Protective effect of exogenous polyamines on root tonoplast function against salt stress in barley seedlings

Salinity stress is one of the most serious factors limiting the productivity of agricultural crops. A possible survival strategy of plants under saline conditions is to sequester excess Na⁺ in the vacuole by vacuolar Na⁺/H⁺ antiport using a pH gradient generated by H⁺-ATPasc (EC 3.6.1.35) and H⁺-Pyrophosphatase (H⁺-PPase; EC 3.6.1.1) to maintain a higher K⁺/Na⁺ ratio in cytoplasm. The effect of exogenously applied polyamines (PAs) in stabilizing root tonoplast integrity and function against salt stress in the barley (Hordeum vulgare L.) seedlings was investigated. The NaCl-induced reductions in the contents of phospholipids and PAs in tonoplast vesicles isolated from barely seedling roots, as well as the activities of H⁺-ATPase, H⁺-PPase and vacuolar Na⁺/H⁺ antiport were all partially restored by the application of 0.5 mM putrescine and 0.5 mM spermidine, especially the former. The above results indicated that one of the mechanisms involved in attenuating salt injury in barley seedlings by exogenous PAs application was to maintain tonoplast integrity and function under saline conditions. Moreover, the possible mechanism involved in counteracting detrimental effects of salt on the barley seedlings by the application of exogenous PAs was discussed.     

Nitric oxide enhances salt tolerance in maize seedlings through increasing activities of proton-pump and Na+/H+ antiport in the tonoplast

Nitric oxide (NO), an endogenous signaling molecule in animals and plants, mediates responses to abiotic and biotic stresses. Our previous work demonstrated that 100 μM sodium nitroprusside (SNP, an NO donor) treatment of maize seedlings increased K⁺ accumulation in roots, leaves and sheathes, while decreasing Na⁺ accumulation (Zhang et al. in J Plant Physiol Mol Biol 30:455–459, 2004b). Here we investigate how NO regulates Na⁺, K⁺ ion homeostasis in maize. Pre-treatment with 100 μM SNP for 2 days improved later growth of maize plants under 100 mM NaCl stress, as indicated by increased dry matter accumulation, increased chlorophyll content, and decreased membrane leakage from leaf cells. An NO scavenger, methylene blue (MB-1), blocked the effect of SNP. These results indicated that SNP-derived NO enhanced maize tolerance to salt stress. Further analysis showed that NaCl induced a transient increase in the NO level in maize leaves. Both NO and NaCl treatment stimulated vacuolar H⁺-ATPase and H⁺-PPase activities, resulting in increased H⁺-translocation and Na⁺/H⁺ exchange. NaCl-induced H⁺-ATPase and H⁺-PPase activities were diminished by MB-1. 1-Butanol, an inhibitor of phosphatidic acid (PA) production by phospholipase D (PLD), reduced NaCl- and NO-induced H⁺-ATPase activation. In contrast, applied PA stimulated H⁺-ATPase activity. These results suggest that NO acts as a signal molecule in the NaCl response by increasing the activities of vacuolar H⁺-ATPase and H⁺-PPase, which provide the driving force for Na⁺/H⁺ exchange. PLD and PA play an important role in this process.     

Protective effects of exogenous fatty acids on root tonoplast function against salt stress in barley seedlings

Salinity stress is one of the most serous factors limiting the productivity of agricultural crops. Previous studies have shown that exogenous fatty acids (EFAs) enhanced plant performance in saline environment. However, the mechanisms remained unclear. This study aimed to investigate whether EFAs (palmitic and linoleic acids) had ameliorating effects on salt injury in NaCl-treated barley (Hordeum vulgare L.) seedlings, and to explore the possible mechanisms by determining tonoplast composition and function. The results showed that linoleic acid at 1 mmol l⁻¹ in culture solution possessed protective effects on root tonoplast function against salt stress in the barley seedlings; this was accompanied with a significant suppression of the degradation of phospholipids and PAs in tonoplast vesicles. Moreover, these salt-ameliorating effects of linoleic acid on tonoplast function were also indicated by the increase in H⁺-ATPase and H⁺-PPase activities. In response to the changes in membrane bound enzyme activities, an augmentation in the activity of a vacuolar Na⁺/H⁺ antiport was occurred by the application of linoleic acid under saline conditions. These findings suggested that the application of linoleic acid exhibited protective effects on tonoplast function in the barley seedlings under salt stress, perhaps due partly to suppress the degradation of phospholipids and PAs in tonoplast vesicles, thus leading partial restorations in the activities of vacuolar H⁺-ATPase, H⁺-PPase and Na⁺/H⁺ antiport.     

