Intracellular Na and K distribution in Debaryomyces hansenii. Cloning and expression in Saccharomyces cerevisiae of DhNHX1

Debaryomyces hansenii is a salt-tolerant yeast that contains high amounts of internal Na(+). Debaryomyces hansenii kept more sodium than Saccharomyces cerevisiae in both the cytoplasm and vacuole when grown under a variety of NaCl concentrations. These results indicate a higher tolerance of Debaryomyces to high internal Na(+), and, in addition, suggest the existence of a transporter driving Na(+) into the vacuole. Moreover, a gene encoding a Na(+) (K(+))/H(+) antiporter from D. hansenii was cloned and sequenced. The gene, designated DhNHX1, exhibited significant homology with genes of the NHE/NHX family. DhNHX1 expression was induced neither at low pH nor by extracellular NaCl. A mutant of S. cerevisiae lacking its own Na(+) transporters (ena1-4Delta nha1 Delta nhx1 Delta), when transformed with DhNHX1, partially recovered cation tolerance as well as the ability to accumulate Na(+) and K(+) into the vacuole. Our analysis provides evidence that DhNhx1p transports Na(+) (and K(+)) into the vacuole and that it can play an important role in ion homeostasis and salt tolerance.     

Proteomic changes in response to potassium starvation in the extremophilic yeast Debaryomyces hansenii

In this work, we performed for the first time a proteomic approach to the processes induced by long-term potassium starvation in the halotolerant yeast Debaryomyces hansenii. The proteomic profile under this ionic stress conditions shows that important changes in gene expression take place as an adaptive response. We found a significant protein expression repression as well as metabolic changes such as the inhibition of the upper part of the glycolysis, the amino acid synthesis, and the Krebs cycle. On the other hand, genes related to stress responses, protein degradation, and sterols synthesis were upregulated in response to potassium deprivation. The findings in this study provide important information about how this particular yeast copes with ionic stress at molecular levels, which might further enrich the global understanding of salt tolerance processes in eukaryal systems and moreover highlighting the importance of the 'omics' approaches as a complement to the classical physiological studies.     

Arabidopsis sos1 mutant in a salt-tolerant accession revealed an importance of salt acclimation ability in plant salt tolerance

An analysis of the salinity tolerance of 354 Arabidopsis thaliana accessions showed that some accessions were more tolerant to salt shock than the reference accession, Col-0, when transferred from 0 to 225 mM NaCl. In addition, several accessions, including Zu-0, showed marked acquired salt tolerance after exposure to moderate salt stress. It is likely therefore that Arabidopsis plants have at least two types of tolerance, salt shock tolerance and acquired salt tolerance. To evaluate a role of well-known salt shock tolerant gene SOS1 in acquired salt tolerance, we isolated a sos1 mutant from ion-beam-mutagenized Zu-0 seedlings. The mutant showed severe growth inhibition under salt shock stress owing to a single base deletion in the SOS1 gene and was even more salt sensitive than Col-0. Nevertheless, it was able to survive after acclimation on 100 mM NaCl for 7 d followed by 750 mM sorbitol for 20 d, whereas Col-0 became chlorotic under the same conditions. We propose that genes for salt acclimation ability are different from genes for salt shock tolerance and play an important role in the acquisition of salt or osmotic tolerance.     

Sterol content, fatty acid composition of phospholipids, and permeability of labeled ethylene glycols in relation to salt-tolerance of yeasts

Two yeasts, the salt-tolerant Debaryomyces hansenii and the non-tolerant Saccharomyces cerevisiae were grown in basal media (4 m M NaCl) and also a high salinities that produced a similar salt stress in the two species in terms of growth rate reduction (i.e., 1.4 M NaCl for S. cerevisae and 2.7 M NaCl for D. hansenii ). A study was made of the sterol content, the fatty acid composition of the phospholipids, and the permeation of a series of tritiated ethylene glycols of graded molecular weights. On the basis of cell dry weight the amount of total and free sterols increased in both species when cultured at high salinity. Irrespective of growth medium salinity, the molar ratio of free sterols to phospholipids was higher in D. hansenii than in S. cerevisiae . Increased salinity produced only minor changes in the fatty acid composition of the phospholipids in D. hansenii , whereas in S. cerevisiae there was a marked decrease of linolenic acid with a concomitant increase of linoleic acid.
In both yeasts there was an energy linked component in the uptake of ethylene glycol, which component could be inhibited by sodium azide and N -ethylmaleimide. The passive permeability for ethylene-, diethylene- and triethylene glycol increased for both species at increased salinity. This increase was more pronounced for S. cerevisiae than for D. hansenii . Polyethylene glycol of M , 200 as well as higher polyethylene glycols appeared to be excluded or very slowly admitted by the yeasts.     

