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Characterization of the gene for delta1-pyrroline-5-carboxylate synthetase and correlation between the expression of the gene and salt tolerance in Oryza sativa L.

A cDNA for delta1-pyrroline-5-carboxylate (P5C) synthetase (cOsP5CS), an enzyme involved in the biosynthesis of proline, was isolated and characterized from a cDNA library prepared from 14-day-old seedlings of Oryza sativa cv. Akibare. The deduced amino acid sequence of the P5CS protein (OsP5CS) from O. sativa exhibited 74.2% and 75.5% homology to that of the P5CS from Arabidopsis thaliana and Vigna aconitifolia, respectively. Northern blot analysis revealed that the gene for P5CS (OsP5CS) was induced by high salt, dehydration, treatment of ABA and cold treatment, while it was not induced by heat treatment. Simultaneously, accumulation of proline was observed as a result of high salt treatment in O. sativa. Moreover, the levels of expression of OsP5CS mRNA and content of proline under salt stress condition were compared between a salt-tolerant cultivar, Dee-gee-woo-gen (DGWG) and a salt-sensitive breeding line, IR28. It was observed that the expression of the P5CS gene and the accumulation of proline in DGWG steadily increased, whereas those in IR28 increased slightly.

1-Pyrroline-5-Carboxylate Dehydrogenase↗

Hydroxylamine sensitivity and salt tolerance in screening and identifying mycobacteria.

Hydroxylamine sensitivity offered no advantage over p-nitrobenzoic acid in distinguishing between tubercle bacilli and other (opportunist and non-significant) mycobacteria. It was of limited use in identification procedures. Salt tolerance assisted in the identification of strains which may be Mycobacterium triviale and M. chelonei var borstelense.

Bacteriological Techniques↗

Effect of conjugated bile salts on antibiotic susceptibility of bile salt-tolerant Lactobacillus and Bifidobacterium isolates.

Virtually every antibiotic may cause in vivo alterations in the number, level, and composition of the indigenous microbiotae. The degree to which the microbiotae are disturbed depends on many factors. Although bile may augment antibiotic activity, studies on the effect of bile on the antibiotic susceptibility of indigenous and exogenous probiotic microorganisms are lacking. It was against this background that the antibiotic susceptibility of 37 bile salt-tolerant Lactobacillus and 11 Bifidobacterium isolates from human and other sources was determined in the presence of 0.5% wt/wt oxgall (conjugated bile salts). Oxgall did not affect the intrinsic resistance of lactobacilli to metronidazole (5 microg), vancomycin (30 microg), and cotrimoxazole (25 microg), whereas it resulted in a complete loss of resistance to polymyxin B (300 microg) and the aminoglycosides gentamicin (10 microg), kanamycin (30 microg), and streptomycin (10 microg) for most strains studied (P < 0.001). Oxgall did not affect the intrinsic resistance of bifidobacteria to metronidazole and vancomycin, whereas polymyxin B and co-trimoxazole resistance was diminished (P < 0.05) and aminoglycoside resistance was lost (P < 0.001). Seven lactobacilli, but no bifidobacteria strain, showed unaltered intrinsic antibiotic resistance profiles in the presence of oxgall. Oxgall affected the extrinsic susceptibility of lactobacilli and bifidobacteria to penicillin G (10 microg), ampicillin (10 microg), tetracycline (30 microg), chloramphenicol (30 microg), erythromycin (15 microg), and rifampicin (5 microg) in a source- and strain-dependent manner. Human strain-drug combinations of lactobacilli (P < 0.05) and bifidobacteria (P < 0.01) were more likely to show no change or decreased susceptibility compared with other strain-drug combinations. The antimicrobial activity spectra of polymyxin B and the aminoglycosides should not be considered limited to gram-negative bacteria but extended to include gram-positive genera of the indigenous and transiting microbiotae in the presence of conjugated bile salts. Those lactobacilli (7 of 37) that show unaltered intrinsic and diminished extrinsic antibiotic susceptibility in the presence of oxgall may possess greater upper gastrointestinal tract transit tolerance in the presence of antibiotics.