Hydrogen peroxide and nitric oxide mediate K+/Na+ homeostasis and antioxidant defense in NaCl-stressed callus cells of two contrasting poplars

Using callus cells of a salt-tolerant Populus euphratica Oliver and a salt-sensitive P. popularis 35–44 (P. popularis), the effects of NaCl stress on hydrogen peroxide (H₂O₂) and nitric oxide (NO) production and the relevance to ionic homeostasis and antioxidant defense were investigated. Results show that P. euphratica exhibited a greater capacity to tolerate NaCl stress in terms of cell viability, membrane permeability and K⁺/Na⁺ relations. NaCl salinity (150 mM) caused a rapid increase of H₂O₂ and NO in P. euphratica cells, but not in P. popularis. Moreover, salinised P. euphratica cells retained a high and stable level of H₂O₂ and NO during the period of 24-h salt stress. Noteworthy, P. eupratica cells increased activities of superoxide dismutase, ascorbate peroxidase, catalase and glutathione reductase under salinity stress, but these antioxidant enzymes were significantly inhibited by the salt treatment in P. popularis cells. Pharmacological experiments proved that the NaCl-induced H₂O₂ and NO was interdependent and contributed to the mediation of K⁺/Na⁺ homeostasis and antioxidant defense in P. euphratica cells. Given these results, we conclude that the increased H₂O₂ and NO enable P. euphratica cells to regulate ionic and ROS (reactive oxygen species) homeostasis under salinity stress in the longer term.     

Polyamines, polyamine oxidases and nitric oxide in development, abiotic and biotic stresses

Nitric oxide (NO), polyamines (PAs), diamine oxidases (DAO) and polyamine oxidases (PAO) play important roles in wide spectrum of physiological processes such as germination, root development, flowering and senescence and in defence responses against abiotic and biotic stress conditions. This functional overlapping suggests interaction of NO and PA in signalling cascades. Exogenous application of PAs putrescine, spermidine and spermine to Arabidopsis seedlings induced NO production as observed by fluorimetry and fluorescence microscopy using the NO-binding fluorophores DAF-2 and DAR-4M. The observed NO release induced by 1 mM spermine treatment in the Arabidopsis seedlings was very rapid without apparent lag phase. These observations pave a new insight into PA-mediated signalling and NO as a potential mediator of PA actions. When comparing the functions of NO and PA in plant development and abiotic and biotic stresses common to both signalling components it can be speculated that NO may be a link between PA-mediated stress responses filing a gap between many known physiological effects of PAs and amelioration of stresses. NO production indicated by PAs could be mediated either by H₂O₂, one reaction product of oxidation of PAs by DAO and PAO, or by unknown mechanisms involving PAs, DAO and PAO.     

Herbivore exposure alters ion fluxes and improves salt tolerance in a desert shrub

Plants have evolved complex mechanisms that allow them to withstand multiple environmental stresses, including biotic and abiotic stresses. Here, we investigated the interaction between herbivore exposure and salt stress of Ammopiptanthus nanus, a desert shrub. We found that jasmonic acid (JA) was involved in plant responses to both herbivore attack and salt stress, leading to an increased NaCl stress tolerance for herbivore-pretreated plants and increase in K⁺/Na⁺ ratio in roots. Further evidence revealed the mechanism by which herbivore improved plant NaCl tolerance. Herbivore pretreatment reduced K⁺ efflux and increased Na⁺ efflux in plants subjected to long-term, short-term, or transient NaCl stress. Moreover, herbivore pretreatment promoted H⁺ efflux by increasing plasma membrane H⁺-adenosine triphosphate (ATP)ase activity. This H⁺ efflux creates a transmembrane proton motive force that drives the Na⁺/H⁺ antiporter to expel excess Na⁺ into the external medium. In addition, high cytosolic Ca²⁺ was observed in the roots of herbivore-treated plants exposed to NaCl, and this effect may be regulated by H⁺-ATPase. Taken together, herbivore exposure enhance s A. nanus tolerance to salt stress by activating the JA-signalling pathway, increasing plasma membrane H ⁺ - ATPase activity, promoting cytosolic Ca²⁺ accumulation, and then restricting K⁺ leakage and reducing Na⁺ accumulation in the cytosol.     

