The effect of salt on protein synthesis in the halophyte Suaeda maritima

Summary An amino acid-incorporating microsomal fraction has been isolated from the leaves of the halophyte Suaeda maritima and the characteristics of the incorporation described. There were no differences in the properties of the microsomes isolated from plants grown in saline and non-saline conditions. The incorporation was severely inhibited by high concentrations of sodium or potassium ions. The results are discussed in relation to the mechanism of salt tolerance in halophytes and the localization of salt in the cells.     

Accumulation of low molecular weight solutes in water-stressed tropical legumes

Fourteen species of tropical legumes, representing 10 genera, were subjected to water stress. Youngest fully expanded leaves of stressed and unstressed plants were analysed for inorganic ions, sugars, inositols, organic acids, betaines and amino acids. The major compounds which accumulated with water stress were O-methyl-inositols (14 species), 2-methyl-2,3,4-trihydroxybutanoic acid-1,4-lactone (10 species) and proline (9 species). Concentrations of inorganic ions, sugars and organic acids decreased or were unchanged in the majority of the stressed species. The betaines, glycinebetaine, trigonelline and stachydrine were detected in low concentrations in most of the legumes but did not accumulate to any degree during water stress. All the legumes which tolerated low leaf water potentials accumulated the O-methyl-inositol, pinitol. The other species, with the exception of Siratro, contained ononitol or O-methyl-scyllo-inositol but no pinitol. It is suggested that pinitol accumulation may indicate a legume able to tolerate low leaf water potentials.     

Effect of salinity stress on organic and mineral constituents in the leaves of pigeonpea (Cajanus cajan L. Var. C-11)

Summary Effects of sodium chloride and sodium sulphate on the content of some organic and inorganic constituents in the leaves of pigeonpea (Cajanus cajan L. Var. C-11) were studied. Increased water content under saline conditions made the leaves succulent. The concentration of reducing sugars appeared to be higher while that of total sugars and starch was lower. The plants also failed to accumulate proline at higher salinity levels. Phosphorus and potassium content were lowered while those of calcium, magnesium, sodium, chloride and sulphate were increase under both salinities. This indicates that there is no regulation on the uptake of latter elements under saline conditions.     

Taxonomic and ecological aspects of the distribution of glycinebetaine and related compounds in plants

Summary The concentrations of the major inorganic ions and glycinebetaine, choline and proline and the osmotic pressure of extract sap have been determined in eight salt marsh species and four sand dune species from local habitats. These results together with those previously reported on hydroponically grown plants and data assembled from the literature show that glycinebetaine accumulation is a feature of members of the Chenopodiaceae, Amaranthaceae, many Gramineae and some members of the Solanaceae and Compositae, particularly when exposed to conditions of low soil water potential. It is suggested that in these families betaine is employed as a non-toxic cytoplasmic osmoticum when decreased osmotic potentials are required. In some other plant species proline may fulfil a similar function. Another quaternary ammonium compound may be accumulated in the Plumbaginaceae in addition to proline. Some evidence suggests that the differences in the organic osmoticum used may relate to the different inorganic ion contents of the plants. The accumulation of nitrogen dipoles as cytoplasmic osmotica may make heavy demands on the nitrogen economy of the plants and this problem is discussed briefly.     

The effects of NaCl on growth,water relations,osmolytes and ion content in <Emphasis Type="Italic">Kochia prostrata</Emphasis>

The effects of NaCl (0, 50, 100, 150 and 200 mM) on growth, water relations, glycinebetaine, free proline, ion contents, stomata number and size of Kochia prostrata (L.) Schard were determined. Shoot and root fresh and dry matter, root and shoot length, relative growth rate, net assimilation rate, relative water content, water use efficiency, soluble sugars and glycinebetaine contents were not changed at low NaCl concentrations, but they were significantly decreased at 200 mM NaCl. The K⁺, Mg²⁺ and Ca²⁺ contents, water potential, chlorophyll a+b and carotenoides contents, and stomata number and size were reduced already at low concentrations of NaCl. In contrast, the Na⁺, Cl^– and proline contents increased several times with increasing NaCl concentration. Kochia prostrata is a salt tolerant species, the optimal growth of this plant occurred up to 150 mM NaCl. The mechanisms of salt tolerance in the plant may be balance among ion accumulation and production of glycinebetaine, proline, soluble sugars for maintenance of pressure potential.     

Biochemical,physiological and growth changes in response to salinity in callus cultures of <Emphasis Type="Italic">Sesuvium portulacastrum</Emphasis> L.