Review article. Molecular biology of salt tolerance in the context of whole-plant physiology

The halobacteria are the only organisms that are tolerant of salinity at the molecular level. All other bacteria, all fungi, all plants, and all animals avoid the need for salt tolerance for most of their macromolecules by maintaining defined and conserved conditions in the cytoplasm. These conditions favour potassium over sodium, the limitation of total inorganic ion activity, and the supplementation of this where necessary with organic solutes which are metabolically neutral osmolytes that may also be osmoprotectant. The salt tolerance of an organism depends upon the range of external salinity over which it is able to sustain these conditions in the cytoplasm. There is substantial and increasing knowledge of the molecular biology and molecular genetics of the processes of ion and organic solute transport, solute synthesis, and compartmentation that underpin cell-based tolerance. Much of recent research focuses on the identification of genes and gene products that affect cell-based tolerance, commonly derived from single-cell models. There is commonly the implicit or explicit assumption that incorporation of these genes will benefit the salt tolerance of food crop species. While this essential experimental approach is giving enormous insight there should not be rash or premature expectations. The unique and overriding consideration for the salinity tolerance of terrestrial plants is the net flux of water due to transpiration and so resides at a higher level of organization. Processes that are advantageous to a single cell in an aqueous medium may be lethal to a cell in a leaf in the air. The likely impact of single structural-gene changes in ion and solute transport upon co-ordinated plant response is probably over-estimated, and recent views consider regulatory processes and multiple gene transfers. While the technical ability for plant transformation increases daily, the practicality of using transgenic plants in complex breeding programmes seems rarely to be given enough thought. If intervention at the molecular level is to lead to salt-tolerant crop plants than it will be essential to view this in the contexts of whole plants and of plant breeding. Recent indications that a relatively small number of quantitative trait loci (QTL) may govern complex physiological characters offer the most hope for the future.     

Regulation of the potassium to sodium ratio and of the osmotic potential in relation to salt tolerance in yeasts

By using the isotope pairs (22)Na-(24)Na and (42)K-(86)Rb, the uptake and retention of Na and K was studied in the salt-tolerant Debaryomyces hansenii and in the less tolerant Saccharomyces cerevisiae at NaCl levels of 4 mm and 0.68, 1.35, and 2.7 m in the medium. The ratio of K to Na is much higher in the cells than in the media, and higher in D. hansenii than in S. cerevisiae under comparable conditions. The difference between the two species is due to a better Na extrusion and a better uptake of K in D. hansenii. The kinetics of ion transport show that at about the time when extrusion of Na could be demonstrated in D. hansenii, K-Rb previously lost to an easily washable compartment of the cells was reabsorbed in both organisms. More H(+) was given off from S. cerevisiae than from D. hansenii in the course of these events. The findings fit the working hypothesis tested, which regards salt tolerance as partly dependent on the ability to mobilize energy to extrude Na from the cells and to take up K. The volume changes in S. cerevisiae are greater and are more slowly overcome than those in D. hansenii. The total salt level of the cells is not sufficient to counteract the osmotic potential of the medium, so that additional osmoregulatory mechanisms must be involved in determining halotolerance.     

Species-specific interactions between algal endosymbionts and coral hosts define their bleaching response to heat and light stress

The impacts of warming seas on the frequency and severity of bleaching events are well documented, but the potential for different Symbiodinium types to enhance the physiological tolerance of reef corals is not well understood. Here we compare the functionality and physiological properties of juvenile corals when experimentally infected with one of two homologous Symbiodinium types and exposed to combined heat and light stress. A suite of physiological indicators including chlorophyll a fluorescence, oxygen production and respiration, as well as pigment concentration consistently demonstrated lower metabolic costs and enhanced physiological tolerance of Acropora tenuis juveniles when hosting Symbiodinium type C1 compared with type D. In other studies, the same D-type has been shown to confer higher thermal tolerance than both C2 in adults and C1 in juveniles of the closely related species Acropora millepora. Our results challenge speculations that associations with type D are universally most robust to thermal stress. Although the heat tolerance of corals may be contingent on the Symbiodinium strain in hospite, our results highlight the complexity of interactions between symbiotic partners and a potential role for host factors in determining the physiological performance of reef corals.     

