Intracellular compartmentation of ions in salt adapted tobacco cells

Na⁺ and Cl⁻ are the principal solutes utilized for osmotic adjustment in cells of Nicotiana tabacum L. var Wisconsin 38 (tobacco) adapted to NaCl, accumulating to levels of 472 and 386 millimolar, respectively, in cells adapted to 428 millimolar NaCl. X-ray microanalysis of unetched frozen-hydrated cells adapted to salt indicated that Na⁺ and Cl⁻ were compartmentalized in the vacuole, at concentrations of 780 and 624 millimolar, respectively, while cytoplasmic concentrations of the ions were maintained at 96 millimolar. The morphometric differences which existed between unadapted and salt adapted cells, (cytoplasmic volume of 22 and 45% of the cell, respectively), facilitated containment of the excited volume of the x-ray signal in the cytoplasm of the adapted cells. Confirmation of ion compartmentation in salt adapted cells was obtained based on kinetic analyses of ²²Na⁺ and ³⁶Cl⁻ efflux from cells in steady state. These data provide evidence that ion compartmentation is a component of salt adaptation of glycophyte cells.     

Abscisic Acid Stimulated Osmotic Adjustment and Its Involvement in Adaptation of Tobacco Cells to NaCl

Osmotic adjustment of cultured tobacco (Nicotiana tabacum L. var Wisconsin 38) cells was stimulated by 10 micromolar (±) abscisic acid (ABA) during adaptation to water deficit imposed by various solutes including NaCl, KCl, K₂SO₄, Na₂SO₄, sucrose, mannitol, or glucose. The maximum difference in cell osmotic potential (Ψπ) caused by ABA treatment during adaptation to 171 millimolar NaCl was about 6 to 7 bar. The cell Ψπ differences elicited by ABA were not due to growth inhibition since ABA stimulated growth of cells in the presence of 171 millimolar NaCl. ABA caused a cell Ψπ difference of about 1 to 2 bar in medium without added NaCl. Intracellular concentrations of Na⁺, K⁺, Cl⁻, free amino acids, or organic acids could not account for the Ψπ differences induced by ABA in NaCl treated cells. However, since growth of NaCl treated cells is more rapid in the presence of ABA than in its absence, greater accumulation of Na⁺, K⁺, and Cl⁻ was necessary for ion pool maintenance. Higher intracellular sucrose and reducing sugar concentrations could account for the majority of the greater osmotic adjustment of ABA treated cells. More rapid accumulation of proline associated with ABA treatment was highly correlated with the effects of ABA on cell Ψπ. These and other data indicate that the role of ABA in accelerating salt adaptation is not mediated by simply stimulating osmotic adjustment.     

Abscisic Acid accelerates adaptation of cultured tobacco cells to salt

Adaptation of tobacco (Nicotiana tabacum L. var Wisconsin 38) cells to NaCl was accelerated by (±) abscisic acid (ABA). In medium with 10 grams per liter NaCl, ABA stimulated the growth of cells not grown in medium with NaCl (unadapted, S-0) with an increasing response from 10⁻⁸ to 10⁻⁴ molar. ABA (10⁻⁵ molar) enhanced the growth of unadapted cells in medium with 6 to 22 grams per liter NaCl but did not increase the growth of cells previously adapted to either 10 (S-10) or 25 (S-25) grams per liter NaCl unless the cells were inoculated into medium with a level of NaCl higher than the level to which the cells were adapted. The growth of unadapted cells in medium with Na₂SO₄ (85.5 millimolar), KCl (85.5 or 171 millimolar), K₂SO₄ (85.5 millimolar) was also stimulated by ABA. ABA (10⁻⁸-10⁻⁴ molar) did not accelerate the growth of unadapted cells exposed to water deficits induced by polyethylene glycol (molecular weight 8000) (5-20 grams per 100 milliliters), sorbitol (342 millimolar), mannitol (342 millimolar) or sucrose (342 millimolar). These results suggest that ABA is involved in adaptation of cells to salts, and is not effective in promoting adaptation to water deficits elicited by nonionic osmotic solutes.     

