The Effects of Low Doses of Gamma-Radiation on Growth and Membrane Activity of <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> GRP3 and <Emphasis Type="Italic">Escherichia coli</Emphasis> M17

Microorganisms are part of the natural environments and reflect the effects of different physical factors of surrounding environment, such as gamma (γ) radiation. This work was devoted to the study of the influence of low doses of γ radiation with the intensity of 2.56?μW (m²?s)^?1 (absorbed doses were 3.8 mGy for the radiation of 15?min and 7.2 mGy—for 30?min) on Escherichia coli M-17 and Pseudomonas aeruginosa GRP3 wild type cells. The changes of bacterial, growth, survival, morphology, and membrane activity had been studied after γ irradiation. Verified microbiological (specific growth rate, lag phase duration, colony-forming units (CFU) number, and light microscopy digital image analysis), biochemical (ATPase activity of bacterial membrane vesicles), and biophysical (H⁺ fluxes throughout cytoplasmic membrane of bacteria) methods were used for assessment of radiation implications on bacteria. It was shown that growth specific rate, lag phase duration and CFU number of these bacteria were lowered after irradiation, and average cell surface area was decreased too. Moreover ion fluxes of bacteria were changed: for P. aeruginosa they were decreased and for E. coli—increased. The N,N′-dicyclohexylcarbodiimide (DCCD) sensitive fluxes were also changed which were indicative for the membrane-associated F₀F₁-ATPase enzyme. ATPase activity of irradiated membrane vesicles was decreased for P. aeruginosa and stimulated for E. coli. Furthermore, DCCD sensitive ATPase activity was also changed. The results obtained suggest that these bacteria especially, P. aeruginosa are sensitive to γ radiation and might be used for developing new monitoring methods for estimating environmental changes after γ irradiation.     

Functional Identification of H<Superscript>+</Superscript>-ATPase and Na<Superscript>+</Superscript>/H<Superscript>+</Superscript> Antiporter in the Plasma Membrane Isolated from the Root Cells of Salt-Accumulating Halophyte <Emphasis Type="Italic">Suaeda altissima</Emphasis>

A membrane fraction enriched in plasma membrane (PM) vesicles was isolated from the root cells of a salt-accumulating halophyte Suaeda altissima (L.) Pall. by means of centrifugation in discontinuous sucrose density gradient. The PM vesicles were capable of generating ΔpH at their membrane and the transmembrane electric potential difference (Δψ). These quantities were measured with optical probes, acridine orange and oxonol VI, sensitive to ΔpH and Δψ, respectively. The ATP-dependent generation of ΔpH was sensitive to vanadate, an inhibitor of P-type ATPases. The results contain evidence for the functioning of H⁺-ATPase in the PM of the root cells of S. altissima. The addition of Na⁺ and Li⁺ ions to the outer medium resulted in dissipation of ΔpH preformed by the H⁺-ATPase, which indicates the presence in PM of the functionally active Na⁺/H⁺ antiporter. The results are discussed with regard to involvement of the Na⁺/H⁺ antiporter and the PM H⁺-ATPase in loading Na⁺ ions into the xylem of S. altissima roots.     

Plasmalemma dicarboxylate transporter of <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> is involved in citrate and succinate influx and is modulated by pH and cations

Succinate and citrate transport into yeast (Saccharomyces cerevisiae) cells was studied by measuring substrate oxidation rates in the presence and in the absence of effective impermeable oxidation inhibitors O-palmitoyl-L-malate and 2-undecyl malonate. Linearity of the Dixon plot for 2-undecyl malonate suggests that this inhibitor blocked the rate-limiting step upon oxidation of both substrates, which was, most probably, transport of these substrates across the plasma membrane (due to inability of the inhibitor to penetrate into the membrane). This approach allowed fast (within 30–40 min) measurement of kinetic parameters of the transporter in individual samples without losing control over limiting conditions. In case of succinate transport, the limiting rate of succinate oxidation (V _max) depended on pH and increased monotonously from near-zero at pH 4.5 to the maximum level at pH 7.5. At pH 5.5, succinate and citrate transport was insensitive to the protonophore FCCP, being activated by Na⁺ ions and competitively inhibited by 2-undecyl malonate and K⁺ ions. Values of K _i for 2-undecyl malonate were similar for both substrates. These data suggest that citrate and succinate influx is mediated by a common plasma membrane transporter. This is not typical of fungi. At pH 6.5, Tris⁺, K⁺ and Na⁺ had no effect on succinate oxidation. In monosodium media pH increase was accompanied by a decrease of succinate K _m due to higher proportion of the dianionic form of the substrate. Atypical substrate specificity and mechanisms of functional activity of the dicarboxylate transporter in plasma membrane of S. cerevisiae are discussed.     