野生大豆耐盐性研究进展

野生大豆对于拓宽大豆种质遗传基础和丰富大豆种质基因库具有重要意义.该文从野生大豆的资源概况及优良性状、耐盐机理和利用野生大豆提高栽培大豆耐盐性等三个方面,对近年来国内外有关野生大豆耐盐性的解剖结构、生理基础、分子生物学基础等方面的研究进展进行了系统综述,并提出野生大豆通过茎叶表皮上的"腺体"及对Na+和Cl-的排斥性,实现对盐渍环境的颉颃作用.较强的抗氧化能力、大豆异黄酮代谢和耐盐基因也是其适应盐渍环境的重要原因.今后应对野生大豆耐盐机理的遗传学基础进行深入研究,并通过种群保护以保障野生大豆的发掘鉴定和创新利用.     

Developing salt tolerant plants in a new century: a molecular biology approach

Soil salinity is a major abiotic stress in plant agriculture strongly, influencing plant productivity world-wide. Classical breeding for salt tolerance in crop plants has been attempted to improve field performance without success. Therefore, an alternative strategy is to generate salt tolerant plants through genetic engineering. Several species and experimental approaches have been used in order to identify those genes that are important for salt tolerance. Due to high level of salt tolerance, halophytes are good candidates to identify salt tolerance genes. However, other species such as yeast and glycophytes have also been employed. Three approaches are commonly used to identify genes important for salt tolerance. The first approach is to identify genes involved in processes known to be critical for salt tolerance (osmolyte synthesis, ion homeostasis, etc.). The second approach is to identify genes whose expression is regulated by salt stress. This is relatively simply and applicable to any plant species. Genetic amenability of some species allows the third approach, which consists in the identification of salt tolerance determinants based on functionality. At the moment, there is a large number of reports in the literature claiming that plants with increased salt tolerance have been obtained. The main problem is that different plant species, stage of development, organs, promoters and salt conditions used it is difficult to compare the degree of salt tolerance conferred by different genes. In this review, we discuss progress made towards understanding the molecular elements involved in salt stress responses that have been used in transgenic approaches to improve salt tolerance.     

Improving salinity tolerance in crop plants: a biotechnological view

Salinity limits the production capabilities of agricultural soils in large areas of the world. Both breeding and screening germplasm for salt tolerance encounter the following limitations: (a) different phenotypic responses of plants at different growth stages, (b) different physiological mechanisms, (c) complicated genotype × environment interactions, and (d) variability of the salt-affected field in its chemical and physical soil composition. Plant molecular and physiological traits provide the bases for efficient germplasm screening procedures through traditional breeding, molecular breeding, and transgenic approaches. However, the quantitative nature of salinity stress tolerance and the problems associated with developing appropriate and replicable testing environments make it difficult to distinguish salt-tolerant lines from sensitive lines. In order to develop more efficient screening procedures for germplasm evaluation and improvement of salt tolerance, implementation of a rapid and reliable screening procedure is essential. Field selection for salinity tolerance is a laborious task; therefore, plant breeders are seeking reliable ways to assess the salt tolerance of plant germplasm. Salt tolerance in several plant species may operate at the cellular level, and glycophytes are believed to have special cellular mechanisms for salt tolerance. Ion exclusion, ion sequestration, osmotic adjustment, macromolecule protection, and membrane transport system adaptation to saline environments are important strategies that may confer salt tolerance to plants. Cell and tissue culture techniques have been used to obtain salt tolerant plants employing two in vitro culture approaches. The first approach is selection of mutant cell lines from cultured cells and plant regeneration from such cells (somaclones). In vitro screening of plant germplasm for salt tolerance is the second approach, and a successful employment of this method in durum wheat is presented here. Doubled haploid lines derived from pollen culture of F₁ hybrids of salt-tolerant parents are promising tools to further improve salt tolerance of plant cultivars. Enhancement of resistance against both hyper-osmotic stress and ion toxicity may also be achieved via molecular breeding of salt-tolerant plants using either molecular markers or genetic engineering.     

