The sodium transporter encoded by the HKT1;2 gene modulates sodium/potassium homeostasis in tomato shoots under salinity

Excessive soil salinity diminishes crop yield and quality. In a previous study in tomato, we identified two closely linked genes encoding HKT1‐like transporters, HKT1;1 and HKT1;2, as candidate genes for a major quantitative trait locus (kc7.1) related to shoot Na⁺/K⁺ homeostasis – a major salt tolerance trait – using two populations of recombinant inbred lines (RILs). Here, we determine the effectiveness of these genes in conferring improved salt tolerance by using two near‐isogenic lines (NILs) that were homozygous for either the Solanum lycopersicum allele (NIL17) or for the Solanum cheesmaniae allele (NIL14) at both HKT1 loci; transgenic lines derived from these NILs in which each HKT1;1 and HKT1;2 had been silenced by stable transformation were also used. Silencing of ScHKT1;2 and SlHKT1;2 altered the leaf Na⁺/K⁺ ratio and caused hypersensitivity to salinity in plants cultivated under transpiring conditions, whereas silencing SlHKT1;1/ScHKT1;1 had a lesser effect. These results indicate that HKT1;2 has the more significant role in Na⁺ homeostasis and salinity tolerance in tomato.     

The K+/H+ antiporter LeNHX2 increases salt tolerance by improving K+ homeostasis in transgenic tomato

The endosomal LeNHX2 ion transporter exchanges H⁺ with K⁺ and, to lesser extent, Na⁺. Here, we investigated the response to NaCl supply and K⁺ deprivation in transgenic tomato (Solanum lycopersicum L.) overexpressing LeNHX2 and show that transformed tomato plants grew better in saline conditions than untransformed controls, whereas in the absence of K⁺ the opposite was found. Analysis of mineral composition showed a higher K⁺ content in roots, shoots and xylem sap of transgenic plants and no differences in Na⁺ content between transgenic and untransformed plants grown either in the presence or the absence of 120 mm NaCl. Transgenic plants showed higher Na⁺/H⁺ and, above all, K⁺/H⁺ transport activity in root intracellular membrane vesicles. Under K⁺ limiting conditions, transgenic plants enhanced root expression of the high‐affinity K⁺ uptake system HAK5 compared to untransformed controls. Furthermore, tomato overexpressing LeNHX2 showed twofold higher K⁺ depletion rates and half cytosolic K⁺ activity than untransformed controls. Under NaCl stress, transgenic plants showed higher uptake velocity for K⁺ and lower cytosolic K⁺ activity than untransformed plants. These results indicate the fundamental role of K⁺ homeostasis in the better performance of LeNHX2 overexpressing tomato under NaCl stress.     

Alkali cation selectivity of the wheat root high-affinity potassium transporter HKT1

The wheat root high-affinity K⁺ transporter HKT1 functions as a sodium-coupled potassium co-uptake transporter. At toxic millimolar levels of sodium (Na⁺), HKT1 mediates low-affinity Na⁺ uptake while potassium (K⁺) uptake is blocked. In roots, low-affinity Na⁺ uptake and inhibition of K⁺ uptake contribute to Na⁺ toxicity. In the present study, the selectivity among alkali cations of HKT1 expressed in Xenopus oocytes and yeast was investigated under various ionic conditions at steady state. The data show that HKT1 is highly selective for uptake of the two physiologically significant alkali cations, K⁺ and Na⁺ over Rb⁺, Cs⁺ and Li⁺. In addition, Rb⁺ and Cs⁺, and an excess of extracellular K⁺ over Na⁺, are shown to partially reduce or block HKT1-mediated K⁺-Na⁺ uptake. Furthermore, K⁺, Rb⁺ and Cs⁺ also effectively reduce outward currents mediated by HKT1, thereby causing depolarizations. In yeast, HKT1 can produce high-affinity Rb⁺ uptake at approximately 15-fold lower rates than for K⁺. Rb⁺ influx in yeast can be mediated by the ability of the yeast plasma membrane proton pump to balance the 35-fold lower HKT1 conductance for Rb⁺. A model for HKT1 activity is presented involving a high-affinity K⁺ binding site and a high-affinity Na⁺ binding site, and competitive interactions of K⁺, Na⁺ and other alkali cations for binding to these two sites. Possible implications of the presented results for physiological K⁺ and Na⁺ uptake in plants are discussed.     