Anti-Bacterial Agents↗

Bioremediation and toxicity determination of natural seawater polluted with weathered crude oil by salt-tolerant consortia in a SBR.

The aim of this research was to compare the bioremediation treatment of seawater polluted with two different concentrations of weathered crude oil (0.2 for experiment (a) and 1.14% for experiment (b), v/v) by salt-tolerant consortia enriched from the sludge of a refinery wastewater treatment facility, in a SBR. The use of a commercial bioremediation stimulant (S200) was also evaluated as an alternative to the traditional nitrogen and phosphorus supplement.

Bacteria↗

Genome-Wide Identification of the Soybean GH5 Gene Family and Functional Analysis of GmGH5-22 in Salt Tolerance.

Plant GH5 family genes function in both cell wall biosynthesis and stress responses. However, comprehensive studies on GH5 genes in the soybean remain limited. Here, we identified 28 GmGH5 genes from the soybean genome. Phylogenetic analysis assigned these genes to three subfamilies (I-III), with no representatives in subfamily IV. The GmGH5 family harbors 15 conserved motifs, which are largely similar within subfamilies but differ across subfamilies. Additionally, exon-intron structures (2-7 introns) exhibit clade-specific patterns, with members within the same clade sharing similar intron numbers and lengths, whereas distinct clades show some variation. The promoter regions of GmGH5 genes contained various cis-acting regulatory elements associated with stress responses and developmental processes. Transcriptome-based expression profiling revealed distinct tissue-specific expression patterns of GmGH5 genes. RT-qPCR further confirmed their differential expression under salt, alkaline, cold, and drought stresses, especially a significant increase in GmGH5-22 expression under salt stress (approximately 22-fold at 6 h, **** p < 0.0001). Furthermore, GmGH5-22 was highly expressed in roots, and transient expression in tobacco leaves showed its peripheral localization, which aligns with its predicted extracellular localization, suggesting that GmGH5-22 is highly likely localized to the cell wall. Overexpression of GmGH5-22 in soybean hairy roots significantly improved tolerance to salt stress. These findings establish a foundation for functional characterization of GmGH5 genes and provide viable targets for molecular breeding to enhance salt tolerance in soybeans.

GH5 family↗

Measurement of biodegradable substances using the salt-tolerant yeast Arxula adeninivorans for a microbial sensor immobilized with poly(carbamoyl)sulfonate (PCS). Part II: Application of the novel biosensor to real samples from coastal and island regions.

A microbial sensor for rapid measurement of the amount of biodegradable substances based on the salt-tolerant yeast Arxula adeninivorans LS3 has been developed especially for coastal and island regions. Our parameter, the so-called sensorBOD, that is available after only a few minutes, agrees with the 5-day value for the biochemical oxygen demand (BOD5) very well. We have employed the Arxula sensor in the short-time estimation and supervision of the BOD of both domestic and industrial wastewater with high salinity. The novel sensor makes it possible to monitor the different types of wastewater rapidly without pretreatment, and it can be used for an active process control of sewage treatment works. Compared to a commercially available sensor, the novel sensor achieves better agreement between sensorBOD and BOD5 measurements with salt containing samples.

Ascomycota↗

Analysis of salt-stress-inducible ESTs isolated by PCR-subtraction in salt-tolerant rice.

To clarify the mechanisms of stress tolerance in rice and to search for rice genes associated with these mechanisms, we analyzed genes induced by a high salinity treatment using the PCR-subtractive hybridization method (PCR-subtraction). Seedlings of the salt-tolerant rice cultivar Dee-geo-woo-gen (DGWG) were either treated with 250 mM NaCl for 5 h or left untreated, and PCR-subtraction was then performed using the untreated (control) plants as a driver and the NaCl-treated plants as a tester. We obtained 384 clones of tester-specific cDNAs as salt-inducible candidates. Northern analysis performed with the cDNA fragments showed that 65 clones had been induced by the NaCl treatment. Sequence analysis and database searching indicated that these clones have homology to proteins functional for detoxification, stress response, and signal transduction in plants. Of these clones, 22% coded for unknown proteins and 12% gave no hits. We selected eight clones from each functional category and analyzed their expression pattern in DGWG. For temporal analysis, seedlings were treated with H(2)O or 250 mM NaCl for 0, 0.5, 1, 2, 5, 10 or 24 h. Different patterns of transcript regulation were found. For the analysis of expression in response to various types of stress and abscisic acid (ABA) treatments, seedlings were treated for 5 h or 10 h with H(2)O, dehydration, cold (4 degrees C), heat (40 degrees C), mannitol, ABA, or wounding. All clones were strongly up-regulated by osmotic stress (dehydration and mannitol) and the ABA treatment.

