Na+-K+ transport in roots under salt stress

Salinity causes billion dollar losses in annual crop production. So far, the main avenue in breeding crops for salt tolerance has been to reduce Na⁺ uptake and transport from roots to shoots. Recently we have demonstrated that retention of cytosolic K⁺ could be considered as another key factor in conferring salt tolerance in plants. A subsequent study has shown that Na⁺-induced K⁺ efflux in barley root epidermis occurs primarily via outward rectifying K⁺ channels (KORC). Surprisingly, expression of KORC was similar in salt- tolerant and sensitive genotypes. However, the former were able to better oppose Na⁺-induced depolarization via enhanced activity of plasma membrane H⁺-ATPase (thus minimizing K⁺ leak from the cytosol). In addition to highly K⁺-selective KORC channels, activities of several types of non-selective cation channels were detected at depolarizing potentials. Here we show that the expression of one of them, NORC, was significantly lower in salt-tolerant genotypes. As NORC is capable of mediating K⁺ efflux coupled to Na⁺ influx, we suggest that the restriction of its activity could be beneficial for plants under salt stress.Key words: salinity tolerance, barley, ion flux, K⁺ homeostasis, KOR, non-selective channels, patch-clamp     

等渗盐胁迫下Na^＋和Cl^-对大豆幼苗光合作用的离子效应

研究和比较了等渗(-0.53MPa)的PEG-6000、NaCl、钠盐(无Cl-)和氯化物(无Na )溶液处理6d对栽培大豆品种‘Lee68’(耐盐性较强)和‘N23674’(耐盐性较弱)幼苗光合作用的离子效应。结果表明:PEG-6000处理使两品种叶片叶绿素含量和Rubisco活性较对照低,但降幅不如同样渗透压的NaCl、钠盐(无Cl-)和氯化物(无Na )溶液明显。PSII最大光化学效率(Fv/Fm)、电子传递速率(ETR)和PSII光化学的有效量子产额(Fv'/Fm')在PEG-6000处理2d和6d时显著下降,但在3种等渗盐处理下,多显著下降。两品种叶片气孔导度(Gs)和净光合速率(Pn)在4种胁迫处理下均显著下降,其中在3种盐处理下更明显,但胞间CO2浓度(Ci)仅在PEG-6000处理时下降,在盐处理下反而升高。两品种叶片叶绿素含量、Rubisco活性、Fv/Fm、ETR、Fv'/Fm'、Pn、Gs等在氯化物(无Na )溶液处理的下降幅度和叶绿体中Cl-含量及其与Na 总量的增加幅度均大于钠盐(无Cl-)处理的,在耐盐性弱的‘N23674’品种中更明显。可见,在NaCl胁迫对栽培大豆幼苗光合作用的毒害效应中,渗透胁迫较轻,离子毒害较重,其中Cl-的毒害大于Na 的。     

Dynamic QTL analysis of the Na+ content,K+ content,and Na+/K+ ratio in rice roots during the field growth under salt stress

Rice (Oryza sativa L.) is seriously impacted by global soil salinization. To determine the quantitative trait loci (QTLs) related to salt tolerance in rice roots, F_2:3 and BC₁F_2:3 populations derived from a cross between the cv. Dongnong 425 of high quality and yield and the salt-tolerant cv. Changbai 10, were studied at different development stages. Two genetic linkage maps of F_2:3 and BC1F_2:3 populations were constructed. A 66 mM NaCl solution was used to irrigate the field and to analyze the dynamic QTL of some rice root traits. Using unconditional and conditional QTL mapping methods, 30 unconditional QTLs and 16 conditional QTLs related to the 6 root traits were detected on the 9 rice chromosomes during different developmental stages. Fourteen pairs of unconditional and conditional QTLs were detected at the identical developmental stage in the identical population. A number of QTLs were detected at different developmental stages, however, many did not appear at the last stage. Remarkably, qRKC1 appeared continuously at multiple stages in both the populations suggesting its key role in regulating the salt tolerance of rice roots.     