Early life stage salinity tolerance of wild and hatchery-reared juvenile pink salmon Oncorhynchus gorbuscha

Salinity tolerance in wild (Glendale) and hatchery (Quinsam) pink salmon Oncorhynchus gorbuscha (average mass 0·2 g) was assessed by measuring whole body [Na⁺] and [Cl^?] after 24 or 72 h exposures to fresh water (FW) and 33, 66 or 100% sea water (SW). Gill Na⁺, K⁺‐ATPase activity was measured following exposure to FW and 100% SW and increased significantly in both populations after a 24 h exposure to 100% SW. Whole body [Na⁺] and whole body [Cl^?] increased significantly in both populations after 24 h in 33, 66 and 100% SW, where whole body [Cl^?] differed significantly between Quinsam and Glendale populations. Extending the seawater exposure to 72 h resulted in no further increases in whole body [Na⁺] and whole body [Cl^?] at any salinity, but there was more variability among the responses of the two populations. Per cent whole body water (c. 81%) was maintained in all groups of fish regardless of salinity exposure or population, indicating that the increase in whole body ion levels may have been related to maintaining water balance as no mortality was observed in this study. Thus, both wild and hatchery juvenile O. gorbuscha tolerated abrupt salinity changes, which triggered an increase in gill Na⁺, K⁺‐ATPase within 24 h. These results are discussed in terms of the preparedness of emerging O. gorbuscha for the marine phase of their life cycle.     

Differences in shoot Na+ accumulation between two tomato species are due to differences in ion affinity of HKT1;2

HKT1 has been shown to be essential in Na⁺ homeostasis in plants. In this paper, we report the analysis of Na⁺ accumulation in different plant organs of two tomato species with contrasting salt tolerances: Solanum lycopersicum and Solanum pennellii. Furthermore, we relate these differences in Na⁺ accumulation between the two species to the differences in HKT1;2 transport kinetics and HKT1;2 expression. S. lycopersicum showed “Na⁺ excluder” behaviour, whereas S. pennellii showed “Na⁺ includer” behaviour. SlHKT1;2 expression, in contrast to SpHKT1;2 expression showed a significant effect of NaCl treatment, especially stems had a high increase in SlHKT1;2 expression. SlHKT1;2 promoter-GUS reporter gene analysis showed that SlHKT1;2 is expressed in the vasculature surrounding the roots and shoots of transformed Arabidopsis plants. In this paper, we present HKT1;2 protein sequences of both tomato species and provide evidence that both SlHKT1;2 and SpHKT1;2 are Na⁺ transporters. Our kinetic studies showed that SpHKT1;2, in comparison with SlHKT1;2, had a lower affinity for Na⁺. This low affinity of SpHKT1;2 correlated with higher xylem Na⁺ and higher accumulation of Na⁺ in stems and leaves of S. pennellii. Our findings demonstrate the importance of the understanding of transport characteristics of HKT1;2 transporters to improve the understanding of Na⁺ homeostasis in plants.     

Expressing the yeast HAL1 gene in tomato increases fruit yield and enhances K+/Na+ selectivity under salt stress

The yeast HAL1 gene facilitates K⁺/Na⁺ selectivity and salt tolerance of cells. Ectopic expression of HAL1 in transgenic tomato (Lycopersicon esculentum Mill.) plants minimized the reduction in fruit production caused by salt stress. Maintenance of fruit production by transgenic plants was correlated with enhanced growth under salt stress of calli derived from the plants. The HAL1 transgene enhanced water and K⁺ contents in both leaf calli and leaves in the presence of salt, which indicates that HAL1 functions in plants using a similar mechanism to that in yeast, namely by facilitating K⁺/Na⁺ selectivity under salt stress.     