Molecular characterization and expression analysis of the Na<Superscript>+</Superscript>/H<Superscript>+</Superscript> exchanger gene family in <Emphasis Type="Italic">Medicago truncatula</Emphasis>

One important mechanism plants use to cope with salinity is keeping the cytosolic Na⁺ concentration low by sequestering Na⁺ in vacuoles, a process facilitated by Na⁺/H⁺ exchangers (NHX). There are eight NHX genes (NHX1 through NHX8) identified and characterized in Arabidopsis thaliana. Bioinformatics analyses of the known Arabidopsis genes enabled us to identify six Medicago truncatula NHX genes (MtNHX1, MtNHX2, MtNHX3, MtNHX4, MtNHX6, and MtNHX7). Twelve transmembrane domains and an amiloride binding site were conserved in five out of six MtNHX proteins. Phylogenetic analysis involving A. thaliana, Glycine max, Phaseolus vulgaris, and M. truncatula revealed that each individual MtNHX class (class I: MtNHX1 through 4; class II: MtNHX6; class III: MtNHX7) falls under a separate clade. In a salinity-stress experiment, M. truncatula exhibited ~?20% reduction in biomass. In the salinity treatment, sodium contents increased by 178 and 75% in leaves and roots, respectively, and Cl^? contents increased by 152 and 162%, respectively. Na⁺ exclusion may be responsible for the relatively smaller increase in Na⁺ concentration in roots under salt stress as compared to Cl^?. Decline in tissue K⁺ concentration under salinity was not surprising as some antiporters play an important role in transporting both Na⁺ and K ⁺ . MtNHX1, MtNHX6, and MtNHX7 display high expression in roots and leaves. MtNHX3, MtNHX6, and MtNHX7 were induced in roots under salinity stress. Expression analysis results indicate that sequestering Na⁺ into vacuoles may not be the principal component trait of the salt tolerance mechanism in M. truncatula and other component traits may be pivotal.     

Na<Superscript>+</Superscript>/H<Superscript>+</Superscript> and K<Superscript>+</Superscript>/H<Superscript>+</Superscript> antiporters AtNHX1 and AtNHX3 from <Emphasis Type="Italic">Arabidopsis</Emphasis> improve salt and drought tolerance in transgenic poplar

The tonoplast and plasma membrane localized sodium (potassium)/proton antiporters have been shown to play an important role in plant resistance to salt stress. In this study, AtNHX1 and AtNHX3, two tonoplast Na⁺(K⁺)/H⁺ antiporter encoding genes from Arabidopsis thaliana, were expressed in poplar to investigate their biological functions in the resistance to abiotic stresses in woody plants. Transgenic poplar plants expressing either gene exhibited increased resistance to both salt and water-deficit stresses. Compared to the wild type (WT) plants, transgenic plants accumulated more sodium and potassium ions in the presence of 100 mM NaCl and showed reduced electrolyte leakage in the leaves under water stress. Furthermore, the proton-translocating and cation-dependent H⁺ (Na⁺/H⁺ or K⁺/H⁺) exchange activities in the tonoplast vesicles isolated from the leaves of transgenic plants were higher than in those isolated from WT plants. Therefore, constitutive expression of either AtNHX1 or AtNHX3 genetically modified the salt and water stress tolerance of transgenic poplar plants, providing a potential tool for engineering tree species with enhanced resistance to multiple abitotic stresses.     