Rapid osmotic adjustment by a succulent halophyte to saline shock

The objective of this research was to measure the short term osmotic adjustment of Salicornia europaea L. ssp. rubra (A. Nels) Breitung when suddenly exposed to 100 millimolar NaCl. Plants were grown hydroponically, shocked with 100 millimolar NaCl added to the culture solution, and stem tips analyzed for free inorganic ions and small organic molecules at intervals up to 72 hours. In the first 2 hours, the calculated leaf osmoticum showed a net increase of 158.8 millimolar most of which was free Mg²⁺ (+135.3 millimolar). Total sugars increased almost 5-fold by the 6th hour, enough to provide sufficient osmoticum for the cytoplasm if only partially confined there. By 24 hours, all measured osmotica had decreased except Na⁺, Mg²⁺, Cl⁻, and proline, with the net increase being 208 millimolar. By 72 hours, there was a net gain of 356 millimolar in osmotica of the stem tips, due to Na⁺ (+233.3 millimolar), Cl⁻ (+306.7 millimolar), and a small increase in sugar and proline (+3.5 millimolar), with all other osmotica decreasing in concentration. Compatible osmotica did not change sufficiently to account for osmotic balance between vacuole and cytoplasm; consequently, there must have been a reapportionment of osmotica within the cell in the short time duration of this experiment.     

Maintenance of low cl concentrations in mesophyll cells of leaf blades of barley seedlings exposed to salt stress

The concentrations of vacuolar Na⁺ and Cl⁻ in the epidermal and mesophyll cells of the leaf blade and sheath of Hordeum vulgare seedlings (cv California Mariout and Clipper) were measured by means of quantitative electron probe x-ray microanalysis. A preferential accumulation of Cl⁻ in vacuoles of epidermal cells in both blade and sheath and a low level in mesophyll cells of the blade were evident in plants grown in full strength Johnson solution. The concentration of Cl⁻ in the mesophyll cells of the blade remained at a low level after exposure to 50 or 100 millimolar NaCl for 1 day or to 50 millimolar for 4 days, while at the same time the concentration of Cl⁻ in the epidermis and mesophyll of the sheath showed a dramatic increase. Clipper generally contained more Cl⁻ in the mesophyll cells of the blade than California Mariout. A greater accumulation of Na⁺ in the mesophyll of the sheath relative to that of the blade was only apparent after treatment with 100 millimolar NaCl for 1 day or 50 millimolar for 4 days. These results confirm the suggestion that sheath tissue is capable of accumulating excess Cl⁻ (and to a lesser extent Na⁺) and suggest that the site of regulation of Cl⁻ concentration in the barley leaf is located in the mesophyll cells of the blade.     

Solutes contributing to osmotic adjustment in cultured plant cells adapted to water stress

Osmotic adjustment was studied in cultured cells of tomato (Lycopersicon esculentum Mill cv VFNT-Cherry) adapted to different levels of external water potential ranging from −4 bar to −28 bar. The intracellular concentrations of reducing sugars, total free amino acids, proline, malate, citrate, quaternary ammonium compounds, K⁺, NO₃⁻, Na⁺, and Cl⁻ increased with decreasing external water potential. At any given level of adaptation, the maximum contribution to osmotic potential was from reducing sugars followed by potassium ions. The sucrose levels in the cells were 3- to 8-fold lower than reducing sugar levels and did not increase beyond those observed in cells adapted to −16 bar water potential. Concentrations of total free amino acids were 4- to 5-fold higher in adapted cells. Soluble protein levels declined in the adapted cell lines, but the total reduced nitrogen was not significantly different after adaptation. Uptake of nitrogen (as NH₄⁺ or NO₃⁻) from the media was similar for adapted and unadapted cells. Although the level of quaternary ammonium compounds was higher in the nonadapted cells than that of free proline, free proline increased as much as 500-fold compared to only a 2- to 3-fold increase observed for quaternary ammonium compounds. Although osmotic adjustment after adaptation was substantial (up to −36 bar), fresh weight (volume increase) was restricted by as much as 50% in the adapted cells. Altered metabolite partitioning was evidenced by an increase in the soluble sugars and soluble nitrogen in adapted cells which occurred at the expense of incorporation of sugar into cell walls and nitrogen into protein. Data indicate that the relative importance of a given solute to osmotic adjustment may change depending on the level of adaptation.     