Coordination of AtHKT1;1 and AtSOS1 facilitates Na+ and K+ homeostasis in Arabidopsis thaliana under salt stress

Reducing Na⁺ accumulation and maintaining K⁺ stability in plant is one of the key strategies for improving salt tolerance. AtHKT1;1 and AtSOS1 are not only the salt tolerance determinants themselves, but also mediate K⁺ uptake and transport indirectly. To assess the contribution of AtHKT1;1 and AtSOS1 to Na⁺ homeostasis and K⁺ nutrition in plant, net Na⁺ and K⁺ uptake rate, Na⁺ and K⁺ distributions in Arabidopsis thaliana wild type (WT), hkt1;1 mutant (athkt1;1) and sos1 mutant (atsos1) were investigated. Results showed that under 2.5 mM K⁺ plus 25 or 100 mM NaCl, athkt1;1 shoot concurrently accumulated more Na⁺ and less K⁺ than did WT shoot, suggesting that AtHKT1;1 was critical for controlling Na⁺ and K⁺ distribution in plant; while atsos1 root accumulated more Na⁺ and absorbed lower K⁺ than did WT root, implying that AtSOS1 was determiner of Na⁺ excretion and K⁺ acquisition. Under 0.01 mM K⁺, athkt1;1 absorbed lower Na⁺ than did WT with 100 mM NaCl, suggesting that AtHKT1;1 is involved in Na⁺ uptake in roots; while atsos1 shoot accumulated less Na⁺ than did WT shoot no matter with 25 or 100 mM NaCl, implying that AtSOS1 played a key role in controlling long-distance Na⁺ transport from root to shoot. We present a model in which coordination of AtHKT1;1 and AtSOS1 facilitates Na⁺ and K⁺ homeostasis in A. thaliana under salt stress: under the normal K⁺, the major function of AtHKT1;1 is Na⁺ unloading and AtSOS1 is mainly involved in Na⁺ exclusion, whereas under the low K⁺, AtHKT1;1 may play a dominant role in Na+ uptake and AtSOS1 may be mainly involved in Na⁺ loading into the xylem.     

Effect of Divalent Cations on Na+ Permeability of Chara corallina and Freshwater Grown Chara buckellii

The effect of elevated Na⁺ concentration on Na⁺ permeability(P_Na) and Na⁺ influx in the presence of two levels of externaldivalent cations was determined in Chara corallina and freshwater-culturedChara buckellii. When Na⁺ in the medium was increased from 1.0to 70 mol m^–3, Na⁺ influx increased in both species ifCa²⁺ was low (0.1 mol m–3). If Ca²⁺ was increased to 7.0mol m^–3 when Na⁺ was increased, Na⁺ influx remained atthe low control level in C. corallina, and showed only a temporaryincrease in C. buckellii. Mg²⁺ was a better substitute for Ca²⁺in C. buckellii than in C. corallina. Na⁺ permeability data suggest that when the external Ca²⁺ concentrationis low, P_Na does not increase in the presence of elevated NaCl;the increase in Na⁺ influx appears to be due to the increasein external Na⁺ concentration alone. Ca^{2 +} supplementation appearsto decrease P_Na whereas supplemental Mg²⁺ has no effect. Na⁺ effluxes were computed from previously determined net fluxesand the influxes. It was found that for both species, fluxesin both directions were stimulated in response to all experimentaltreatments, but Na⁺ influx always exceeded efflux. This resultedin net Na⁺ accumulation in the vacuoles of both species. The results are discussed with reference to net flux and electrophysiologicaldata obtained previously under identical conditions, as wellas the comparative salinity tolerance of both species and theNa⁺/divalent cation ratio. Key words: Na⁺ influx, Na⁺ tolerance, membrane potential, permeability, Chara     

盐胁迫对4树种叶片中K^＋和Na^＋的影响及其耐盐能力的评价

通过盆栽试验,对我国南方银杏（Ginkgo biloba L.)、侧柏[Patycladus orentalis(L.)Franco]、火炬松（Pinus taeda L.)和剌槐(Robinia Pseudoacacia L.)4造林树种苗木叶片中K^ 、N^ 浓度浓度和N^ ／K^ 比进行测定,盐处理水平为0.0％（CK）、0.1%、0.3%和0.5％。随着盐浓度的提高,各树种叶片中Na^ 浓度和N^ ／K^ 有逐渐升高的趋势,但K^ 浓度变化较小;随着盐胁迫时间的推移,各树种叶片中Na^ 、K^ 浓度和Na^ /K^ 比都没有明显的变化规律。方差分析和多层比较表明,侧柏、火炬松和剌槐叶片中Na^ 、K^ 浓度和N^ ／K^ 比在4组处理间的差异均达显著或极显著水平。4树种中剌槐和侧柏的耐盐能力最强,银杏次之,火炬松最弱。     

22Na+ fluxes in thymic lymphocytes. I. Na+/Na+ and Na+/H+ exchange through an amiloride-insensitive pathway

