhizobium sp. maintained a higher K⁺/Na⁺ ratio and K⁺–Na⁺ selectivity.     

Overexpression of <Emphasis Type="Italic">OsVP1</Emphasis> and <Emphasis Type="Italic">OsNHX1</Emphasis> Increases Tolerance to Drought and Salinity in Rice

Drought and salinity are major abiotic stresses affecting rice production. To improve plant tolerance to salinity and drought, we overexpressed rice Na⁺/H⁺ exchangers (OsNHX1) and H⁺-pyrophosphatase in tonoplasts (OsVP1) in a japonica elite rice cultivar, Zhonghua 11. Compared with our wild-type control, transgenic plants overexpressing both genes incurred less damage when exposed to long-term treatment with 100 mM NaCl or water deprivation. Under high-saline conditions, the transformants accumulated less Na⁺ and malondialdehyde in the leaves, thereby allowing the plants to maintain a low level of leaf water potential and reduce stress-induced damage. Those transgenics also had higher photosynthetic activity during the stress period. Under those conditions, they also showed an increase in root biomass, which enabled more water uptake. These results suggest that OsVP1 and OsNHX1 improve the tolerance of rice crops against drought and salt by employing multiple strategies in addition to osmotic regulation.     

The wheat gene <Emphasis Type="Italic">TaST</Emphasis> can increase the salt tolerance of transgenic <Emphasis Type="Italic">Arabidopsis</Emphasis>

On the basis of the results of gene chip analysis of the salt-tolerant wheat mutant RH8706-49 under conditions of salt stress, we identified and cloned an unknown salt-induced gene TaST (Triticum aestivum salt-tolerant). Real-time quantitative PCR analysis showed that the expression of the gene was induced by salt stress. Transgenic Arabidopsis plants overexpressing the TaST gene showed higher salt tolerance than the wild-type controls. Subcellular localization studies revealed that the protein encoded by this gene was in the nucleus. In comparison with wild-type controls, transgenic Arabidopsis plants accumulated more Ca²⁺, soluble sugar, and proline and less Na⁺ under salt stress. Real-time quantitative PCR analysis showed that Arabidopsis plants overexpressing TaST also showed increased expression of many stress-related genes. All these findings indicated that TaST can enhance the salt tolerance of transgenic Arabidopsis plants.     

Effect of salt stress on physiological and antioxidative responses in two species of <Emphasis Type="Italic">Salicornia</Emphasis> (<Emphasis Type="Italic">S. persica</Emphasis> and <Emphasis Type="Italic">S. europaea</Emphasis>)

The effects of salt stress on growth parameters, free proline content, ion accumulation, lipid peroxidation, and several antioxidative enzymes activities were investigated in S. persica and S. europaea. The seedlings were grown for 2 months in half-strength Hoagland solution and treated with different concentrations of NaCl (0, 85, 170, 340, and 510 mM) for 21 days. The fresh and dry weights of both species increased significantly at 85 and 170 mM NaCl and decreased at higher concentrations. Salinity increased proline content in both the species as compared to that of control. Sodium (Na⁺) content in roots and shoots increased, whereas K⁺ and P_i content in both organs decreased. At all NaCl concentrations, the total amounts of Na⁺ and K⁺ were higher in shoots than in roots. Malondialdehyde (MDA) content declined at moderate NaCl concentrations (85 and 170 mM) and increased at higher levels. With increased salinity, superoxide dismutase (SOD), catalase (CAT), and guaiacol peroxidase (GPX) activities also increased gradually in both species. In addition, it seems that GPX, CAT, and SOD activities play an essential protective role in the scavenging reactive oxygen species (ROS) in both species. Native polyacrylamide gel electrophoresis (PAGE) indicated different isoform profiles between S. persica and S. europaea concerning antioxidant enzymes. These results showed that S. persica exhibits a better protection mechanism against oxidative damage and it is more salt-tolerant than S. europaea possibly by maintaining and/or increasing growth parameters, ion accumulation, and antioxidant enzyme activities.     