In vitro-grown cells of Sesuvium portulacastrum L., an important ‘salt accumulator’ mangrove associate, were incubated on a medium containing different levels of salt, including 0, 100, 200, or 400 mM NaCl, in order to evaluate biochemical, physiological, and growth responses. A significant decrease in callus growth, water status, and cell membrane damage was observed under salt stress. Osmotic adjustment was revealed by the accumulation of inorganic ions, such as sodium (Na⁺), and organic osmolytes (proline, glycine betaine, and total soluble sugars) in NaCl-treated calli compared to control. However, accretion of osmolytes and inorganic ions did not support growth of calli under NaCl stress conditions. The observed reduced growth rate in calli subjected to stress, up to 200 mM NaCl, was coupled with lower catalase and ascorbate peroxidase activities and with a significantly higher superoxide dismutase activity. These findings suggested that S. portulacastrum cell cultures exhibited higher osmotic adjustment to salt stress.     

Transformation of Arabidopsis thaliana with the codA gene for choline oxidase; accumulation of glycinebetaine and enhanced tolerance to salt and cold stress

Glycinebetaine is one of the compatible solutes that accumulate in the chloroplasts of certain halotolerant plants when these plants are exposed to salt or cold stress. The codA gene for choline oxidase, the enzyme that converts choline into glycinebetaine, has previously been cloned from a soil bacterium, Arthrobacter globiformis. Transformation of Arabidopsis thaliana with the cloned codA gene under the control of the 35S promoter of cauliflower mosaic virus enabled the plant to accumulate glycinebetaine and enhanced its tolerance to salt and cold stress. At 300 mM NaCl, considerable proportions of seeds of transformed plants germinated well, whereas seeds of wild-type plants failed to germinate. At 100 mM NaCl, transformed plants grew well whereas wild-type plants did not do so. The transformed plants tolerated 200 mM NaCl, which was lethal to wild-type plants. After plants had been incubated with 400 mM NaCl for two days, the photosystem II activity of wild-type plants had almost completely disappeared, whereas that of transformed plants remained at more than 50% of the original level. When exposed to a low temperature in the light, leaves of wild-type plants exhibited symptoms of chlorosis, whereas those of transformed plants did not. These observations demonstrate that the genetic modification of Arabidopsis thaliana that allowed it to accumulate glycinebetaine enhanced its ability to tolerate salt and cold stress.     

Assessment of glycinebetaine and proline compartmentation by analysis of isolated beet vacuoles

Vacuoles isolated from storage root tissue of red beet (Beta vulgaris L.) do not leak significant quantities of betanin, sucrose, Na⁺ or K⁺ during isolation. This indicates that analysis of vacuoles in vitro gives meanigful information about the compartmentation of solutes in vivo. Preparations of vacouoles were used to determine the distribution of glycinebetaine and proline between vacuole and cytoplasm in beet cells. Both compounds were detected in preparations of isolated beet vacuoles. In the case of glycinebetaine it was shown that this solute was associated with the vacuoles, not with the small number of other organelles which contaminated the preparations. The vacuolar pool accounted for 26 to 84% of the total tissue glycinebetaine and 17 to 57% of the proline. Concentrations of these compounds in vacuole and cytoplasm were calculated and were always higher in the cytoplasm than in the vacuole. The concentration gradient across the tonoplast varied considerably. The significance of these results is discussed in relation to the hypothesis that glycinebetaine and proline function as benign cytoplasmic osmotica.Abbreviations A₅₃₇ absorbance at 537 nm - MES 2-(N-morpholino)-ethanesulphonic acid - Na₂EDTA ethylenediaminetetraacetic acid, disodium salt - SDS sodium dodecyl sulphate - Tris tris(hydroxymethyl)methylamine     

Low-molecular-weight carbohydrates in some salt-stressed plants

A study was made of the effects of salinity on the concentrations of free sugars, glycinebetaine, proline and other chemical components of Aster tripolium L., Daucus carota L., Honkenya peploides (L.) Ehr. and Plantago coronopus L. (Dicotyledones); and Carex extensa Good., Eleocharis uniglumis (Link) Schutt., Juncus maritima Lam. and Schoenoplectus tabernaemontani (C. C. Gmel.) Palla (Monocotyledones) grown in the laboratory. In Plantago coronopus the level of the polyol sorbitol increased when the plants were subjected to NaCl stress, while in Honkenya peploides the cyclitol pinitol accumulated. No consistent pattern emerged with respect to the changes in free sugar contents in either the monocotyledonous or dicotyledonous plants, though the monocotyledonous plants generally had higher sugar contents.     