Sodium,potassium, chloride and proline concentrations of chloroplasts isolated from a halophyte,Mesembryanthemum crystallinum L.

Concentrations of four major solutes (Na⁺, K⁺, Cl^-, proline) were determined in isolated, intact chloroplasts from the halophyte Mesembryanthemum crystallinum L. following long-term exposure of plants to three levels of NaCl salinity in the rooting medium. Chloroplasts were obtained by gentle rupture of leaf protoplasts. There was either no or only small leakage of inorganic ions from the chloroplasts to the medium during three rapidly performed washing steps involving precipitation and re-suspension of chloroplast pellets. Increasing NaCl salinity of the rooting medium resulted in a rise of Na⁺ und Cl^- in the total leaf sap, up to approximately 500 and 400 mM, respectively, for plants grown at 400 mM NaCl. However, chloroplast levels of Na⁺ und Cl^- did not exceed 160–230 and 40–60 mM, respectively, based upon a chloroplast osmotic volume of 20–30 l per mg chlorophyll. At 20 mM NaCl in the rooting medium, the Na⁺/K⁺ ratio of the chloroplasts was about 1; at 400 mM NaCl the ratio was about 5. Growth at 400 mM NaCl led to markedly increased concentrations of proline in the leaf sap (8 mM) compared with the leaf sap of plants grown in culture solution without added NaCl (proline 0.25 mM). Although proline was fivefold more concentrated in the chloroplasts than in the total leaf sap of plants treated with 400 mM NaCl, the overall contribution of proline to the osmotic adjustment of chloroplasts was small. The capacity to limit chloroplast Cl^- concentrations under conditions of high external salinity was in contrast to an apparent affinity of chloroplasts for Cl^- under conditions of low Cl^- availability.Abbreviation Chl chlorophyll     

Sodium chloride toxicity and the cellular basis of salt tolerance in halophytes

Background Halophytes are the flora of saline soils. They adjust osmotically to soil salinity by accumulating ions and sequestering the vast majority of these (generally Na⁺ and Cl⁻) in vacuoles, while in the cytoplasm organic solutes are accumulated to prevent adverse effects on metabolism. At high salinities, however, growth is inhibited. Possible causes are: toxicity to metabolism of Na⁺ and/or Cl⁻ in the cytoplasm; insufficient osmotic adjustment resulting in reduced net photosynthesis because of stomatal closure; reduced turgor for expansion growth; adverse cellular water relations if ions build up in the apoplast (cell walls) of leaves; diversion of energy needed to maintain solute homeostasis; sub-optimal levels of K⁺ (or other mineral nutrients) required for maintaining enzyme activities; possible damage from reactive oxygen species; or changes in hormonal concentrations.Scope This review discusses the evidence for Na⁺ and Cl⁻ toxicity and the concept of tissue tolerance in relation to halophytes.Conclusions The data reviewed here suggest that halophytes tolerate cytoplasmic Na⁺ and Cl⁻ concentrations of 100–200 mm, but whether these ions ever reach toxic concentrations that inhibit metabolism in the cytoplasm or cause death is unknown. Measurements of ion concentrations in the cytosol of various cell types for contrasting species and growth conditions are needed. Future work should also focus on the properties of the tonoplast that enable ion accumulation and prevent ion leakage, such as the special properties of ion transporters and of the lipids that determine membrane permeability.     