Abscisic Acid↗

Isolation of a salt tolerant pleomorphic Klebsiella strain from a case of diabetic periodontitis.

An unusual Klebsiella strain was isolated from a deep periodontal pocket of a diabetic patient. According to its biochemical reactions the new strain differed from Klebsiella pneumoniae and other described biotypes. In addition, the new isolate was very salt tolerant; in the presence of 7.5% sodium chloride the bacterium changed into a spirillum-like form, with highly pleomorphic filaments, and reverted immediately to the short rod form at lower sodium chloride concentrations or in the absence of sodium chloride. Optimum growth was observed in the presence of 2% sodium chloride. A wide range of carbohydrates was fermented with a large amount of gas production. It appears that the adaptation of the isolated Klebsiella strain to sodium chloride allows it to enter the ecological niche of periodontal lesions.

Anti-Bacterial Agents↗

Exogenous proline and glycinebetaine increase NaCl-induced ascorbate-glutathione cycle enzyme activities, and proline improves salt tolerance more than glycinebetaine in tobacco Bright Yellow-2 suspension-cultured cells.

Up-regulation of the antioxidant system provides protection against NaCl-induced oxidative damage in plants. Antioxidants and activity of enzymes involved in the ascorbate-glutathione (ASC-GSH) cycle in tobacco Bright Yellow-2 (BY-2) were investigated to assess the antioxidant protection offered by exogenous proline and glycinebetaine (betaine from now on) against salt stress using cells grown in suspension culture. Reduced ascorbate (ASC) was detected in BY-2 cells but dehydroascorbate (DHA) was not. Large quantities of a reduced form of glutathione (GSH) and smaller quantities of an oxidized form of glutathione (GSSG) were detected in BY-2 cells. Salt stress significantly reduced the contents of ASC and GSH as well as activities of ASC-GSH cycle enzymes such as ascorbate peroxidase (APX), monodehydroascorbate reductase (MDHAR), dehydroascorbate reductase (DHAR), and glutathione reductase (GR). Exogenous proline or betaine increased the activities of all enzymes except MDHAR involved in NaCl-induced ASC-GSH cycle. Levels of ASC and GSH in BY-2 cells under salt stress were lower in the presence of proline or betaine than in the absence of proline or betaine whereas there was no difference in redox status. Proline proved more effective than betaine in maintaining the activity of enzymes involved in NaCl-induced ASC-GSH cycle. Neither proline nor betaine had any direct protective effect on NaCl-induced enzyme activity involved in the antioxidant system; however, both improved salt tolerance by increasing enzyme activity. The present study, together with our earlier findings [Hoque MA, Okuma E, Banu MNA, Nakamura Y, Shimoishi Y, Murata Y. Exogenous proline mitigates the detrimental effects of salt stress more than exogenous betaine by increasing antioxidant enzyme activities. J Plant Physiol 2006;164:553-61.], suggests that proline offered greater protection against salt stress than betaine did because proline was more effective in increasing the activity of enzymes involved in the antioxidant system.

Antioxidants↗

[Construction of fused osmoregulation proBA gene from a salt-tolerant mutant of Bacillus subtilis and its influence on the osmotolerance of Escherichia coli].

The osmoregulation proB and proA genes from Bacillus subtilis 93151 are overlapping genes, which encode two proteins ProB and ProA. A restriction enzyme site was inserted in the overlapping region of proB and proA genes from a salt-tolerant mutant of B. subtilis 93151, and a fusion gene was constructed by cloning proB and proA genes respectively. SDS-PAGE analysis showed that a novel protein with molecular mass of 85 kD was observed. When expressed in E. coli, enhanced intracellular concentrations of free proline and osmotolerance of the strain carrying the fusion gene were observed, compared with the control host cell harbouring a plasmid encoding the separate ProB and ProA.