The Na+ transport pathways of normal rat thymocytes were investigated. Na+ conductance was found to be lower than K+ conductance, which is consistent with reported values of membrane potential. In contrast, the isotopically measured Na+ permeability was greater than 10-fold higher than that of K+, which indicates that most of the flux is electroneutral. Cotransport with Cl- (or K+ and Cl-) and countertransport with Ca2+ were ruled out by ion substitution experiments and use of inhibitors. Countertransport for Na+ or H+ through the amiloride-sensitive antiport accounts for only 15-20% of the resting influx. In the presence of amiloride, 22Na+ uptake was increased in Na+-loaded cells, which suggests the existence of Na+/Na+ countertransport. Cytoplasmic pH determinations using fluorescent probes indicated that under certain conditions this amiloride-resistant system will also exchange Na+ for H+, as evidenced by an internal Na+- dependent acidification is proportional to internal [Na+] but inversely related to extracellular [Na+]. Moreover, 22Na+ uptake is inhibited by increasing external [H+]. The results support the existence of a substantial amiloride-insensitive, electroneutral cation exchange system capable of transporting Na+ and H+.     

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.     

Severe salt stress in Vaccinium myrtillus (L.) in response to Na+ ion toxicity

NaCl is spread on the northern roads as a winter de-icing agent. Na⁺ stress may occur in roadside forests especially during growing season, because snowmelt increases Na⁺ concentrations in the roadside forest soil. To simulate summertime Na⁺ stress and its effect on viability and anthocyanin concentrations, bilberry (Vaccinium myrtillus L.) plants were subjected to NaCl (0, 6, 30 and 60 g m⁻²) in a boreal mesic heath forest in northern Finland (65°N). It was hypothesised that Na⁺ stress decreases tissue water content (TWC) in below- and aboveground stems, where resulting water stress increases anthocyanin level for osmotic regulation. Uptake of Na⁺ from the soil to the below- and aboveground stems was detected by a fast sequential atomic absorption spectrometry. Na⁺ accumulated in belowground stems, but it was translocated into the aboveground stems to a lesser extent. At the end of the growing season, TWC increased in belowground stems and decreased in aboveground stems. Also anthocyanin concentrations decreased in aboveground stems. The viability of the aboveground stems decreased to 30% along with the Na⁺ accumulation. Despite the strong viability response in aboveground stems, TWC was constant in the middle of the growing season and decreased relatively little at the end of the growing season. It is thus proposed that direct effect of Na⁺ ions, rather than Na⁺-induced water stress, is the primary reason behind the strong viability response and a severe salt stress in bilberry.     

Na+- and cGMP-induced Ca2+ fluxes in frog rod photoreceptors

We have examined the Ca2+ content and pathways of Ca2+ transport in frog rod outer segments using the Ca2+-indicating dye arsenazo III. The experiments employed suspensions of outer segments of truncated, but physiologically functional, frog rods (OS-IS), intact isolated outer segments (intact OS), and leaky outer segments (leaky OS with a plasma membrane leaky to small solutes, but with sealed disk membranes). We observed the following. Intact OS or OS-IS isolated and purified in Percoll-Ringer's solution contained an average of 2.2 mM total Ca2+, while leaky OS contained 2.0 mM total Ca2+. This suggests that most of the Ca2+ in OS-IS is contained inside OS disks. Phosphodiesterase inhibitors increased the Ca2+ content to approximately 4.2 mM in intact OS or OS-IS, whereas the Ca2+ content of leaky OS was not altered. Na-Ca exchange was the dominant pathway for Ca2+ efflux in both intact and leaky OS/OS-IS. The rate of Na-Ca exchange in intact OS/OS-IS was half-maximal between 30 and 50 mM Na+; at 50 mM Na+, this amounted to 5.8 X 10(7) Ca2+/OS X s or 0.05 mM total Ca2+/s. This is much larger than the Ca2+ component of the dark current. Other alkali cations could not replace Na+ in Na-Ca exchange in either OS-IS or leaky OS. They inhibited the rate of Na-Ca exchange (K greater than or equal to Rb greater than Cs greater than or equal to Li greater than TMA) and, as the inhibition became greater, a delay developed in the onset of Na-Ca exchange. The inhibition of Na-Ca exchange by alkali cations correlates with the prolonged duration of the photoresponse induced by these cations (Hodgkin, A. L., P. A. McNaughton, and B. J. Nunn. 1985. Journal of Physiology. 358:447-468). In addition to Na-Ca exchange, disk membranes in leaky OS showed a second pathway of Ca2+ transport activated by cyclic GMP (cGMP). The cGMP-activated pathway required the presence of alkali cations and had a maximal rate of 9.7 X 10(6) Ca2+/OS X s. cGMP caused the release of only 30% of the total Ca2+ from leaky OS. The rate of Na-Ca exchange in leaky OS amounted to 1.9 X 10(7) Ca2+/OS X s.(ABSTRACT TRUNCATED AT 400 WORDS)     