Sodium Conductance Activation without Inactivation in Pronase-perfused Axons

A controversy of long standing in membrane electrophysio-logy is whether the sodium ion current (I_Na) and potassium ion current (I_K) pass through the membrane in separate channels, or through a single set of channels which conduct first I_Na and then I_K. In support of the latter hypothesis it has been noted that the sodium conductance (g_Na) decline, called inactivation, proceeds with about the same time course as the potassium conductance (g_K) increase. This could mean that Na⁺ selective channels are being converted into K⁺ selective channels. The hypothesis is especially interesting because of the possibility that the carrier postulated in active transport is convertible from Na⁺ to K⁺ selectivity¹. An explicit statement of the single channel hypothesis and the means for disproving it were given by Mullins². Because a single channel could not simultaneously conduct I_Na and I_K, disproof requires that membrane conductance (g_m) be made somehow to exceed the maximum value of g_Na or g_K. We report here that inactivation of g_Na can be destroyed fairly selectively by the action from inside the axon of the unspecific proteolytic enzymes of pronase. In many cases g_m after pronase treatment is greater than maximum g_K before treatment, making untenable the single channel hypothesis.     

Check of Gene Number during the Process of rDNA Magnification

THE multiple sequences of rDNA (DNA complementary to ribosomal RNA) of the Drosophila genome are localized at the bobbed locus, located in the X chromosome, position 66 and in the short arm of the Y chromosome^1,2. Wild bobbed (bb⁺) is that locus which, without a partner, gives rise to a normal phenotype. That locus which in similar conditions is incapable of giving rise to a normal phenotype is called a bobbed mutation (bb) and contains fewer genes for rRNA. The number of genes for rRNA in different individuals can vary considerably. One mechanism for rDNA variation is unequal crossing over³. Another mechanism, described by Tartof⁴, becomes apparent when individual flies, carrying only one bobbed locus, are constructed and only if such a locus is on the X chromosome; that is, if one constructs Xbb⁺/O males (and also Xbb/O males) or Xbb⁺/X_NO- females. Such individuals show a higher rDNA content than expected from the analysis of the same locus in Xbb⁺/Xbb⁺ females or in Xbb⁺/Ybb⁺ males. The increase of rDNA in this case is not inheritable⁴.     

Effect of cumulative polymery of tomato keeping life genes

Results from works involving the study of hetero-and homozygous interaction of the keeping quality genes alc, nor, and rin in tomato plants are presented. It is shown that cumulative polymery leading to the formation of a new “long ripening” phenotype is observed in the double heterozygotes alc/alc ⁺//nor/nor ⁺, nor/nor ⁺//rin/rin ⁺, and alc/alc ⁺//rin/rin ⁺. Strong inhibition of processes of ripeness and synthesis of carotenoids occurs with nonallelic interaction in the double homozygotes alc/alc//rin/rin, nor/nor//rin/rin, and alc/alc//nor/nor. This promotes the formation of a new “non-ripening” phenotype with the absence of visual signs of ripeness, i.e., a whitish-green coloring of the fruit.     

Improved salt tolerance in a wheat stay-green mutant <Emphasis Type="Italic">tasg1</Emphasis>

Salt stress inhibited the growth of both tasg1 and wild-type (WT) wheat seedlings, but the inhibition in tasg1 plants was relatively weaker than that of WT. Compared to the WT, the chlorophyll content, thylakoid membrane polypeptides, Hill reaction activity, actual photochemical efficiency of PSII (ΦPSII), and Mg²⁺- and Ca²⁺-ATPase activities were higher in tasg1 under salt stress. At the same time, the photosynthetic activity of the tasg1 was significantly higher than that of WT. In addition, tasg1 plants displayed relatively less accumulation of reactive oxygen species and oxidative damage accompanied by higher activity of some antioxidant enzymes, and the up-regulation of antioxidant genes further demonstrated the improvement of antioxidant activity in tasg1 under salt stress. Furthermore, tasg1 plants also showed relatively weaker Na⁺ fluorescence and lower Na⁺ content, but relatively higher content of K⁺ in their roots and shoots, and then, the roots of tasg1 plants enhanced net outward Na⁺ flux and a correspondingly increased net inward K⁺ flux during salt stress. This might be associated with the relatively higher activity of H⁺-ATPase in tasg1 plants. These results suggest that the improved antioxidant competence and Na⁺/K⁺ ion homeostasis play an important role in the enhanced salinity tolerance of tasg1 plants.     