Adaptation, Physiological↗

The WD-40 repeat motif of Lgl tumor suppressor proteins associated with salt tolerance and temperature sensitivity.

We have recently identified mammalian homologues of lethal giant larvae (Lgl) tumor suppressor gene, rat Rgl-1 and bovine Bgl-1, and demonstrated that they can complement yeast double mutants lacking Sop1 and Sop2, yeast homologues of Lgl. These gene products are capable of regulating cellular viability in restrictive salt and temperature environments. Since Lgl family members contain the WD-40 repeat motif, we investigated its cellular functions using mouse homologue Mgl-1 in the absence of Sop1 and Sop2 in yeasts by complementation. Interestingly, mutant forms of Mgl-1 at the conserved glycine at position 450 and aspartic acid at position 453 in the most conserved WD-40 repeat motif were not able to complement, indicating that these amino acids are critical for regulating salt tolerance and temperature sensitivity in yeast. These results shed light on the important regulation of cytoskeletal complex for cellular polarity within eukaryotic cells.

Adaptation, Physiological↗

A novel mechanism of ion homeostasis and salt tolerance in yeast: the Hal4 and Hal5 protein kinases modulate the Trk1-Trk2 potassium transporter.

The regulation of intracellular ion concentrations is a fundamental property of living cells. Although many ion transporters have been identified, the systems that modulate their activity remain largely unknown. We have characterized two partially redundant genes from Saccharomyces cerevisiae, HAL4/SAT4 and HAL5, that encode homologous protein kinases implicated in the regulation of cation uptake. Overexpression of these genes increases the tolerance of yeast cells to sodium and lithium, whereas gene disruptions result in greater cation sensitivity. These phenotypic effects of the mutations correlate with changes in cation uptake and are dependent on a functional Trk1-Trk2 potassium transport system. In addition, hal4 hal5 and trk1 trk2 mutants exhibit similar phenotypes: (i) they are deficient in potassium uptake; (ii) their growth is sensitive to a variety of toxic cations, including lithium, sodium, calcium, tetramethylammonium, hygromycin B, and low pH; and (iii) they exhibit increased uptake of methylammonium, an indicator of membrane potential. These results suggest that the Hal4 and Hal5 protein kinases activate the Trk1-Trk2 potassium transporter, increasing the influx of potassium and decreasing the membrane potential. The resulting loss in electrical driving force reduces the uptake of toxic cations and improves salt tolerance. Our data support a role for regulation of membrane potential in adaptation to salt stress that is mediated by the Hal4 and Hal5 kinases.

Biological Transport↗

Natural selection for salt tolerance quantitative trait loci (QTLs) in wild sunflower hybrids: implications for the origin of Helianthus paradoxus, a diploid hybrid species.

For a new diploid or homoploid hybrid species to become established, it must diverge ecologically from parental genotypes. Otherwise the hybrid neospecies will be overcome by gene flow or competition. We initiated a series of experiments designed to understand how the homoploid hybrid species, Helianthus paradoxus, was able to colonize salt marsh habitats, when both of its parental species (H. annuusxH. petiolaris) are salt sensitive. Here, we report on the results of a quantitative trait locus (QTL) analysis of mineral ion uptake traits and survivorship in 172 BC2 hybrids between H. annuus and H. petiolaris that were planted in H. paradoxus salt marsh habitat in New Mexico. A total of 14 QTLs were detected for mineral ion uptake traits and three for survivorship. Several mineral ion QTLs mapped to the same position as the survivorship QTLs, confirming previous studies, which indicated that salt tolerance in Helianthus is achieved through increased Ca uptake, coupled with greater exclusion of Na and related mineral ions. Of greater general significance was the observation that QTLs with effects in opposing directions were found for survivorship and for all mineral ion uptake traits with more than one detected QTL. This genetic architecture provides an ideal substrate for rapid ecological divergence in hybrid neospecies and offers a simple explanation for the colonization of salt marsh habitats by H. paradoxus. Finally, selection coefficients of +0.126, -0.084 and -0.094 for the three survivorship QTLs, respectively, are sufficiently large to account for establishment of new, homoploid hybrid species.