Exclusion of Na+ via sodium ATPase (PpENA1) ensures normal growth of Physcomitrella patens under moderate salt stress

The bryophyte Physcomitrella patens is unlike any other plant identified to date in that it possesses a gene that encodes an ENA-type Na(+)-ATPase. To complement previous work in yeast (Saccharomyces cerevisiae), we determined the importance of having a Na(+)-ATPase in planta by conducting physiological analyses of PpENA1 in Physcomitrella. Expression studies showed that PpENA1 is up-regulated by NaCl and, to a lesser degree, by osmotic stress. Maximal induction is obtained after 8 h at 60 mm NaCl or above. No other abiotic stress tested led to significant increases in PpENA1 expression. In the gametophyte, strong expression was confined to the rhizoids, stem, and the basal part of the leaf. In the protonemata, expression was ubiquitous with a few filaments showing stronger expression. At 100 mm NaCl, wild-type plants were able to maintain a higher K(+)-to-Na(+) ratio than the PpENA1 (ena1) knockout gene, but at higher NaCl concentrations no difference was observed. Although no difference in chlorophyll content was observed between ena1 and wild type at 100 mm NaCl, the impaired Na(+) exclusion in ena1 plants led to an approximately 40% decrease in growth.     

NaCl胁迫下棉花体内 Na~+ 、K~+分布与耐盐性

采用盐化土壤方法 ,选择苗期耐盐性较强的陆地棉品种枝棉 3号和中棉所 1 9及耐盐性较弱的品种泗棉 2号和苏棉 1 2号 ,研究了盐胁迫下棉苗体内 Na+、K+的运输和分配与耐盐性的关系。结果表明 ,耐盐品种根系具有一定的截留 Na+作用。棉花地上部盐分器官水平上的区域化分布特征明显 :2 0 0 mmol/L Na Cl胁迫下 ,枝棉 3号叶片中的 Na+含量显著低于泗棉 2号 ,茎及叶柄中的 Na+含量显著高于泗棉 2号 ;棉株地上部茎、叶柄、叶片中的 Na+含量分别由下而上逐渐减小 ,相同节位的茎、叶柄中的 Na+含量大于叶片 ,枝棉 3号更显著。1 0 0 mmol/L和 1 50 mmol/L Na Cl胁迫下 ,枝棉 3号和中棉所 1 9K+/Na+显著高于泗棉 2号和苏棉 1 2号。Na+在茎和叶柄中滞留和积累 ,根中的 K+向地上部选择性运输 ,以维持叶片中较高的 K+/Na+,是棉花耐盐性的一个重要特点     

Improved measurements of Na+ fluxes in plants using calixarene-based microelectrodes

Ion-selective microelectrodes are a powerful tool in studying adaptive responses of plant cells and tissues to various abiotic stresses. However, application of this technique in Na⁺ flux measurements was limited due to poor selectivity for Na⁺ ions of commercially available Na⁺ cocktails. Often, these cocktails cannot discriminate between Na⁺ and other interfering ions such as K⁺ and Ca²⁺, leading to inaccurate measurements of Na⁺ concentration and, consequently, inaccurate Na⁺ flux calculations. To overcome this problem, three Na⁺-selective cocktail mixtures were prepared using tetramethoxyethyl ester derivative of p-t-butyl calix[4]arene. These cocktail mixtures were compared with commercially available ETH 227-based Na⁺ cocktail for selectivity for Na⁺ ions over other ions (particularly K⁺ and Ca²⁺). Among the three calixarene-based Na⁺ cocktails tested, cocktail 2 [in % w/w: Na⁺ ionophore (4-tert-butylcalix[4]arene-tetra acetic acid tetraethyl ester) 3.5, the plasticizer (2-nitrophenyl octyl ether) 95.9 and lipophilic anion (potassium tetrakis (4-chlorophenyl) borate) 0.6] showed the best selectivity for Na⁺ ions over K⁺ and Ca²⁺ ions and was highly stable over time (up to 10 h). Na⁺ flux measurements under a wide range of NaCl concentrations (25-150 mM) using Na⁺ cocktail 2 established a clear dose-response relationship between severity of salt stress and magnitude of Na⁺ influx at the distal elongation and mature zones of Arabidopsis thaliana roots. Furthermore, Na⁺ cocktail 2 was compared with commercially available ETH 227-based Na⁺ cocktail by measuring Na⁺ fluxes at the two Arabidopsis root zones in response to 100 mM NaCl treatment. With calixarene-based Na⁺ cocktail 2, a large decreasing Na⁺ influx (0-15 min) followed by small Na⁺ influx (15-45 min) was measured, whereas with ETH-based Na⁺ cocktail Na⁺ influx was short-lived (1-3 min) and was followed by Na⁺ efflux (3-45 min) that might have been due to K⁺ and Ca²⁺ efflux measured together with Na⁺ influx. In conclusion, Na⁺-selective calixarene-based microelectrodes have excellent potential to be used in real-time Na⁺ flux measurements in plants.     