Physiological and molecular mechanisms of plant salt tolerance

Salt tolerance is an important economic trait for crops growing in both irrigated fields and marginal lands. The plant kingdom contains plant species that possess highly distinctive capacities for salt tolerance as a result of evolutionary adaptation to their environments. Yet, the cellular mechanisms contributing to salt tolerance seem to be conserved to some extent in plants although some highly salt-tolerant plants have unique structures that can actively excrete salts. In this review, we begin by summarizing the research in Arabidopsis with a focus on the findings of three membrane transporters that are important for salt tolerance: SOS1, AtHKT1, and AtNHX1. We then review the recent studies in salt tolerance in crops and halophytes. Molecular and physiological mechanisms of salt tolerance in plants revealed by the studies in the model plant, crops, and halophytes are emphasized. Utilization of the Na⁺ transporters to improve salt tolerance in plants is also summarized. Perspectives are provided at the end of this review.     

Comparative roles of brassinosteroids and polyamines in salt stress tolerance

Salt stress is among the major abiotic stresses that adversely affect the global crop production and its adverse impacts are getting more serious in the regions where saline water is used for irrigation. It induces reactive oxygen species, alters the activity of antioxidant system and adversely affects the process of photosynthesis. Various strategies have been employed to mitigate the deleterious effects of salt stress. Presently, the recommended strategies to overcome the adverse effects of salt stress include the use of tolerant cultivars, ameliorative water management and diverse cultural practices. However, none of these approaches have been found to be fully effective under salt stress conditions. An alternative and technically simpler approach to induce salt stress tolerance is the exogenous application of plant growth regulators (PGRs). This technique has gained significant importance during the past decade. PGRs have been implicated to regulate a wide range of metabolic and physiological activities in plants, ranging from cell division and organogenesis to protection against biotic and abiotic stresses. One of the important factors for enhanced plant productivity by PGRs is their efficiency to overcome the salt-induced stress conditions. Recent findings on the effects of brassinosteroids and polyamines on the salt stress tolerance of crops open new avenues to address the salinity problems. This review enlightens the role of brassinosteroids and polyamines in different plant processes like their role in regulation of photosynthesis, antioxidant systems and other related aspects, thereby improving overall performance of plants.     

碱蓬属植物耐盐机理研究进展

碱蓬属(Suaeda)植物是一类典型的真盐生植物,属于重要的盐生植物资源,全球广泛分布.人们已经对20种碱蓬属植物进行了观察和盐胁迫实验,研究了不同器官或组织的生理生化特征及其对盐胁迫的反应,并基于这些研究分析了盐胁迫的应答机制.叶片肉质化、细胞内离子区域化、渗透调节物质增加和抗氧化系统能力增强是碱蓬属植物响应和适应盐胁迫的重要方式和途径.但迄今为止的研究工作尚有一定的局限性,主要包括:研究工作主要集中在植物地上部分,而对植物地下部分的研究较少;多是少数生物学指标或生理学现象的单独观察,而缺乏对生理代谢过程的整体和综合分析;针对某种碱蓬的独立分析较多,而与近缘种的比较研究较少;植物对中性盐胁迫的反应研究较多,而对碱性盐的研究较少.为进一步系统阐明碱蓬属植物的耐盐机制,今后的工作应注重碱蓬属植物响应和适应盐胁迫的信号网络和调控机制研究,基于系统生物学研究思路,采用现代组学技术探索该属植物响应盐胁迫的由复杂信号网络调控的特殊生理特征和特异代谢途径.     

Membrane phospholipids, lipoxidative damage and molecular integrity: A causal role in aging and longevity

Nonenzymatic molecular modifications induced by reactive carbonyl species (RCS) generated by peroxidation of membrane phospholipids acyl chains play a causal role in the aging process. Most of the biological effects of RCS, mainly α,β-unsaturated aldehydes, di-aldehydes, and keto-aldehydes, are due to their capacity to react with cellular constituents, forming advanced lipoxidation end-products (ALEs). Compared to reactive oxygen and nitrogen species, lipid-derived RCS are stable and can diffuse within or even escape from the cell and attack targets far from the site of formation. Therefore, these soluble reactive intermediates, precursors of ALEs, are not only cytotoxic per se, but they also behave as mediators and propagators of oxidative stress and cellular and tissue damage. The consequent loss-of-function and structural integrity of modified biomolecules can have a wide range of downstream functional consequences and may be the cause of subsequent cellular dysfunctions and tissue damage. The causal role of ALEs in aging and longevity is inferred from the findings that follow: a) its accumulation with aging in several tissues and species; b) physiological interventions (dietary restriction) that increase longevity, decrease ALEs content; c) the longer the longevity of a species, the lower is the lipoxidation-derived molecular damage; and finally d) exacerbated levels of ALEs are associated with pathological states.     