Ion Relations of Symplastic and Apoplastic Space in Leaves from Spinacia oleracea L. and Pisum sativum L. under Salinity

Salt tolerant spinach (Spinacia oleracea) and salt sensitive pea (Pisum sativum) plants were exposed to mild salinity under identical growth conditions. In order to compare the ability of the two species for extra- and intracellular solute compartmentation in leaves, various solutes were determined in intercellular washing fluids and in aqueously isolated intact chloroplasts. In pea plants exposed to 100 millimolar NaCl for 14 days, apoplastic salt concentrations in leaflets increased continuously with time up to 204 (Cl⁻) and 87 millimolar (Na⁺), whereas the two ions reached a steady concentration of only 13 and 7 millimolar, respectively, in spinach leaves. In isolated intact chloroplasts from both species, sodium concentrations were not much different, but chloride concentrations were significantly higher in pea than in spinach. Together with data from whole leaf extracts, these measurements permitted an estimation of apoplastic, cytoplasmic, and vacuolar solute concentrations. Sodium and chloride concentration gradients across the tonoplast were rather similar in both species, but spinach was able to maintain much steeper sodium gradients across the plasmamembrane compared with peas. Between day 12 and day 17, concentrations of other inorganic ions in the pea leaf apoplast increased abruptly, indicating the onset of cell disintegration. It is concluded that the differential salt sensitivity of pea and spinach cannot be traced back to a single plant performance. Major differences appear to be the inability of pea to control salt accumulation in the shoot, to maintain steep ion gradients across the leaf cell plasmalemma, and to synthesize compatible solutes. Perhaps less important is a lower selectivity of pea for K⁺/Na⁺ and NO₃⁻/Cl⁻ uptake by roots.     

Solutes Involved in Osmotic Adjustment to Increasing Salinity in Suspension Cells of Alternanthera philoxeroides Griseb

Cell recovery from osmotic stress was studied in suspension cell cultures from Alternanthera philoxeroides [Mart.] Griseb. Changes in different classes of cellular solutes were measured after cells were transferred from 0 to 200 mM NaCl (high salt) to obtain an integrated picture of the solute pools involved in osmotic adjustment. By 2 h, cellular [Na⁺] and [Cl⁻] had increased several-fold, potentially accounting for the osmotic adjustment that produced a rapid recovery of cell turgor. There was a four-fold increase in the concentration of quaternary ammonium compounds (QAC) by 12 h and a slower increase for several days afterward. Betaine aldehyde dehydrogenase (BADH) is required for synthesis of glycine betaine, a QAC produced by a range of organisms in response to osmotic stress. Western-blot analysis for BADH suggested that glycine betaine was a significant component of the QAC solutes. The amount of BADH was generally similar at different sampling times for control and high salt cells, unlike previous reports of stimulation by osmotic stress in intact plants of some species. Between 3 and 7 days after cell transfer to high salt, other organic solutes increased in concentration and [Na⁺] and [Cl⁻] decreased. In A. philoxeroides, high [Na⁺] and [Cl⁻] produce rapid osmotic adjustment but organic solutes apparently replace these potentially harmful inorganic ions after the recovery of turgor.     

Accumulation of inorganic and organic osmolytes and their role in osmotic adjustment in NaCl-stressed vetiver grass seedlings

The accumulation of inorganic and organic osmolytes and their role in osmotic adjustment were investigated in roots and leaves of vetiver grass (Vetiveria zizanioides) seedlings stressed with 100, 200, and 300 mM NaCl for 9 days. The results showed that, although the contents of inorganic (K⁺, Na⁺, Ca²⁺, Mg²⁺, Cl⁻, NO₃⁻, SO₄²⁻ and H₂PO₃⁻)) and organic (soluble sugar, organic acids, and free amino acids) osmolytes all increased with NaCl concentration, the contribution of inorganic ions (mainly Na⁺, K⁺, and Cl⁻) to osmotic adjustment was higher (71.50–80.56% of total) than that of organic solutes (19.43–28.50%). The contribution of inorganic ions increased and that of organic solutes decreased in roots with the enhanced NaCl concentration, whereas the case in leaves was opposite. On the other hand, the osmotic adjustment was only effective for vetiver grass seedlings under moderate saline stress (less than 200 mM NaCl).     