Spermine accumulation under salt stress

Polyamines have long been recognized to be linked to stress situations, and it is generally accepted that they have protective characteristics. However, little is known about their physiological relevance in plants subjected to long-term salt stress. In order to precise their importance, two rice (Oryza sativa) cultivars differing in their salt tolerance were salinized for 7, 14 and 21 days. The activities of some of the enzymes involved in polyamine metabolism, free polyamines and proline contents were evaluated. Arginine decarboxylase and S-adenosyl-L-methionine decarboxylase activities were reduced in both cultivars as a consequence of salt treatment. However, spermidine synthase activity was reduced in the salt tolerant cultivar (var Giza) but not in the salt sensitive (var El Paso), while no polyamine oxidase activity was detected. During the salinization period, putrescine and spermidine levels decreased in both cultivars, although less dramatically in Giza. Simultaneously, spermine accumulations occur in both varieties, while proline accumulation was major in the sensitive one. However, spermine accumulation induced by treatment with spermidine synthase inhibitor cyclohexylamine, determined no reduction in leaf injury associated with salt stress in both cultivars. The data presented suggest that spermine accumulation is not a salt tolerance trait.     

Antioxidative defense under salt stress

Salt tolerance is a complex trait involving the coordinated action of many gene families that perform a variety of functions such as control of water loss through stomata, ion sequestration, metabolic adjustment, osmotic adjustment and antioxidative defense. In spite of the large number of publications on the role of antioxidative defense under salt stress, the relative importance of this process to overall plant salt tolerance is still a matter of controversy. In this article, the generation and scavenging of reactive oxygen species (ROS) under normal and salt stress conditions in relation to the type of photosynthesis is discussed. The CO₂ concentrating mechanism in C4 and CAM plants is expected to contribute to decreasing ROS generation. However, the available data supports this hypothesis in CAM but not in C4 plants. We discuss the specific roles of enzymatic and non enzymatic antioxidants in relation to the oxidative load in the context of whole plant salt tolerance. The possible preventive antioxidative mechanisms are also discussed.Key words: salt stress, generation of ROS, type of photosynthesis, scavenging of ROS, preventive antioxidative defense     

Ca2+-activated Na+ fluxes in human red cells. Amiloride sensitivity

The effect of Ca2+ on the ouabain- and bumetanide-resistant Na+ fluxes in intact red cells was studied at relatively constant internal Ca2+, membrane potential, and cell volume. The red cell calcium concentration was modified using the ionophore A23187. In fresh red cells, the Na+ influx and efflux (1.2 +/- 0.13 and 0.26 +/- 0.07 mmol/liter cells x h, respectively) were not affected by amiloride (1 mM). When external Ca2+ was raised from 0 to 150 microM, in the presence of A23187, both the Na+ influx and efflux were stimulated (about 3.5-fold). The Ca2+-activated Na+ efflux and influx had an apparent Km for activation by Ca2+o of about 25 microM. The Ca2+-dependent Na+ transport was inhibited 30-60% by amiloride (ID50 = 17.3 +/- 8 microM). Amiloride, however, had no effect on the Ca2+-dependent K+ influx. The amiloride-sensitive (AS) transport pathway was a linear function of the Na+o concentration in the range from 0 to 75 mM. The Ca2+i activation seems to depend on the metabolic integrity of red cells. 1) It does not take place in ATP-depleted red cells; 2) ATP-repletion of ATP-depleted red cells fully restored AS Na influx; and 3) ATP-enrichment (ATP-red cells) enhanced the AS Na influx by about 100%. The Ca2+-activated AS Na+ influx was not affected by either DIDS or trifluoperazine. The present results indicate that in human erythrocytes an increase in internal Ca2+ activates on otherwise silent AS Na+-transport system, which is dependent on the metabolic integrity of the red cells.