Contribution of <Emphasis Type="Italic">Glomus intraradices</Emphasis> inoculation to nutrient acquisition and mitigation of ionic imbalance in NaCl-stressed <Emphasis Type="Italic">Trigonella foenum-graecum</Emphasis>

The study aimed to investigate the effects of an AM fungus (Glomus intraradices Schenck and Smith) on mineral acquisition in fenugreek (Trigonella foenum-graecum) plants under different levels of salinity. Mycorrhizal (M) and non-mycorrhizal (NM) fenugreek plants were subjected to four levels of NaCl salinity (0, 50, 100, and 200 mM NaCl). Plant tissues were analyzed for different mineral nutrients. Leaf senescence (chlorophyll concentration and membrane permeability) and lipid peroxidation were also assessed. Under salt stress, M plants showed better growth, lower leaf senescence, and decreased lipid peroxidation as compared to NM plants. Salt stress adversely affected root nodulation and uptake of NPK. This effect was attenuated in mycorrhizal plants. Presence of the AM fungus prevented excess uptake of Na⁺ with increase in NaCl in the soil. It also imparted a regulatory effect on the translocation of Na⁺ ions to shoots thereby maintaining lower Na⁺ shoot:root ratios as compared to NM plants. Mycorrhizal colonization helped the host plant to overcome Na⁺-induced Ca²⁺ and K⁺ deficiencies. M plants maintained favorable K⁺:Na⁺, Ca²⁺:Na⁺, and Ca²⁺:Mg²⁺ ratios in their tissues. Concentrations of Cu, Fe, and Zn²⁺ decreased with increase in intensity of salinity stress. However, at each NaCl level, M plants had higher concentration of Cu, Fe, Mn²⁺, and Zn²⁺ as compared to NM plants. M plants showed reduced electrolyte leakage in leaves as compared to NM plants. The study suggests that AM fungi contribute to alleviation of salt stress by mitigation of NaCl-induced ionic imbalance thus maintaining a favorable nutrient profile and integrity of the plasma membrane.     

Increase in Salt Tolerance of <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis> by <Emphasis Type="Italic">TaDi19</Emphasis>

The gene expression profile chip of salt-resistant wheat mutant RH8706-49 under salt stress was investigated. The overall length of the cDNA sequence of the probe was obtained using electronic cloning and RT-PCR. An unknown gene induced by salt was obtained, cloned, and named TaDi19 (Triticum aestivum drought-induced protein). No related report or research on the protein is available. qPCR analysis showed that gene expression was induced by many stresses, such as salt. Arabidopsis thaliana was genetically transferred using the overexpressing gene, which increased its salt tolerance. After salt stress, the transgenic plant demonstrated better physiological indicators (higher Ca²⁺ and lower Na⁺) than those of the wild-type plant. Results of non-invasive micro-test technology indicate that TaDi19-overexpressing A. thaliana significantly effluxed Na⁺ after salt treatment, whereas the wild-type plant influxed Na⁺. Chelating extracellular Ca²⁺ resulted in insignificant differences in salt tolerance between overexpressing and wild-type A. thaliana. Subcellular localization showed that the gene encoding protein was mainly located in the cell membrane and nucleus. TaDi19 was overexpressed in wild-type A. thaliana, and the transgenic lines were more salt-tolerant than the control A. thaliana. Thus, the wheat gene TaDi19 could increase the salt tolerance of A. thaliana.     

Overexpression of a <Emphasis Type="Italic">Malus</Emphasis> vacuolar Na<Superscript>+</Superscript>/H<Superscript>+</Superscript> antiporter gene (<Emphasis Type="Italic">MdNHX1</Emphasis>) in apple rootstock M.26 and its influence on salt tolerance

Soil salinity is a major factor limiting apple production in some areas. Tonoplast Na⁺/H⁺ antiporters play a critical role in salt tolerance. Here, we isolated MdNHX1, a vacuolar Na⁺/H⁺ antiporter from Luo-2, a salt-tolerant rootstock of apple (Malus × domestica Borkh.), and introduced it into apple rootstock M.26 by Agrobacterium-mediated transformation. PCR and DNA gel blot analyses confirmed successful integration of MdNHX1. RT-PCR analysis indicated that the gene was highly expressed in transgenic plants, but the degree of this expression varied among lines. Its overexpression conferred high tolerance to salt stress. Analysis of ion contents showed that, when exposed to salinity stress, the transgenics compartmentalized more Na⁺ in the roots and also maintained a relatively high K⁺/Na⁺ ratio in the leaves compared with non-transformed plants. Under normal conditions, however, amounts of potassium and sodium did not differ significantly between transgenic and control plants.     