Adaptation, Biological↗

Alfin1 transcription factor overexpression enhances plant root growth under normal and saline conditions and improves salt tolerance in alfalfa.

Plant root development is an essential determinant of plant growth and crop yield that could be enhanced by induced changes in the expression of root-specific regulatory factors. We reported previously that Alfin1 binds DNA in a sequence-specific manner and that Alfin1 overexpression in transgenic alfalfa (Medicago sativa L.) enhances expression of the salt-inducible MsPRP2 gene in roots, suggesting that Alfin1 functions to regulate gene expression in roots. Here we show that Alfin1 is an essential gene for root growth and that its overexpression in transgenic plants confers a many-fold increase in root growth under normal and saline conditions. Alfin1-binding sites occur in promoters of genes expressed in roots of a wide variety of plant species and we propose that it is a general root growth regulator. Even though Alfin1 overexpression was under the control of the CaMV 35S promoter, plant shoot growth was not adversely affected. We show further that introduction of the Alfin1 transgene in plants confers a dominant characteristic that significantly increases plant growth and salt tolerance.

Caulimovirus↗

A putative plasma membrane cation/proton antiporter from soybean confers salt tolerance in Arabidopsis.

Cation transport is thought to be an important process for ion homeostasis in plant cells. Here, we report that a soybean putative cation/proton antiporter GmCAX1 may be a mediator of this process. GmCAX1 is expressed in all tissues of the soybean plants but at a lower level in roots. Its expression was induced by PEG, ABA, Ca(2+), Na(+) and Li(+) treatments. The GmCAX1-GFP fusion protein was mainly localized in plasma membrane of the transgenic Arabidopsis plant cells and onion epidermal cells. Transgenic Arabidopsis plants overexpressing GmCAX1 accumulated less Na(+), K(+), and Li(+), and were more tolerant to elevated Li(+) and Na(+) levels during germination when compared with the controls. These results suggest that GmCAX1 may function as an antiporter for Na(+), K(+) and Li(+). Modulation of this antiporter may be beneficial for regulation of ion homeostasis and thus plant salt tolerance.

Amino Acid Sequence↗

The Candida albicans antiporter gene CNH1 has a role in Na+ and H+ transport, salt tolerance, and morphogenesis.

The isolation and functional characterization of a Candida albicans Na+/H+ antiporter gene, CNH1, is reported here. The gene encodes a protein of 840 amino acids that exhibits high levels of similarity in sequence, size, and structural and functional domains to a group of known Na+/H+ antiporters of fungi. The CNH1 gene is able to functionally complement the salt-sensitivity of a Saccharomyces cerevisiae ena1 nha1 mutant, and mutations of two conserved aspartate residues to asparagines in the putative Na+-binding site abolish this activity. Deletion of CNH1 results in retardation of growth and a highly elongated morphology in a significant fraction of cells under conditions that normally support yeast growth. These results indicate that CNH1 has a role in Na+ and H+ transport, salt-tolerance, and morphogenesis.

Adenosine Triphosphatases↗

Physiological basis of salt tolerance in plants.

For most plants, and under most field conditions, osmotic effects of salinity greatly predominate in restricting growth and yields. In certain cases, however, specific ion effects may be decisive. These may involve either nutrition, as in calcium deficiency in some lettuce varieties, tomato, and bell peppers, or direct toxicity (chloride or sodium toxicity, or both) in tree and vine crops. Rootstocks, or varieties that restrict the uptake of toxic ions, increase the salt tolerance of some susceptible fruit crops. Salinity-induced nutritional imbalance can, in some cases, be corrected by selecting better adapted varieties and in others by the use of foliar nutrient sprays. Recent evidence indicates simple single-gene control over uptakes of chloride and sodium, but the more general osmotic effects appear to be complex and under multigenic control.

Biological Transport↗