Physiology of halophytes

Summary The cellular basis of salt tolerance in halophytes depends upon the compartmentation of ions necessary for osmoregulation in vacuoles and upon osmotic adjustment of the cytoplasm by compatible solutes. The central role played by Na⁺ and Cl^– in osmotic adjustment suggests that the transport of these ions and its regulation must be of primary importance in the physiology of the plant as a whole. There have been few investigations into the regulation of leaf ion concentrations, but such data as are in the literature suggest that limiting xylem Na⁺ (and Cl^–) concentrations, together with continued leaf expansion, are particularly important. The role of phloem in retranslocation is uncertain due to lack of data. Decreases in transpiration rate per unit area of leaf help to lower the ion input into leaves. Any linked reductions in photosynthesis appear to be due to decreases in stomatal frequency.     

Physiological and growth changes in micropropagated Citrus macrophylla explants due to salinity

Salinity is one of the major abiotic stresses affecting arable crops worldwide, and is the most stringent factor limiting plant distribution and productivity. In the present study, the possible use of in vitro culture to evaluate the growth and physiological responses to salt-induced stress in cultivated explants of Citrus macrophylla was analyzed. For this purpose, micropropagated adult explants were grown in proliferation and rooting media supplemented with different concentrations of NaCl. All growth parameters were decreased significantly by these NaCl treatments; this was accompanied by visible symptoms of salt injury in the proliferated shoots from 60 mM NaCl and in the rooted shoots from 40 mM NaCl. Malondialdehyde (MDA) increased with increasing salinity in proliferated shoots, indicating a rising degree of membrane damage. The concentration of total chlorophyll significantly decreased in the presence of NaCl, and this effect was more pronounced in the rooted explants. The Na⁺ and Cl⁻ concentrations in the explants increased significantly with the salinity level, but Cl⁻ levels were higher in the proliferated explants than in the rooted explants. For osmotic adjustment, high concentrations of compatible solutes (proline and quaternary ammonium compounds—QAC) accumulated in salt-stressed plants in proliferation, but differences were not observed in rooted explants. In proliferation, proline and QAC were highly correlated with the sodium and chloride concentrations in the explants, indicating a possible role of these compounds in osmotic adjustment. The plant concentrations of NO₃⁻, K⁺, Mg²⁺, Ca⁺ and Fe were also affected by the NaCl concentration of the medium. We suggest that the important deleterious effects in the in vitro explants of Citrus macrophylla grown at increasing NaCl concentrations were due mainly to toxic effects of saline ions, particularly Cl⁻, at the cellular level.     

Relation between Ion Accumulation of Salt-Sensitive and Isolated Stable Salt-Tolerant Cell Lines of Citrus aurantium

Four selected NaCl-tolerant cell lines of Sour orange (Citrus aurantium) were compared with the nonselected cell line in their growth and internal ion content of Na⁺, K⁺, and Cl⁻ when exposed to increasing NaCl concentrations. No difference was found among the various NaCl-tolerant cell lines in Na⁺ and Cl⁻ uptake, and all these cell lines took up similar or even larger amounts of Na⁺ and Cl⁻ than the NaCl-sensitive cell line. Exposure of cells of NaCl-sensitive and NaCl-tolerant lines to equal external concentrations of NaCl, resulted in a greater loss of K⁺ from the NaCl-sensitive cell line. This observation leads to the conclusion that growth and ability to retain high levels of internal K⁺ are correlated. Exposure of the NaCl-tolerant cell lines to salts other than NaCl resulted in even greater tolerance to Na₂SO₄, but rather poor tolerance to K⁺ introduced as either K₂SO₄ or KCl; the latter has a stronger inhibitory effect. The NaCl-sensitive cell line proved to be more sensitive to replacement of Na⁺ by K⁺. Analyses of internal Na⁺, K⁺, and Cl⁻ concentrations failed to identify any particular internal ion concentration which could serve as a reliable marker for salt tolerance.     