Contributions of inorganic ions,soluble carbohydrates,and multiatomic alcohols to water homeostasis in <Emphasis Type="Italic">Artemisia lerchiana</Emphasis> and <Emphasis Type="Italic">A. pauciflora</Emphasis>

Two morphological forms of wormwood Artemisia lerchiana (f. erecta and f. nutans) and A. pauciflora Web. (morphological form erecta) were grown on sand culture at a range of NaCl concentrations in the nutrient medium and then assayed for Na⁺, K⁺, and Cl^? content in various organs. In addition, the content of mono-, di-, and trisaccharides and multiatomic alcohols (mannitol and glycerol); water content; and organ biomass were determined. All plants examined showed high NaCl tolerance, comparable to that of halophytes. They were able to maintain high tissue hydration under conditions of salinity-induced growth suppression. The intracellular osmotic pressure in wormwood organs was mainly determined by the presence of Na⁺, K⁺, and Cl^?, as well as by mono-, di-, and trisaccharides, mannitol, and glycerol. The high content of Na⁺ and Cl^? in wormwood organs was also observed in the absence of salinity, which implies the ability of these organs to absorb ions from diluted NaCl solutions and accumulate ions in cells of their tissues. With the increase in salinity, the content of Na⁺ and Cl^? in roots and leaves increased to even higher levels. It is concluded that the ability of wormwood plants to absorb and accumulate inorganic ions provides for sustainable high intracellular osmotic pressure and, accordingly, low water potential under drought and salinity conditions. Growing plants under high salinity lowered the content of monosaccharides in parallel with accumulation of the trisaccharide raffinose. It is supposed that soluble carbohydrates and multiatomic alcohols are not only significant for osmoregulation but also perform a protective function in wormwood plants. The lower osmotic pressure in root cells compared to that in leaf cells of all plants examined was mainly due to the gradient distribution of K⁺ and Cl^? between roots and leaves. The two Artemisia species and two morphological forms of A. lerchiana did not differ appreciably in the ways of water balance regulation. It is found that different morphologies of two A. lerchiana forms are unrelated to variations in intracellular osmotic and turgor pressures.     

Comparative effectiveness of <Emphasis Type="Italic">Pseudomonas</Emphasis> and <Emphasis Type="Italic">Serratia</Emphasis> sp. containing ACC-deaminase for improving growth and yield of wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.) under salt-stressed conditions

Ethylene synthesis is accelerated in response to various environmental stresses like salinity. Ten rhizobacterial strains isolated from wheat rhizosphere taken from different salt affected areas were screened for growth promotion of wheat under axenic conditions at 1, 5, 10 and 15 dS m⁻¹. Three strains, i.e., Pseudomonas putida (N21), Pseudomonas aeruginosa (N39) and Serratia proteamaculans (M35) showing promising performance under axenic conditions were selected for a pot trial at 1.63 (original), 5, 10 and 15 dS m⁻¹. Results showed that inoculation was effective even in the presence of higher salinity levels. P. putida was the most efficient strain compared to the other strains and significantly increased the plant height, root length, grain yield, 100-grain weight and straw yield up to 52, 60, 76, 19 and 67%, respectively, over uninoculated control at 15 dS m⁻¹. Similarly, chlorophyll content and K⁺/Na⁺ of leaves also increased by P. putida over control. It is highly likely that under salinity stress, 1-aminocyclopropane-1-carboxylic acid-deaminase activity of these microbial strains might have caused reduction in the synthesis of stress (salt)-induced inhibitory levels of ethylene. The results suggested that these strains could be employed for salinity tolerance in wheat; however, P. putida may have better prospects in stress alleviation/reduction.     