Early effects of salt stress on the physiological and oxidative status of Cakile maritima (halophyte) and Arabidopsis thaliana (glycophyte)

Early changes in physiological and oxidative status induced by salt stress were monitored in two Brassicaceae plants differing in their tolerance to salinity, Cakile maritima (halophyte) and Arabidopsis thaliana (glycophyte). Growth response and antioxidant defense of C. maritima under 400 mM NaCl were compared with those of A. thaliana exposed to 100 mM NaCl. Salinity induced early growth reduction that is less pronounced in C. maritima than in A. thaliana. Maximum hydrogen peroxide (H₂O₂) level occurred in the leaves of both species 4 h after the onset of salt treatment. A rapid decline in H₂O₂ concentration was observed thereafter in C. maritima, whereas it remained high in A. thaliana. Correlatively, superoxide dismutase, catalase and peroxidase activities increased at 4 h of treatment in C. maritima and decreased thereafter. However, the activity of these enzymes remained higher in treated plants than that in controls, regardless of the duration of treatment, in A. thaliana. The concentrations of malondialdehyde (MDA) reached maximum values at 24 h of salt stress in both species. Again, MDA levels decreased later in C. maritima, but remained high in A. thaliana. The contents of α‐tocopherol remained constant during salt stress in C. maritima and decreased during the first 24 h of salt stress and then remained low in A. thaliana. The results clearly showed that C. maritima, in contrast to A. thaliana, can rapidly evolve physiological and antioxidant mechanisms to adapt to salt and manage the oxidative stress. This may explain, at least partially, the difference in salt tolerance between halophytes and glycophytes.     

Compatibility of glycinebetaine in rice plants: evaluation using transgenic rice plants with a gene for peroxisomal betaine aldehyde dehydrogenase from barley

Glycinebetaine is synthesized in plants by the two‐step oxidation of choline, with betaine aldehyde as the intermediate. The reactions are catalyzed by choline mono‐oxygenase and betaine aldehyde dehydrogenase. Rice plants, which do not accumulate glycinebetaine, possess a gene encoding betaine aldehyde dehydrogenase, whose activity is detectable at low levels. To evaluate the compatibility in rice of glycinebetaine on growth and tolerance to salt, cold and heat, we produced transgenic rice plants by introduction of a cDNA for betaine aldehyde dehydrogenase of barley, which is localized in peroxisomes unlike the chloroplast‐specific localization of betaine aldehyde dehydrogenase in spinach and sugar beet. The transgenic rice plants converted high levels of exogenously applied betaine aldehyde (up to 10 mol m^–3) to glycinebetaine more efficiently than did wild‐type plants. The elevated level of glycinebetaine in transgenic plants conferred significant tolerance to salt, cold and heat stress. However, very high levels of glycinebetaine, resulting from conversion of applied betaine aldehyde to glycinebetaine or from exogenous application, inhibited increases in length of rice plants but not increases in dry weight. Our results suggested that the benefits of accumulation of glycinebetaine by rice plants might be considerable under high light conditions.     

Salt regulation in halophytes

Summary Ion concentration and saturation water content were measured in various aged leaves of halophytes growing in saline soils east of lake Neusiedlersee (Austria).All species investigated showed a substantial sodium accumulation within the maturing organs accompanied by a considerable potassium decline. In most species chloride concentration rises distinctly with increasing leaf age, too, whereas concentration shifts of alkaline earth ions and of sulfate (except in Plantago maritima, Lepidium crassifolium and Crypsis aculeata) are of comparably less importance.Saturation water increases markedly in succulent species (Suaeda maritima, Chenopodium glaucum, Spergularia media, Lepidium crassifolium) and to a less degree in xerophytic monocotyledons (Puccinellia distans, Crypsis aculeata, Bolboschoenus maritimus). However, this surplus of water in older leaves is not sufficient to dilute the salt to such an extent that a rise in concentration can be prevented (except chloride in Suaeda maritima and Chenopodium glaucum).Rosette plants (Triglochin maritimum, Plantago maritima, Scorzonera parviflora, Aster tripolium) with the ability to renew their leaves continuously throughout the growth period are characterized by only insignificant changes of saturation water content with increasing leaf age. In these plants, shedding of old salt-saturated leaves is thought to be the main strategy for salt regulation.A modification of Steiner's classical concept of different salt regulation types is proposed, based on original findings about salt regulation in Austrian halophytes and on new bibliographical data upon additionally revealed regulatory principles in halophytes and saltaffected nonhalophytes.Respectfully and thankfully dedicated to Prof. Dr. H. Kinzel on the occasion of his 50th birthday.     