Kinetics of chloride transport in the cyanobacterium Synechococcus R-2 (Anacystis nidulans, S. leopoliensis) PCC 7942

The kinetics of the light-driven Cl^? uptake pump of Synechococcus R-2 (PCC 7942) were investigated. The kinetics of Cl^? uptake were measured in BG-11 medium (pH_o, 7·5; [K⁺]_o, 0·35 mol m^?3; [Na⁺]_o, 18 mol m^?3; [Cl^?]_o, 0·508 mol m^?3) or modified media based on the above. Net³⁶Cl^? fluxes (?Cl^?_o,i) followed Michaelis-Menten kinetics and were stimulated by Na⁺ [18 mol m^?3 Na⁺ BG-11 ?Cl^?_max= 3·29±0·60 (49) nmol m^?2 s^?1 versus Na⁺-free BG-11 ?Cl^?_max= 1·02±0·13 (54) nmol m^?2 s^?1] but the Km was not significantly different in the presence or absence of Na⁺ at pH_o 10; the Km was lower, but not affected by the presence or absence of Na⁺ [Km = 22·3±3·54 (20) mmol m^?3]. Na⁺ is a non-competitive activator of net ?Cl^?_o,i. High [K⁺]_o (18 mol m^?3) did not stimulate net ?Cl^?_o,i or change the Km in Na⁺-free medium. High [K⁺]_o (18 mol m^?3) added to Na⁺ BG-11 medium decreased net ?Cl^?_o,i [18 mol m^?3K⁺ BG-11; ?Cl^?_max= 2·50±0·32 (20) nmol m^?2 s^?1 versus BG-11 medium; ?Cl^?_max= 3·35±0·56 (20) nmol m^?2 s^?1] but did not affect the Km 55·8±8·100 (40) mmol m^?3]. Na⁺-stimulation of net ?Cl^?_o,i followed Michaelis-Menten kinetics up to 2–5 mol m^?3 [Na⁺]_o but higher concentrations were inhibitory. The Km for Na⁺-stimulation of net ?Cl^?_o,i [K_1/2(Na⁺)] was different at 47 mmol m^?3 [Cl^?]_o (K_1/2[Na⁺] = 123±27 (37) mmol m^?3]. Li⁺ was only about one-third as effective as Na⁺ in stimulating Cl^? uptake but the activation constant was similar [K_1/2(Li⁺) = 88±46 (16) mmol m^?3]. Br^? was a competitive inhibitor of Cl^? uptake. The inhibition constant (Ki) was not significantly different in the presence and absence of Na⁺. The overall Ki was 297±23 (45) mmol m^?3. The discrimination ratio of Cl^? over Br^? (δCl^?/δBr^?) was 6·38±0·92 (df = 147). Synechococcus has a single Na⁺-stimulated Cl^? pump because the Km of the Cl^? transporter and its discrimination between Cl^? and Br^? are not significantly different in the presence and absence of Na⁺. The Cl^? pump is probably driven by ATP.     

Modeling of the Interaction of Na+ and K+ with the Binding of the Cocaine Analogue 3β-(4-[125I]Iodophenyl)tropane-2β-Carboxylic Acid Isopropyl Ester to the Dopamine Transporter

Abstract: The present study examines the interaction of Na⁺ and K⁺ with the binding of the cocaine analogue 3β-(4-[¹²⁵I]iodophenyl)tropane-2β-carboxylic acid isopropyl ester to dopamine transporters (DATs) in rat striatal synaptosomal membranes at 37°C. The binding increases with [Na⁺] from 10 to 100 mM and decreases with higher [Na⁺]. The presence of K⁺ reduces the maximal stimulatory effect of Na⁺ and causes a nonlinear EC₅₀ shift for Na⁺. K⁺ strongly inhibits the binding at low [Na⁺]. Increasing [Na⁺] produces a linear IC₅₀ shift for K⁺. Saturation analysis indicates a single binding site changing its affinity for the radioligand depending on [K⁺]/[Na⁺] ratio in the assay buffer. A reduced B_max was observed in the presence of 10 mM Na⁺ and 30 mM K⁺. Both high [Na⁺] and high [K⁺] accelerate the dissociation of the binding, and K⁺-induced acceleration was abolished by increasing [Na⁺]. Least squares model fitting of equilibrium data and kinetic analysis of dissociation rates reveal competitive interactions between Na⁺ and K⁺ at two sites allosterically linked on the DAT: One site mediates the stimulatory effect of Na⁺, and the other site involves the radioligand binding and the inhibitory effect of cations on the binding. Various uptake blockers and substrates, dopamine in particular, display reduced potency in inhibiting the binding at a higher [K⁺]/[Na⁺] ratio.     