Ethylene and nitric oxide are involved in maintaining ion homeostasis in <Emphasis Type="Italic">Arabidopsis</Emphasis> callus under salt stress

In the present study, the role of ethylene in nitric oxide (NO)-mediated protection by modulating ion homeostasis in Arabidopsis callus under salt stress was investigated. Results showed that the ethylene-insensitive mutant etr1-3 was more sensitive to salt stress than the wild type (WT). Under 100 mM NaCl, etr1-3 callus displayed a greater electrolyte leakage and Na⁺/K⁺ ratio but a lower plasma membrane (PM) H⁺-ATPase activity compared to WT callus. Application of exogenous 1-aminocyclopropane-1-carboxylic acid (ACC, an ethylene precursor) or sodium nitroprusside (SNP, a NO donor) alleviated NaCl-induced injury by maintaining a lower Na⁺/K⁺ ratio and an increased PM H⁺-ATPase activity in WT callus but not in etr1-3 callus. The SNP actions in NaCl stress were attenuated by a specific NO scavenger or an ethylene biosynthesis inhibitor in WT callus. Under 100 mM NaCl, the NO accumulation and ethylene emission appeared at early time, and NO production greatly stimulated ethylene emission in WT callus. In addition, ethylene induced the expression of PM H⁺-ATPase genes under salt stress. The recovery experiment showed that NaCl-induced injury was reversible, as signaled by the similar recovery of Na⁺/K⁺ ratio and PM H⁺-ATPase activity in WT callus. Taken together, the results indicate that ethylene and NO cooperate in stimulating PM H⁺-ATPase activity to modulate ion homeostasis for salt tolerance, and ethylene may be a part of the downstream signal molecular in NO action.     

Leaf cell membrane stability‐based mechanisms of zinc nutrition in mitigating salinity stress in rice

• Excess salt affects about 955 million ha of arable land worldwide, and 49% of agricultural land is Zn‐deficient. Soil salinity and zinc deficiency can intensify plant abiotic stress. The mechanisms by which Zn can mitigate salinity effects on plant functions are not well understood.
• We conducted an experiment to determine how Zn and salinity effects on rice plant retention of Zn, K⁺ and the salt ion Na⁺ affect chlorophyll formation, leaf cell membrane stability and grain yield. We examined the mechanisms of Zn nutrition in mitigating salinity stress by examining plant physiology and nutrition. We used native Zn‐deficient soils (control), four salinity (EC ) and Zn treatments – Zn 10 mg·kg^?1 (Zn₁₀), EC 5 dS ·m^?1 (EC ₅), Zn₁₀+EC ₅ and Zn₁₅+EC ₅, a coarse rice (KS ‐282) and a fine rice (Basmati‐515) in the study.
• Our results showed that Zn alone (Zn₁₀) significantly increased rice tolerance to salinity stress by promoting Zn/K⁺ retention, inhibiting plant Na⁺ uptake and enhancing leaf cell membrane stability and chlorophyll formation in both rice cultivars in native alkaline, Zn‐deficient soils (P  <  0.05). Further, under the salinity treatment (EC ₅), Zn inputs (10–15 mg·kg^?1) could also significantly promote rice plant Zn/K⁺ retention and reduce plant Na⁺ uptake, and thus increased leaf cell membrane stability and grain yield. Coarse rice was more salinity‐tolerant than fine rice, having significantly higher Zn/K⁺ nutrient retention.
• The mechanistic basis of Zn nutrition in mitigating salinity impacts was through promoting plant Zn/K⁺ uptake and inhibiting plant Na⁺ uptake, which could result in increased plant physiological vigour, leaf cell membrane stability and rice productivity.