Development of callus and suspension cultures of potato resistant to NaCl and mannitol and their response to stress

Callus and suspension cultures adapted to various concentrations of NaCl or mannitol were developed from the cultivated potato Solanum tuberosum cv. Desire. Growth of the calli was less inhibited by mannitol than by iso-osmotic concentrations of NaCl. Reduction of growth by both NaCl and mannitol was considerably lower in osmotically adapted calli than in non-adapted ones. Salt-adapted suspension cultures that grew in the medium to which they had been originally adapted had a shorter lag in growth as well as a shorter time required to achieve the maximum growth, as compared with non-adapted cells. Suspension cultures adapted to NaCl concentrations higher than 150 mM were obtained only after preadaptation to osmotic stress. Adaptation of these cells was found to be stable. Accumulation of Na⁺ was lower and level of K⁺ was more stable in osmotically adapted than in non-adapted calli, when both were exposed to salt. Potassium level in NaCl-adapted calli exposed to saline medium was lower than that in non-adapted calli in standard medium. The maximum of Cl^– and Na⁺ accumulation was reached at higher external salt concentration in salt-adapted than in non-adapted suspension cultures. In both callus and suspension cultures, Cl^– accumulated more than Na⁺. Potassium level decreased more in non-adapted than in NaCl-adapted suspension cultures. The decrease of osmotic potential in osmotically adapted calli exposed to mannitol and in salt-adapted calli and suspension cultures exposed to salt was correlated to the increase of the external concentration. Such a correlation was not found in osmotically adapted calli exposed to salt. Non-electrolytes were found to be the main contributors to the decrease is osmotic potential in both callus and suspension cultures.     

Role of Internal Potassium in Maintaining Growth of Cultured Citrus Cells on Increasing NaCl and CaCl(2) Concentrations

Shamouti orange (Citrus sinensis L. Osbeck) salt-tolerant cells were grown under low water potential conditions induced by polyethylene glycol (PEG), NaCl, and CaCl₂. On the basis of equal osmotic potentials, PEG was the least inhibitory, NaCl next, and CaCl₂ the most inhibitory. The relation between growth capacity and ion content can be summarized as follows. (a) Internal K⁺ concentration was a major factor which changed in the presence of PEG, NaCl, and CaCl₂ and probably played a key role in determining growth capacity. (b) Internal concentrations of Na⁺, Ca²⁺, or Cl⁻ could not be directly correlated with growth. (C) Internal Mg²⁺ concentration could be significant only in the presence of high external Ca²⁺ concentrations. (d) The contribution of nitrate and phosphate to the internal osmoticum was negligible. The ratio of external (Ca²⁺)/(Na⁺)² concentration is crucial for growth. Ratios above 0.5 × 10⁻⁴ per millimolar gave maximal protection from adverse effects of NaCl. Growth capacity was found to be determined by the combination of (Ca²⁺)/(Na⁺)² ratio and the absolute external concentration of NaCl. However, a correlation between internal K⁺ concentration and growth capacity seemed independent of external NaCl concentration.     

Enhanced H Transport Capacity and ATP Hydrolysis Activity of the Tonoplast H-ATPase after NaCl Adaptation

Tonoplast enriched membrane vesicle fractions were isolated from unadapted and NaCl (428 millimolar) adapted tobacco cells (Nicotiana tabacum L. var Wisconsin 38). Polypeptides from the tonoplast enriched vesicle fractions were separated by SDS-PAGE and analyzed by Western blots using polyclonal antibodies to the 70 kilodalton subunit of the red beet tonoplast H⁺-ATPase. These antibodies cross-reacted exclusively to a tobacco polypeptide of an apparent molecular weight of 69 kilodaltons. The antibodies inhibited ATP-dependent, NO₃⁻ sensitive H⁺ transport into vesicles in tonoplast enriched membrane fractions from both unadapted and NaCl adapted cells. The relative H⁺ transport capacity per unit of 69 kilodalton subunit of the tonoplast ATPase of vesicles from NaCl adapted cells was fourfold greater than that observed for vesicles from unadapted cells. The increase in specific H⁺ transport capacity after adaptation was also observed for ATP hydrolysis.     