ZxAKT1 is essential for K+ uptake and K+/Na+ homeostasis in the succulent xerophyte Zygophyllum xanthoxylum

The inward‐rectifying K⁺ channel AKT1 constitutes an important pathway for K⁺ acquisition in plant roots. In glycophytes, excessive accumulation of Na⁺ is accompanied by K⁺ deficiency under salt stress. However, in the succulent xerophyte Zygophyllum xanthoxylum, which exhibits excellent adaptability to adverse environments, K⁺ concentration remains at a relatively constant level despite increased levels of Na⁺ under salinity and drought conditions. In this study, the contribution of ZxAKT1 to maintaining K⁺ and Na⁺ homeostasis in Z. xanthoxylum was investigated. Expression of ZxAKT1 rescued the K⁺‐uptake‐defective phenotype of yeast strain CY162, suppressed the salt‐sensitive phenotype of yeast strain G19, and complemented the low‐K⁺‐sensitive phenotype of Arabidopsis akt1 mutant, indicating that ZxAKT1 functions as an inward‐rectifying K⁺ channel. ZxAKT1 was predominantly expressed in roots, and was induced under high concentrations of either KCl or NaCl. By using RNA interference technique, we found that ZxAKT1‐silenced plants exhibited stunted growth compared to wild‐type Z. xanthoxylum. Further experiments showed that ZxAKT1‐silenced plants exhibited a significant decline in net uptake of K⁺ and Na⁺, resulting in decreased concentrations of K⁺ and Na⁺, as compared to wild‐type Z. xanthoxylum grown under 50 mm NaCl. Compared with wild‐type, the expression levels of genes encoding several transporters/channels related to K⁺/Na⁺ homeostasis, including ZxSKOR, ZxNHX, ZxSOS1 and ZxHKT1;1, were reduced in various tissues of a ZxAKT1‐silenced line. These findings suggest that ZxAKT1 not only plays a crucial role in K⁺ uptake but also functions in modulating Na⁺ uptake and transport systems in Z. xanthoxylum, thereby affecting its normal growth.     

Sodium exclusion QTL associated with improved seedling growth in bread wheat under salinity stress

Worldwide, dryland salinity is a major limitation to crop production. Breeding for salinity tolerance could be an effective way of improving yield and yield stability on saline-sodic soils of dryland agriculture. However, this requires a good understanding of inheritance of this quantitative trait. In the present study, a doubled-haploid bread wheat population (Berkut/Krichauff) was grown in supported hydroponics to identify quantitative trait loci (QTL) associated with salinity tolerance traits commonly reported in the literature (leaf symptoms, tiller number, seedling biomass, chlorophyll content, and shoot Na⁺ and K⁺ concentrations), understand the relationships amongst these traits, and determine their genetic value for marker-assisted selection. There was considerable segregation within the population for all traits measured. With a genetic map of 527 SSR-, DArT- and gene-based markers, a total of 40 QTL were detected for all seven traits. For the first time in a cereal species, a QTL interval for Na⁺ exclusion (wPt-3114-wmc170) was associated with an increase (10%) in seedling biomass. Of the five QTL identified for Na⁺ exclusion, two were co-located with seedling biomass (2A and 6A). The 2A QTL appears to coincide with the previously reported Na⁺ exclusion locus in durum wheat that hosts one active HKT1;4 (Nax1) and one inactive HKT1;4 gene. Using these sequences as template for primer design enabled mapping of at least three HKT1;4 genes onto chromosome 2AL in bread wheat, suggesting that bread wheat carries more HKT1;4 gene family members than durum wheat. However, the combined effects of all Na⁺ exclusion loci only accounted for 18% of the variation in seedling biomass under salinity stress indicating that there were other mechanisms of salinity tolerance operative at the seedling stage in this population. Na⁺ and K⁺ accumulation appear under separate genetic control. The molecular markers wmc170 (2A) and cfd080 (6A) are expected to facilitate breeding for salinity tolerance in bread wheat, the latter being associated with seedling vigour.     