Osmoregulation in halophytic higher plants: a comparative study of soluble carbohydrates, polyols, betaines and free proline

Abstract. The levels of soluble carbohydrates, polyols, betaines and free proline have been determined in different organs of sixteen plants collected on coastal salt marshes. Among the carbohydrates, sucrose, fructose and glucose accumulated at quite high levels. Sucrose is particularly abundant in the monocots Juncus maritima, Phragmites communis and Scirpus maritimus. Maltose is quite abundant in Atriplex hastata and rhamnose in Plantago maritima roots. High levels of polyols were detected in Aster tripolium, Juncus maritimus, Plantago maritima and Phragmites communis. According to their capacity to accumulate carbohydrates and (or) nitrogenous solutes, halophytic higher plants can be divided into three main groups: (1) species producing high levels of soluble carbohydrates only, (2) species accumulating both carbohydrates and nitrogenous compounds, (3) species producing more nitrogenous solutes than soluble carbohydrates. The possible functions of these various organic solutes, including a role in equalizing the relative water potentials of cytoplasm and vacuole and to lower the internal potential, are discussed.     

Halophytes--an emerging trend in phytoremediation

Halophytic plants are of special interest because these plants are naturally present in environments characterized by an excess of toxic ions, mainly sodium and chloride. Several studies have revealed that these plants may also tolerate other stresses including heavy metals based on the findings that tolerance to salt and to heavy metals may, at least partly, rely on common physiological mechanisms. In addition, it has been shown that salt-tolerant plants may also be able to accumulate metals. Therefore, halophytes have been suggested to be naturally better adapted to cope with environmental stresses, including heavy metals compared to salt-sensitive crop plants commonly chosen for phytoextraction purposes. Thus, potentially halophytes are ideal candidates for phytoextraction orphytostabilization of heavy metal polluted soils and moreover of heavy metal polluted soils affected by salinity. Some halophytes use excretion processes in order to remove the excess of salt ions from their sensitive tissues and in some cases these glandular structures are not always specific to Na+ and Cl- and other toxic elements such as cadmium, zinc, lead, or copper are accumulated and excreted by salt glands or trichomes on the surface of the leaves--a novel phytoremediation process called "phytoexcretion". Finally, the use of halophytes has also been proposed for soil desalination through salt accumulation in the plant tissue or dissolution of soil calcite in the rhizosphere to provide Ca2+ that can be exchanged with Na+ at cation exchange sites.     

ABI4 downregulates expression of the sodium transporter HKT1;1 in Arabidopsis roots and affects salt tolerance

A plant's ability to cope with salt stress is highly correlated with their ability to reduce the accumulation of sodium ions in the shoot. Arabidopsis mutants affected in the ABSCISIC ACID INSENSITIVE (ABI) 4 gene display increased salt tolerance, whereas ABI4‐overexpressors are hypersensitive to salinity from seed germination to late vegetative developmental stages. In this study we demonstrate that abi4 mutant plants accumulate lower levels of sodium ions and higher levels of proline than wild‐type plants following salt stress. We show higher HKT1;1 expression in abi4 mutant plants and lower levels of expression in ABI4‐overexpressing plants, resulting in reduced accumulation of sodium ions in the shoot of abi4 mutants. HKT1;1 encodes a sodium transporter which is known to unload sodium ions from the root xylem stream into the xylem parenchyma stele cells. We have shown recently that ABI4 is expressed in the root stele at various developmental stages and that it plays a key role in determining root architecture. Thus ABI4 and HKT1;1 are expressed in the same cells, which suggests the possibility of direct binding of ABI4 to the HKT1;1 promoter. In planta chromatin immunoprecipitation and in vitro electrophoresis mobility shift assays demonstrated that ABI4 binds two highly related sites within the HKT1;1 promoter. These sites, GC(C/G)GCTT(T), termed ABI4‐binding element (ABE), have also been identified in other ABI4‐repressed genes. We therefore suggest that ABI4 is a major modulator of root development and function.     