Debaryomyces singareniensis sp. nov., a novel yeast species isolated from a coal mine soil in India

Three strains (AP19, AP19-4 and AP19-6) of a novel yeast species were isolated from soil from the Singareni coal mines, Andhra Pradesh, India. They were morphologically, physiologically and phylogenetically identical and produced one to four spherical ascospores per ascus. Phylogenetic analysis using the D1/D2 variable domain of the large-subunit rRNA gene indicated that the closest relative of these strains is Debaryomyces etchellsii (2.6% divergence). Other species related to these strains are D. mycophilus (5.1% divergence) and species of the D. hansenii cluster (4.9-5.6% divergence). The novel species differs by 20 and 15 physiological tests from D. etchellsii and D. mycophilus, respectively. Phylogenetic analysis of the internal transcribed spacer (ITS) region also indicated that strains of the new species are related to D. etchellsii (7.7% divergence), followed by species of the D. hansenii cluster (9-10% divergence). In the small-subunit rRNA gene sequences, they differed from D. etchellsii by seven substitutions and one insertion or deletion of a base in a sequence (indel) and from D. mycophilus by 17 substitutions and 1 indel. The physiological, biochemical and molecular data suggest that these strains belong to a novel species, for which we propose the name Debaryomyces singareniensis sp. nov. The type strain of AP19(T) (=MTCC 7061(T)=CBS 10405(T)). The Mycobank number of the new species is MB510046.     

Genes from Debaryomyces hansenii increase salt tolerance in Saccharomyces cerevisiae W303

The yeast Debaryomyces hansenii has been chosen as a model for molecular studies of tolerance to NaCl. A gene library was built and transformants of Saccharomyces cerevisiae W303 containing genes from D. hansenii were selected for their ability to grow in the presence of high concentrations of NaCl and/or low concentrations of KCl. In three of these transformants 500 mM NaCl improved growth at pH 7.6 like in D. hansenii but not in S. cerevisiae. One of the plasmids restored growth at 50 microM KCl and K(+) uptake in a mutant of S. cerevisiae lacking genes that encode K(+) transporters.     

Salt tolerance in crop plants: new approaches through tissue culture and gene regulation

Recent approaches to study of salinity tolerance in crop plants have ranged from genetic mapping to molecular characterization of gene products induced by salt/drought stress. Transgenic plant design has allowed to test the effects of overexpression of specific prokaryotic or plant genes that are known to be up-regulated by salt/drought stress. This review summarizes current progress in the field in the context of adaptive metabolic and physiological responses to salt stress and their potential role in long term tolerance. Specifically considered are gene activation by salt, in view of proposed avenues for improved salt tolerance and the need to ascertain the additional influences of developmental regulation of such genes. Discussion includes the alternate genetic strategy we have pursued for improving salinity tolerance in alfalfa (Medicago sativa L.) and rice (Oryza sativa L.). This strategy combines single-step selection of salt-tolerant cells in culture, followed by regeneration of salt-tolerant plants and identification of genes important in conferring salt tolerance. We have postulated that activation or improved expression of a subset of genes encoding functions that are particularly vulnerable under conditions of salt-stress could counteract the molecular effects of such stress and could provide incremental improvements in tolerance. We have proceeded to identify the acquired specific changes in gene regulation for our salt-tolerant mutant cells and plants. One particularly interesting and novel gene isolate from the salt-tolerant cells is Alfin1, which encodes a putative zinc-finger regulatory protein, expressed predominantly in roots. We have demonstrated that this protein binds DNA in a sequence specific manner and may be potentially important in gene regulation in roots in response to salt and an important marker for salt tolerance in crop plants.     

Changes in RACK1 expression induce defects in nodulation and development in Phaseolus vulgaris

RACK1 is a scaffold protein with the ability to interact in a regulated manner with a diverse number of ligands from distinct signal-transduction pathways. This assessment allowed us to infer that it may be involved in different processes such as nodulation. In a recent study we showed by silencing, that PvRACK1 has a pivotal role in cell expansion and in symbiosome and bacteroid integrity during nodule development in Phaseolus vulgaris. On the other hand, we have also observed that its overexpression provokes a dramatic phenotype in: (a) seedlings that have been exposed to heat, in which systemic necrosis is induced; and (b) in Agrobacterium rhizogenes-transformed roots, where nodulation is strongly inhibited and nodules show early senescent symptoms. These findings indicate that PvRACK1 may be an integrator of diverse signal-transduction pathways in processes as varied as nodulation, cell expansion, heat stress responses, and systemic activation of necrosis.