Cytosolic sodium regulation in mouse cortical astrocytes and its dependence on potassium and bicarbonate

Sodium plays a major role in different astrocytic functions, including maintenance of ion homeostasis and uptake of neurotransmitters and metabolites, which are mediated by different Na⁺-coupled transporters. In the current study, the role of an electrogenic sodium-bicarbonate cotransporter (NBCe1), a sodium-potassium-chloride transporter 1 (NKCC1) and sodium-potassium ATPase (Na⁺-K⁺-ATPase) for the maintenance of [Na⁺]_i was investigated in cultured astrocytes of wild-type (WT) and of NBCe1-deficient (NBCe1-KO) mice using the Na⁺-sensitive dye, asante sodium green-2. Our results suggest that cytosolic Na⁺ was higher in the presence of CO₂/HCO₃⁻ (15 mM) than CO₂/HCO₃⁻-free, HEPES-buffered solution in WT, but not in NBCe1-KO astrocytes (12 mM). Surprisingly, there was a strong dependence of cytosolic [Na⁺] on the extracellular [HCO₃⁻] attributable to NBCe1 activity. Pharmacological blockage of NKCC1 with bumetanide led to a robust drop in cytosolic Na⁺ in both WT and NBCe1-KO astrocytes by up to 6 mM. There was a strong dependence of the cytosolic [Na⁺] on the extracellular [K⁺]. Inhibition of the Na⁺-K⁺-ATPase led to larger increase in cytosolic Na⁺, both in the absence of K⁺ as compared with the presence of ouabain and in NBCe1-KO astrocytes as compared with WT astrocytes. Our results show that cytosolic Na⁺ in mouse cortical astrocytes can vary considerably and depends greatly on the concentrations of HCO₃⁻ and K⁺, attributable to the activity of the Na⁺-K⁺-ATPase, of NBCe1 and NKCC1.     

The AtNHX1 exchanger mediates potassium compartmentation in vacuoles of transgenic tomato

NHX‐type antiporters in the tonoplast have been reported to increase the salt tolerance of various plants species, and are thought to mediate the compartmentation of Na⁺ in vacuoles. However, all isoforms characterized so far catalyze both Na⁺/H⁺ and K⁺/H⁺ exchange. Here, we show that AtNHX1 has a critical involvement in the subcellular partitioning of K⁺, which in turn affects plant K⁺ nutrition and Na⁺ tolerance. Transgenic tomato plants overexpressing AtNHX1 had larger K⁺ vacuolar pools in all growth conditions tested, but no consistent enhancement of Na⁺ accumulation was observed under salt stress. Plants overexpressing AtNHX1 have a greater capacity to retain intracellular K⁺ and to withstand salt‐shock. Under K⁺‐limiting conditions, greater K⁺ compartmentation in the vacuole occurred at the expense of the cytosolic K⁺ pool, which was lower in transgenic plants. This caused the early activation of the high‐affinity K⁺ uptake system, enhanced K⁺ uptake by roots, and increased the K⁺ content in plant tissues and the xylem sap of transformed plants. Our results strongly suggest that NHX proteins are likely candidates for the H⁺‐linked K⁺ transport that is thought to facilitate active K⁺ uptake at the tonoplast, and the partitioning of K⁺ between vacuole and cytosol.     