Changes in the water status and osmotic solute contents in response to drought and salicylic acid treatments in four different cultivars of wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis>)

Salicylic acid (SA) controls growth and stress responses in plants. It also induces drought tolerance in plants. In this paper, four wheat (Triticum aestivum L.) cultivars with different drought responses were treated with SA in three levels of drain (90, 60, 30% of maximum field capacity) to examine its interactive effects on drought responses and contents of osmotic solutes that may be involved in growth and osmotic adjustment. Under drought condition, the cultivars Geza 164 and Sakha 69 had the plant biomass and leaf relative water content (LRWC) greater than the cultivars Gemaza 1 and Gemaza 3. In all cultivars, drought stress decreased the biomass, LRWC, and the contents of inorganic solutes (Ca, K, Mg) and largely increased the contents of organic solutes (soluble sugars and proline). By contrast, SA increased the biomass, LRWC and the inorganic and organic solute contents, except proline. Correlation analysis revealed that the LRWC correlated positively with the inorganic solute contents but negatively with proline in all cultivars. SA caused maximum accumulations of soluble sugars in roots under drought. These results indicated that SA-enhanced tolerance might involve solute accumulations but independently of proline biosynthesis. Drought-sensitive cultivars had a trait lowering Ca and K levels especially in shoots. Possible functions of the ions and different traits of cultivars were discussed.     

Role of antioxidant systems in wild plant adaptation to salt stress

Wild plants differing in the strategies of adaptation to salinity were grown for six weeks in the phytotron and then subjected to salt stress (100 mM NaCl, 24 h). The activities of principal antioxidant enzymes and the accumulation of sodium ions and proline were studied. Independently of the level of constitutive salt tolerance, plants of all species tested accumulated sodium ions under salinity conditions but differed in their capability of stress-dependent proline accumulation and superoxide dismutase (SOD) and guaiacol-dependent peroxidase activities. Proline-accumulating species were found among both halophytes (Artemisia lerchiana and Thellungiella halophila) and glycophytes (Plantago major and Mycelis muralis). The high activities of ionically-bound and covalently bound peroxidases were characteristic of Th. halophila plants. High constitutive and stress-induced SOD activities were, as a rule, characteristic of glycophytes with the low constitutive proline level: Geum urbanum and Thalictrum aquilegifolium. Thus, a negative correlation was found between proline content and SOD activity in wild species tested; it was especially bright in the halophyte Th. halophila and glycophyte G. urbanum. An extremely high constitutive and stress-induced levels of proline and peroxidase activity in Th. halophila maybe compensate SOD low activity in this plant, and this contributed substantially into its salt resistance. Thus, monitoring of stress-dependent activities of some antioxidant enzymes and proline accumulation in wild plant species allowed a supposition of reciprocal interrelations between SOD activity and proline accumulation. It was also established that the high SOD activity is not obligatory trait of species salt tolerance. Moreover, plants with the high activity of peroxidase and active proline accumulation could acclimate to salts stress (100 mM NaCl, 24 h) independently of SOD activity.     

Salt Tolerance in the Halophyte, Suaeda maritima (L.) Dum.: The Influence of the Salinity of the Culture Solution on the Content of Various Organic Compounds

Plants of the halophyte Suaeda maritima were grown in tap wateror in a culture solution in the presence or absence of sodiumchloride and the levels of sugars, amino acids, organic acidsand quaternary ammonium compounds determined in relation tothe balance between cytoplasmic and vacuolar water potentials.The sugar content (some 7 µmol. g f. wt^–1) was unaffectedby the salinity of the growth medium as was the overall contentof amino acids (about 4 µmol. g f. wt^–1). The organicacid content was maximal in plants kept in tap water alone wherethe dominant acid was malic. Plants grown in culture solutioncontained the same acids, although addition of sodium chlorideto the medium brought about the apparent loss of glycolic acidand the appearance of oxalic acid. Only a single quaternaryammonium compound, glycinebetaine, was apparently present inthe tissues: the content of betaine doubled (to 37·5µrmol. g f. wt^–) when sodium chloride was addedto the culture solution. The content of these various compoundsis discussed in relation to the relative values of the cytoplasmicand vacuolar components of the overall tissue water potential Suaeda maritima, halophyte, salt tolerance, betaine, organic compounds, water potential     

INFLUENCE OF HORMONAL TREATMENTS ON THE GROWTH OF TWO HALOPHYTIC SPECIES OF SUAEDA

Experiments were made to determine the interaction between the growth regulating substances, gibberellic acid and kinetin, and salinity on the growth and development of two species of Suaeda. We found that the growth of Suaeda maritima var. macrocarpa, an obligate halophyte, was greatly stimulated with treatments by both of these hormones in controls not treated with sodium chloride. These results suggest that halophytes grow poorly under nonsaline conditions because of some type of hormonal imbalance. GA₃ was found to stimulate growth at all salinities for both S. maritima var. macrocarpa and S. depressa, while kinetin proved to be inhibitory to growth and elongation of plants at higher salinities. Chlorophyll content decreased with salt treatments and was not significantly influenced by hormonal treatments. GA₃ had little influence on water content in roots and shoots, whereas plants treated with kinetin generally had reduced water contents.