     

外源SA和NO对NaCl胁迫下番茄幼苗生长、光合及离子分布的影响

以番茄(Lycopersicon esculentum Mill.)品种‘秦丰保冠’为试材,采用营养液培养法,研究单独和复配施用外源水杨酸(SA)、一氧化氮(NO)供体硝普钠(SNP)对100mmol/L NaCl胁迫下番茄幼苗生长、光合及离子分布的影响。结果表明:(1)单独和复配外施SA、SNP均能有效抑制NaCl胁迫下番茄幼苗叶片光合色素(Chla、Chlb、Chla+b和Car)含量、Chla/b值、净光合速率(Pn)、蒸腾速率(Tr)、气孔导度(Gs)、瞬时水分利用效率(WUEt)、表观光能利用效率(LUEapp)和表观CO2利用效率(CUEapp)的下降及Car/Chla+b值和胞间CO2浓度(Ci)的升高,并以SA和SNP复配处理效果最明显。(2)NaCl胁迫下,外源SA、SNP单独和复配处理的番茄幼苗各器官(叶、茎和根)中Cl-、Na+含量和Na+/K+、Na+/Ca2+、Na+/Mg2+值显著降低,而K+、Ca2+和Mg2+的含量却不同程度提高,其中以SA和SNP复配处理效果最好。(3)单独和复配外施SA、SNP均能有效减轻NaCl胁迫对番茄幼苗生长的抑制作用,并促进各器官生物量的积累和壮苗的形成,且以SA和SNP复配处理效果更佳。研究表明,复配外施SA和SNP在诱导番茄幼苗提高抗(耐)盐能力方面具有协同增效作用。     

Exogenous application of free polyamines enhance salt tolerance of pistachio (Pistacia vera L.) seedlings

The protective effects of free polyamines (PAs) against salinity stress were investigated for pistachio seedlings (Pistacia vera cv. Badami-Zarand) in a controlled greenhouse. Seedlings were treated with 25, 50, 100 and 150 mM of salts including NaCl, CaCl₂ and MgCl₂. Foliar treatments of putrescine, spermidine (Spd) and spermine (Spm) (0.1 and 1 mM) were applied during the salinity period. Results showed that growth characteristics of pistachio seedlings decreased under salinity stress and the application of PAs efficiently reduced the adverse effects of salt stress. PAs reduced the severe effects of salt stress in pistachio seedlings neither by increasing the activities of peroxidase and ascorbate peroxidase nor by increasing the proline content but by increasing the activities of superoxide dismutase and catalase and decreasing the hydrogen peroxide (H₂O₂) activity. PAs treated seedlings showed a lower Na⁺:K⁺ ratio and Cl^? in leaves suggesting the role of PAs in balancing the ion exchange and better Na⁺:K⁺ discrimination under salt stress condition. These results showed the promising potential use of PAs especially Spm and Spd for reducing the negative effects of salinity stress and improving the growth of pistachio seedlings.     

Heterologous expression of Arabidopsis H+‐pyrophosphatase enhances salt tolerance in transgenic creeping bentgrass (Agrostis stolonifera L.)

The Arabidopsis vacuolar H⁺‐pyrophosphatase (AVP1), when over‐expressed in transgenic (TG) plants, regulates root and shoot development via facilitation of auxin flux, and enhances plant resistance to salt and drought stresses. Here, we report that TG perennial creeping bentgrass plants over‐expressing AVP1 exhibited improved resistance to salinity than wild‐type (WT) controls. Compared to WT plants, TGs grew well in the presence of 100 mm NaCl, and exhibited higher tolerance and faster recovery from damages from exposure to 200 and 300 mm NaCl. The improved performance of the TG plants was associated with higher relative water content (RWC), higher Na⁺ uptake and lower solute leakage in leaf tissues, and with higher concentrations of Na⁺, K⁺, Cl^‐ and total phosphorus in root tissues. Under salt stress, proline content was increased in both WT and TG plants, but more significantly in TGs. Moreover, TG plants exhibited greater biomass production than WT controls under both normal and elevated salinity conditions. When subjected to salt stress, fresh (FW) and dry weights (DW) of both leaves and roots decreased more significantly in WT than in TG plants. Our results demonstrated the great potential of genetic manipulation of vacuolar H⁺‐pyrophosphatase expression in TG perennial species for improvement of plant abiotic stress resistance.     