Ion fluxes and abscisic Acid-induced proline accumulation in barley leaf segments

The increase in proline induced by ABA, a process stimulated by NaCl or KCl in barley leaves, did not occur when Na⁺ (or K⁺) was present in the external medium as the gluconate salt, namely with an anion unable to permeate the plasma membrane. However, proline increase was restored, to different extents, by the addition of various chloride salts but not by ammonium chloride. Moreover, it was shown that the stimulation of the process by NaCl (or KCl) was variously affected by the presence of different salts; all the ammonium salts (10 millimolar NH₄⁺ concentration) inhibited this stimulation almost completely. Inhibition by NH₄⁺ was accompanied by a decreased Na⁺ influx (−40%). Also, in the case of Na-gluconate, Na⁺ uptake was reduced and the addition of Cl⁻ as the calcium or magnesium salt (but not as ammonium salt) restored both the ion influxes and the increase in proline typical of NaCl treatments. Both 4,4′-diisothiocyano-2,2′-disulfonic acid stilbene (DIDS), an anion transport inhibitor, and tetraethylammonium chloride (TEA), a K⁺ channels-blocking agent, caused, as well as with a reduction of ion influxes, an inhibition of the proline accumulation. The inhibition was practically total with 1 millimolar DIDS and about 80% with 20 millimolar TEA. A possible role of ion influxes in the process leading to the increase in proline induced by ABA is proposed.     

Induction of Increased Salt Tolerance in Sorghum bicolor by NaCl Pretreatment

Following 20 d of exposure to 75 or 150 mol m^–3 NaCl Sorghumbicolor (L.) Moench plants become capable of growing in mediumcontaining 300 mol m^–3 NaCl. Control plants, which havenot been pretreated, or plants pretreated for less than 20 ddie within 2 weeks when exposed to 300 mol m^–3 NaCl. Weconsider this induction of a capacity to survive in and toleratea high NaCl concentration as an adaptation to salinity. We suggestthat adaptation to salinity is more than osmotic adjustmentand that it takes longer to develop than osmotic adjustment.Concomitantly with the appearance of the ability to grow inhigh salinity, adaptation also comprises the development ofa capacity to regulate internal Na⁺ and Cl^– concentrations,even when external salinity is high. Shoot mean relative growthrates are similar for both control plants and for adapted plantsgrowing in 300 mol m^–3 NaCl, although their shoot Na⁺and Cl^– concentrations are quite different. Based on thesedata, we propose that adaptation of Sorghum to high salinityresults from a modulation of genome expression occurring duringextended exposure to non-lethal NaCl concentrations. Key words: Sorghum bicolor (L.) Moench, NaCl, salt tolerance, adaptation to salinity     

Properties of the Isolated Intact Chloroplast at Cytoplasmic K Concentrations : I. Light-Induced Cation Uptake into Intact Chloroplasts is Driven by an Electrical Potential Difference

Photosynthesis, stroma-pH, and internal K⁺ and Cl⁻ concentrations of isolated intact chloroplasts from Spinacia oleracea, as well as ion (K⁺, H⁺, Cl⁻) movements across the envelope, were measured over a wide range of external KCl concentrations (1-100 millimolar).

Isolated intact chloroplasts are a Donnan system which accumulates cations (K⁺ or added Tetraphenylphosphonium⁺) and excludes anions (Cl⁻) at low ionic strength of the medium. The internally negative dark potential becomes still more negative in the light as estimated by Tetraphenylphosphonium⁺ distribution. At 100 millimolar external KCl, potentials both in the light and in the dark and also the light-induced uptake of K⁺ or Na⁺ and the release of protons all become very small. Light-induced K⁺ uptake is not abolished by valinomycin suggesting that the K⁺ uptake is not primarily active. Intact chloroplasts contain higher K⁺ concentrations (112-157 millimolar) than chloroplasts isolated in standard media. Photosynthetic activity of intact chloroplasts is higher at 100 millimolar external KCl than at 5 to 25 millimolar. The pH optimum of CO₂ fixation at high K⁺ concentrations is broadened towards low pH values. This can be correlated with the observation that high external KCl concentrations at a constant pH of the suspending medium produce an increase of stroma-pH both in the light and in the dark. These results demonstrate a requirement of high external concentrations of monovalent cations for CO₂ fixation in intact chloroplasts.