Effects of potassium on the anion and cation contents of primary cultures of mouse astrocytes and neurons

Inastrocytes, as [K⁺]_o was increased from 1.2 to 10 mM, [K⁺]_i and [Cl^–]_i were increased, whereas [Na⁺]_i was decreased. As [K⁺]_o was increased from 10 to 60 mM, intracellular concentration of these three ions showed no significant change. When [K⁺]_o was increased from 60 to 122 mM, an increase in [K⁺]_i and [Cl^–]_i and a decrease in [Na⁺]_i were observed.Inneurons, as [K⁺]_o was increased from 1.2 to 2.8 mM, [Na⁺]_i and [Cl^–]_i were decreased, whereas [K⁺]_i was increased. As [K⁺]_o was increased from 2.8 to 30 mM, [K⁺]_i, [Na⁺]_i and [Cl^–]_i showed no significant change. When [K⁺]_o was increased from 30 to 122 mM, [K⁺]_i and [Cl^–]_i were increased, whereas [Na⁺]_i was decreased. Inastrocytes, pH_i increased when [K⁺]_o was increased. Inneurons, there was a biphasic change in pH_i. In lower [K⁺]_o (1.2–2.8 mM) pH_i decreased as [K⁺]_o increased, whereas in higher [K⁺]_o (2.8–122 mM) pH_i was directly related to [K⁺]_o. In bothastrocytes andneurons, changes in [K⁺]_o did not affect the extracellular water content, whereas the intracellular water content increased as the [K⁺]_o increased. Transmembrane potential (E_m) as measured with Tl-204 was inversely related to [K⁺]_o between 1.2 and 90 mM, a ten-fold increase in [K⁺]_o depolarized the astrocytes by about 56 mV and the neurons about 52 mV. The E_m values measured with Tl-204 were close to the potassium equilibrium potential (E_k) except those in neurons at lower [K⁺]_o. However, they were not equal to the chloride equilibrium potential (E_Cl) at [K⁺]_o lower than 30 mM in both astrocytes and neurons. Results of this study demonstrate that alteration of [K⁺]_o produced different changes in [K⁺]_i, [Na⁺]_i, [Cl^–]_i, and pH_i in astrocytes and neurons. The data show that astrocytes can adapt to alterations in [K⁺]_o, in such a way to maintain a more suitable environment for neurons.     

Effects of ouabain on proliferation of human endothelial cells correlate with Na<Superscript>+</Superscript>,K<Superscript>+</Superscript>-ATPase activity and intracellular ratio of Na<Superscript>+</Superscript> and K<Superscript>+</Superscript>

Side-by-side with inhibition of the Na⁺,K⁺-ATPase ouabain and other cardiotonic steroids (CTS) can affect cell functions by mechanisms other than regulation of the intracellular Na⁺ and K⁺ ratio ([Na⁺]_i/[K⁺]_i). Thus, we compared the doseand time-dependences of the effect of ouabain on intracellular [Na⁺]_i/[K⁺]_i ratio, Na⁺,K⁺-ATPase activity, and proliferation of human umbilical vein endothelial cells (HUVEC). Treatment of the cells with 1-3 nM ouabain for 24-72 h decreased the [Na⁺]_i/[K⁺]_i ratio and increased cell proliferation by 20-50%. We discovered that the same ouabain concentrations increased Na⁺,K⁺-ATPase activity by 25-30%, as measured by the rate of ⁸⁶Rb⁺ influx. Higher ouabain concentrations inhibited Na⁺,K⁺-ATPase, increased [Na⁺]_i/[K⁺]_i ratio, suppressed cell growth, and caused cell death. When cells were treated with low ouabain concentrations for 48 or 72 h, a negative correlation between [Na⁺]_i/[K⁺]_i ratio and cell growth activation was observed. In cells treated with high ouabain concentrations for 24 h, the [Na⁺]_i/[K⁺]_i ratio correlated positively with proliferation inhibition. These data demonstrate that inhibition of HUVEC proliferation at high CTS concentrations correlates with dissipation of the Na⁺ and K⁺ concentration gradients, whereas cell growth stimulation by low CTS doses results from activation of Na⁺,K⁺-ATPase and decrease in the [Na⁺]_i/[K⁺]_i ratio.     