Reactive Oxygen Species Production in Wheat Roots Is Not Linked with Changes in H+ Fluxes During Acidic and Aluminium Stresses

Aluminium stress induces peroxidation of lipids in the plasma membrane, the effect akin to that caused by reactive oxygen species (ROS). ROS have recently been proposed as regulators of redox-dependent ion transport across the plasma membrane during biotic and abiotic stresses, thus contributing to the plant defence system. The aim of this study was to discover whether ROS production is linked to redox-dependent H⁺ transport system located at the plasma membranes of two near-isogenic lines of wheat (Triticum aestivum L., ET8 = Al-resistant, ES8 = Al-sensitive).The activities of NADPH-dependent ROS synthase and SOD were increased in both wheat lines 15 and 30 min after Al treatments. However, the ROS production was also increased under acidic stress. There was no difference between the two wheat lines in the root-cell plasma membrane capacity to efflux H⁺ in response to potassium ferricyanide after Al and acidic treatments. In ET8, both stresses led to increases in ROS production and H⁺ influx.ROS production in wheat seedlings was activated primarily by low pH exposure rather than by the Al stress. ROS production and breakdown in wheat seedlings under Al and acidic stresses appear to be linked to the intracellular metabolic changes rather than to the increased activity of plasma membrane-based NADPH-dependent ROS synthase.Key Words: ion fluxes, reactive oxygen species (ROS), redox system, superoxide dismutase (SOD), Triticum aestivum L., wheat     

In vivo role of Arabidopsis arginase in arginine metabolism and abiotic stress response

Nitric oxide (NO) and polyamines play essential roles in many developmental processes and abiotic stress responses in plants. NO and polyamines are metabolized from arginine through NO synthase (NOS) and arginine decarboxylase (ADC), respectively. Function of arginase, another important enzyme involved in arginine metabolism, in abiotic stress remains largely unknown. In the recent study, we have dissected the impact of arginase on arginine metabolism and abiotic stress responses through manipulating AtARGAHs expression. The results suggested that manipulation of arginase expression modulated accumulation of arginine and direct downstream products of arginine catabolism. AtARGAHs knockout lines exhibited increased accumulation of polyamines and NO and enhanced abiotic stress tolerance, while AtARGAHs overexpressing lines displayed the opposite results. Notably, we highlighted that Arabidopsis arginase plays distinctive and dual roles in the crosstalk between polyamines and NO signaling during abiotic stress responses, mediating both arginine metabolism and reactive oxygen species (ROS) accumulation. It is likely that accumulation of both NO and polyamines might activate abiotic stress responses in the plant.     

Nitric Oxide Mediates Root K+/Na+ Balance in a Mangrove Plant,Kandelia obovata,by Enhancing the Expression of AKT1-Type K+ Channel and Na+/H+ Antiporter under High Salinity

It is well known that nitric oxide (NO) enhances salt tolerance of glycophytes. However, the effect of NO on modulating ionic balance in halophytes is not very clear. This study focuses on the role of NO in mediating K⁺/Na⁺ balance in a mangrove species, Kandelia obovata Sheue, Liu and Yong. We first analyzed the effects of sodium nitroprusside (SNP), an NO donor, on ion content and ion flux in the roots of K. obovata under high salinity. The results showed that 100 μM SNP significantly increased K⁺ content and Na⁺ efflux, but decreased Na⁺ content and K⁺ efflux. These effects of NO were reversed by specific NO synthesis inhibitor and scavenger, which confirmed the role of NO in retaining K⁺ and reducing Na⁺ in K. obovata roots. Using western-blot analysis, we found that NO increased the protein expression of plasma membrane (PM) H⁺-ATPase and vacuolar Na⁺/H⁺ antiporter, which were crucial proteins for ionic balance. To further clarify the molecular mechanism of NO-modulated K⁺/Na⁺ balance, partial cDNA fragments of inward-rectifying K⁺ channel, PM Na⁺/H⁺ antiporter, PM H⁺-ATPase, vacuolar Na⁺/H⁺ antiporter and vacuolar H⁺-ATPase subunit c were isolated. Results of quantitative real-time PCR showed that NO increased the relative expression levels of these genes, while this increase was blocked by NO synthesis inhibitors and scavenger. Above results indicate that NO greatly contribute to K⁺/Na⁺ balance in high salinity-treated K. obovata roots, by activating AKT1-type K⁺ channel and Na⁺/H⁺ antiporter, which are the critical components in K⁺/Na⁺ transport system.     