Analysis of applicability of triplet-state emission of molecular hydrogen for spectral diagnostics of a DC discharge

The applicability of emission of the N ³Λ_σ triplet states of molecular hydrogen for spectral diagnostics of the positive column of a dc glow discharge in hydrogen at translational gas temperatures of 360–600 K, specific absorbed powers of 0.8–4.25 W/cm, gas pressures of p = 0.3–15.0 Torr, reduced fields of E/N = 30–130 Td, and electron densities of n _e = 4.0 × 10⁹–6.5 × 10¹⁰ cm^–3 is analyzed by using an advanced level-based semi-empirical collisional?radiative model. It is found that secondary processes make the main contribution to the population and decay of the N ³Λ_σ = a ³Σ⁺ _g , c ³Π _u , g ³Σ⁺ _g , h ³Σ⁺ _g , and i ³Π _g triplet states. The dipole-allowed transitions e ³Σ⁺ _g → a ³Σ⁺ _g , f ³Σ⁺ _g → a ³Σ⁺ _g , g ³Σ⁺ _g and k ³Π _u → a ³Σ⁺ _g can be used for spectral diagnostics of a dc discharge within a simplified coronal model.     

Search for genes influencing the maintenance of the [<Emphasis Type="Italic">ISP</Emphasis><Superscript>+</Superscript>] prion-like antisuppressor determinant in yeast with the use of an insertion gene library

The prion-like determinant [ISP ⁺] manifests itself as an antisuppressor of certain sup35 mutations. To establish that [ISP ⁺] is indeed a new yeast prion, it is necessary to identify the gene that codes for the protein whose prion form is [ISP ⁺]. Analysis of the transformants obtained by transformation of an [ISP ⁺] strain with an insertion gene library revealed three genes controlling the [ISP ⁺] maintenance: UPF1, UPF2, and SFP1. SFP1 codes for a potentially prionogenic protein, which is enriched in Asn and Gln residues, and is thereby the most likely candidate for the [ISP ⁺] structural gene. UPF1 and UPF2 code for components of nonsense-mediated mRNA decay. The [ISP ⁺] elimination caused by UPF1 and UPF2 inactivation was reversible, and Upf1p and Upf2p were not functionally related to phosphatase Ppz1p, which influences the [ISP ⁺] manifestation. Possible mechanisms sustaining the influence of UPF1 and UPF2 on [ISP ⁺] maintenance are discussed.     

Mapping of novel salt tolerance QTL in an Excalibur × Kukri doubled haploid wheat population

Key message

Novel QTL for salinity tolerance traits have been detected using non-destructive and destructive phenotyping in bread wheat and were shown to be linked to improvements in yield in saline fields.

Abstract

Soil salinity is a major limitation to cereal production. Breeding new salt-tolerant cultivars has the potential to improve cereal crop yields. In this study, a doubled haploid bread wheat mapping population, derived from the bi-parental cross of Excalibur?×?Kukri, was grown in a glasshouse under control and salinity treatments and evaluated using high-throughput non-destructive imaging technology. Quantitative trait locus (QTL) analysis of this population detected multiple QTL under salt and control treatments. Of these, six QTL were detected in the salt treatment including one for maintenance of shoot growth under salinity (QG_(1–5).asl-7A), one for leaf Na⁺ exclusion (QNa.asl-7A) and four for leaf K⁺ accumulation (QK.asl-2B.1, QK.asl-2B.2, QK.asl-5A and QK:Na.asl-6A). The beneficial allele for QG_(1–5).asl-7A (the maintenance of shoot growth under salinity) was present in six out of 44 mainly Australian bread and durum wheat cultivars. The effect of each QTL allele on grain yield was tested in a range of salinity concentrations at three field sites across 2 years. In six out of nine field trials with different levels of salinity stress, lines with alleles for Na⁺ exclusion and/or K⁺ maintenance at three QTL (QNa.asl-7A, QK.asl-2B.2 and QK:Na.asl-6A) excluded more Na⁺ or accumulated more K⁺ compared to lines without these alleles. Importantly, the QK.asl-2B.2 allele for higher K⁺ accumulation was found to be associated with higher grain yield at all field sites. Several alleles at other QTL were associated with higher grain yields at selected field sites.