Transport of Na<Superscript>+</Superscript> and K<Superscript>+</Superscript> by an antiporter-related subunit from the <Emphasis Type="Italic">Escherichia coli </Emphasis>NADH dehydrogenase I produced in <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis>

The NADH dehydrogenase I from Escherichia coli is a bacterial homolog of the mitochondrial complex I which translocates Na⁺ rather than H⁺. To elucidate the mechanism of Na⁺ transport, the C-terminally truncated NuoL subunit (NuoL_N) which is related to Na⁺/H⁺ antiporters was expressed as a protein A fusion protein (ProtA–NuoL_N) in the yeast Saccharomyces cerevisiae which lacks an endogenous complex I. The fusion protein inserted into membranes from the endoplasmatic reticulum (ER), as confirmed by differential centrifugation and Western analysis. Membrane vesicles containing ProtA–NuoL_N catalyzed the uptake of Na⁺ and K⁺ at rates which were significantly higher than uptake by the control vesicles under identical conditions, demonstrating that ProtA–NuoL_N translocated Na⁺ and K⁺ independently from other complex I subunits. Na⁺ transport by ProtA–NuoL_N was inhibited by EIPA (5-(N-ethyl-N-isopropyl)-amiloride) which specifically reacts with Na⁺/H⁺ antiporters. The cation selectivity and function of the NuoL subunit as a transporter module of the NADH dehydrogenase complex is discussed. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

The role of root apoplastic transport barriers in salt tolerance of rice (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.)

Increasing soil salinity reduces crop yields worldwide, with rice being particularly affected. We have examined the correlation between apoplastic barrier formation in roots, Na⁺ uptake into shoots and plant survival for three rice (Oryza sativa L.) cultivars of varying salt sensitivity: the salt-tolerant Pokkali, moderately tolerant Jaya and sensitive IR20. Rice plants grown hydroponically or in soil for 1 month were subjected to both severe and moderate salinity stress. Apoplastic barriers in roots were visualized using fluorescence microscopy and their chemical composition determined by gas chromatography and mass spectrometry. Na⁺ content was estimated by flame photometry. Suberization of apoplastic barriers in roots of Pokkali was the most extensive of the three cultivars, while Na⁺ accumulation in the shoots was the least. Saline stress induced the strengthening of these barriers in both sensitive and tolerant cultivars, with increase in mRNAs encoding suberin biosynthetic enzymes being detectable within 30 min of stress. Enhanced barriers were detected after several days of moderate stress. Overall, more extensive apoplastic barriers in roots correlated with reduced Na⁺ uptake and enhanced survival when challenged with high salinity. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Molecular characterization and functional analysis of a vacuolar Na<Superscript>+</Superscript>/H<Superscript>+</Superscript> antiporter gene (<Emphasis Type="Italic">HcNHX1</Emphasis>) from <Emphasis Type="Italic">Halostachys caspica</Emphasis>

According to sequences of several vacuolar Na⁺/H⁺ antiporter genes from Xinjiang halophytic plants, a new vacuolar Na⁺/H⁺ antiporter gene (HcNHX1) from the halophyte Halostachys caspica was obtained by RACE and RT-PCR using primers corresponding to conserved regions of the coding sequences. The obtained HcNHX1 cDNA was 1,983 bp and contained a 1,656 bp open reading frame encoding a deduced protein of 551 amino acid residues. The deduced amino acid sequence showed high identity with other NHX1 we have cloned previously from halophyte in Xinjiang desert area. The phylogenetic analysis showed that HcNHX1 formed a clade with NHX homologs of Chenopodiaceae. Expression profiles under salt treatment and ABA induction were investigated, and the results revealed that expression of HcNHX1 was induced by NaCl and ABA. To compare the degree of salt tolerance, we over-expressed HcNHX1 in Arabidopsis. Two transgenic lines grew more vigorously than the wild type (WT) under salt stress. The analysis of ion contents indicated that under salt stress, the transgenic plants compartmentalized more Na⁺ in the leaves compared with wild-type plants. Together, these results suggest that the products of the novel gene HcNHX1 from halophyte Halostachys caspica is a functional tonoplast Na⁺/H⁺ antiporter.