New perspective on the mechanism of alleviating salt stress by spermidine in barley seedlings

Salt stress is considered to be a major limiting factor for plant growth and crop productivity. Salt injuries in plants are mostly due to excess Na⁺ entry. A possible survival strategy of plants under saline environments is the effective compartmentation of excess Na⁺ by sequestering Na⁺ in roots and inhibiting transport of Na⁺ from roots to shoots. Our previous study showed that exogenous application of polyamines (PAs) could attenuate salt injuries in barley plants. In order to further understand such protective roles of PAs against salt stress, the effects of spermidine (Spd) on sodium and potassium distribution in barley (Hordeum vulgare L.) seedlings under saline conditions were investigated. The results showed that exogenous application of Spd induced reductions in Na⁺ levels in roots and shoots with comparison of NaCl-treated plants, while no significant changes in K⁺ levels were observed. Correspondingly, the plants treated with Spd exogenously maintained high values of [K⁺]/[Na⁺] as compared with salt-stressed plants. Moreover, it was shown by X-ray microanalysis that K⁺ and Na⁺ accumulated mainly in the exodermal intercellular space and cortical cells of roots under salinity stress, and low accumulation was observed in endodermal cells and stelar parenchyma, indicating Casparian bands possibly act as ion transport barriers. Most importantly, Spd treatment further strengthened this barrier effects, leading to inhibition of Na⁺ transport into shoots. These results suggest that, by reinforcing barrier effects of Casparian bands, exogenous Spd inhibits Na⁺ transport from roots to shoots under conditions of high salinity which are beneficial for attenuating salt injuries in barley seedlings.     

The molecular mechanism of plasma membrane H+-ATPases in plant responses to abiotic stress

Plasma membrane H⁺-ATPases (PM H⁺-ATPases) are critical proton pumps that export protons from the cytoplasm to the apoplast. The resulting proton gradient and difference in electrical potential energize various secondary active transport events. PM H⁺-ATPases play essential roles in plant growth, development, and stress responses. In this review, we focus on recent studies of the mechanism of PM H⁺-ATPases in response to abiotic stresses in plants, such as salt and high pH, temperature, drought, light, macronutrient deficiency, acidic soil and aluminum stress, as well as heavy metal toxicity. Moreover, we discuss remaining outstanding questions about how PM H⁺-ATPases contribute to abiotic stress responses.     

碱胁迫对黑麦草幼苗根系活性氧代谢和渗透溶质积累的影响

为了探讨牧草对碱胁迫的耐受程度,采用营养液砂培方法,研究了不同浓度NaHCO3（0、50、100、150和200 mmol·L^-1）胁迫对黑麦草幼苗根系生长、活性氧代谢和渗透溶质积累的影响。结果表明：NaHCO3胁迫显著抑制黑麦草幼苗根系的生长,其抑制程度随胁迫浓度提高而增强,黑麦草可耐受的最高NaHCO₃浓度约为150 mmol·L^-1。随着NaHCO₃胁迫浓度的增加,黑麦草根中超氧阴离子(O^-·₂)、过氧化氢(H₂O₂)和丙二醛(MDA)含量明显上升,超氧化物歧化酶(SOD)活性和谷胱甘肽(GSH)含量显著下降,过氧化氢酶(CAT)、过氧化物酶(POD)和抗坏血酸过氧化物酶(APX)活性及抗坏血酸(ASA)含量先升后降。黑麦草根中Na⁺含量随NaHCO₃浓度增大而增加,K⁺含量和K⁺/Na⁺比降低,可溶性糖含量先升后降,脯氨酸含量则先降后升,游离氨基酸含量呈先升后降再升高变化。表明碱胁迫导致的活性氧代谢失调和Na⁺、K⁺失衡及积累有机溶质进行渗透调节时更多能量的消耗可能是黑麦草根系生长受抑的重要因素。