     

<Emphasis Type="Italic">OsNHX2</Emphasis>, an Na<Superscript>+</Superscript>/H<Superscript>+</Superscript> antiporter gene,can enhance salt tolerance in rice plants through more effective accumulation of toxic Na<Superscript>+</Superscript> in leaf mesophyll and bundle sheath cells

The Na⁺/H⁺ antiporters play an important role in salt tolerance in plants. However, the functions of OsNHXs in rice except OsNHX1 have not been well studied. Using the gain- and loss-of-function strategies, we studied the potential role of OsNHX2 in salt tolerance in rice. Overexpression of OsNHX2 (OsNHX2-OE) in rice showed the significant tolerance to salt stress than wild-type plants and OsNHX2 knockdown transgenic plants (OsNHX2-KD). Under salt treatments of 300-mM NaCl for 5 days, the plant fresh weights, relative water percentages, shoot heights, Na⁺ contents, K⁺ contents, and K⁺/Na⁺ ratios in leaves of OsNHX2-OE transgenic plants were higher than those in wild-type plants, while no differences were detected in roots. K⁺/Na⁺ ratios in rice leaf mesophyll cells and bundle sheath cells were higher in OsNHX2-OE transgenic plants than in wild-type plants and OsNHX2-KD transgenic plants. Our data indicate that OsNHX2 plays an important role in salt stress based on leaf mesophyll cells and bundle sheath cells and can be served in genetically engineering crop plants with enhanced salt tolerance.     

A physiological comparative study of acid tolerance of <Emphasis Type="Italic">Lactobacillus plantarum</Emphasis> ZDY 2013 and <Emphasis Type="Italic">L. plantarum</Emphasis> ATCC 8014 at membrane and cytoplasm levels

This study aimed to disclose the acid tolerance mechanism of Lactobacillus plantarum by comparing L. plantarum ZDY 2013 with the type strain L. plantarum ATCC 8014 in terms of cell membrane, energy metabolism, and amino acid metabolism. L. plantarum ZDY 2013 had a superior growth performance under acidic condition with 100-fold higher survival rate than that of L. plantarum ATCC 8014 at pH 2.5. To determine the acid tolerance physiological mechanism, cell integrity was investigated through scanning electron microscopy. The study revealed that L. plantarum ZDY 2013 maintained cell morphology and integrity, which is much better than L. plantarum ATCC 8014 under acid stress. Analysis of energy metabolism showed that, at pH 5.0, L. plantarum ZDY 2013 enhanced the activity of Na⁺/K⁺-ATPase and decreased the ratio of NAD⁺/NADH in comparison with L. plantarum ATCC 8014. Similarly, amino acid metabolism of intracellular arginine, glutamate, and alanine was improved in L. plantarum ZDY 2013. Correspondingly, the activity of arginine deiminase and glutamate decarboxylase of L. plantarum ZDY 2013 increased by 1.2-fold and 1.3-fold compared with L. plantarum ATCC 8014 in acid stress. In summary, it is demonstrated that the special physiological behaviors (integrity of cell membrane, enhanced energy metabolism, increased amino acid and enzyme level) of L. plantarum ZDY 2013 can protect the cells from acid stress.     

Overexpression of <Emphasis Type="Italic">AlTMP2</Emphasis> gene from the halophyte grass <Emphasis Type="Italic">Aeluropus littoralis</Emphasis> in transgenic tobacco enhances tolerance to different abiotic stresses by improving membrane stability and deregulating some stress-related genes

Herein, we report isolation of the AlTMP2 gene from the halophytic C4 grass Aeluropus littoralis. The subcellular localization suggested that AlTMP2 is a plasma membrane protein. In A. littoralis exposed to salt and osmotic stresses, the AlTMP2 gene was induced early and at a high rate, but was upregulated relatively later in response to abscisic acid and cold treatments. Expression of AlTMP2 in tobacco conferred improved tolerance against salinity, osmotic, H₂O₂, heat, and freezing stresses at the germination and seedling stages. Under control conditions, no growth or yield penalty were mentioned in transgenic plants due to the constitutive expression of AlTMP2. Interestingly, under greenhouse conditions, the seed yield of transgenic plants was significantly higher than that of non-transgenic (NT) plants grown under salt or drought stress. Furthermore, AlTMP2 plants had less electrolyte leakage, higher membrane stability, and lower Na⁺ and higher K⁺ accumulation than NT plants. Finally, six stress-related genes were shown to be deregulated in AlTMP2 plants relative to NT plants under both control and stress conditions. Collectively, these results indicate that AlTMP2 confers abiotic stress tolerance by improving ion homeostasis and membrane integrity, and by deregulating certain stress-related genes.