Hydrogen peroxide as a mediator of 5‐aminolevulinic acid‐induced Na+ retention in roots for improving salt tolerance of strawberries

To explore the mechanisms of 5‐aminolevulinic acid (ALA)‐improved plant salt tolerance, strawberries (Fragaria × ananassa Duch. cv. ‘Benihoppe’) were treated with 10 mg l^?1 ALA under 100 mmol l^?1 NaCl stress. We found that the amount of Na⁺ increased in the roots but decreased in the leaves. Laser scanning confocal microscopy (LSCM) observations showed that ALA‐induced roots had more Na⁺ accumulation than NaCl alone. Measurement of the xylem sap revealed that ALA repressed Na⁺ concentrations to a large extent. The electron microprobe X‐ray assay also confirmed ALA‐induced Na⁺ retention in roots. qRT‐PCR showed that ALA upregulated the gene expressions of SOS1 (encoding a plasma membrane Na⁺/H⁺ antiporter), NHX1 (encoding a vacuolar Na⁺/H⁺ antiporter) and HKT1 (encoding a protein of high‐affinity K⁺ uptake), which are associated with Na⁺ exclusion in the roots, Na⁺ sequestration in vacuoles and Na⁺ unloading from the xylem vessels to the parenchyma cells, respectively. Furthermore, we found that ALA treatment reduced the H₂O₂ content in the leaves but increased it in the roots. The exogenous H₂O₂ promoted plant growth, increased root Na⁺ retention and stimulated the gene expressions of NHX1, SOS1 and HKT1. Diphenyleneiodonium (DPI), an inhibitor of H₂O₂ generation, suppressed the effects of ALA or H₂O₂ on Na⁺ retention, gene expressions and salt tolerance. Therefore, we propose that ALA induces H₂O₂ accumulation in roots, which mediates Na⁺ transporter gene expression and more Na⁺ retention in roots, thereby improving plant salt tolerance.     

Kinetic study on the effects of intracellular K++ and Na++ on Na++, K++, Cl+− cotransport of HeLa cells by Rb++ influx determination

Summary The effects of intracellular K⁺ and Na⁺ (K⁺ c, Na⁺ c) on the Na⁺,K⁺,Cl^+– cotransport pathway of HeLa cells were studied by measuring ouabain-insensitive, furosemide-sensitive Rb⁺ influx (JRb) at various intracellular concentrations of K⁺ and Na⁺ ([K⁺]c, [Na⁺]c). When [K⁺]c was increased and [Na⁺]c was decreased, keeping the sums of their concentrations almost constant, JRb as a function of the extracellular Rb⁺ or Na⁺ concentration ([Rb⁺]e, [Na⁺]e) was stimulated. However, the apparent K _0.5 for Rb⁺ e or Na⁺ e remained unchanged and the ratio of the apparent K ^+0.5 for K⁺ c and the apparent K _i for Na⁺ c was larger than 1. When JRb was increased by hypertonicity by addition of 200 mM mannitol, the apparent maximum JRb increased without change in the apparent K _0.5 for Rb⁺ e. These results show that K⁺ c stimulates and Na⁺ c inhibits JRb, without change in the affinities of the pathway for Rb⁺ e and Na⁺ e. The affinity for K⁺ c is slightly lower than that for Na⁺ c. Hypertonicity enhances JRb without any change in the affinity for Rb⁺ e. We derived a kinetic equation for JRb with respect to K⁺ c and Na⁺ c and proposed a general and a special model of the pathway. The special model suggests that, in HeLa cells, JRb takes place when Rb⁺ e binds to the external K⁺ binding site of the pathway after the binding of K⁺ c to the internal regulatory site.We thank Mr. T. Masuya for technical assistance. This study was supported in part by a Grant-in-Aid for Scientific Research on Priority Areas (No. 03202136) from the Japanese Ministry of Education, Science and Culture.