Tolerance of Hordeum marinum accessions to O2 deficiency, salinity and these stresses combined

Background and Aims

When root-zone O₂ deficiency occurs together with salinity, regulation of shoot ion concentrations is compromised even more than under salinity alone. Tolerance was evaluated amongst 34 accessions of Hordeum marinum, a wild species in the Triticeae, to combined salinity and root-zone O₂ deficiency. Interest in H. marinum arises from the potential to use it as a donor for abiotic stress tolerance into wheat.

Methods

Two batches of 17 H. marinum accessions, from (1) the Nordic Gene Bank and (2) the wheat belt of Western Australia, were exposed to 0·2 or 200 mol m⁻³ NaCl in aerated or stagnant nutrient solution for 28–29 d. Wheat (Triticum aestivum) was included as a sensitive check species. Growth, root porosity, root radial O₂ loss (ROL) and leaf ion (Na⁺, K⁺, Cl⁻) concentrations were determined.

Key Results

Owing to space constraints, this report is focused mainly on the accessions from the Nordic Gene Bank. The 17 accessions varied in tolerance; relative growth rate was reduced by 2–38 % in stagnant solution, by 8–42 % in saline solution (aerated) and by 39–71 % in stagnant plus saline treatment. When in stagnant solution, porosity of adventitious roots was 24–33 %; salinity decreased the root porosity in some accessions, but had no effect in others. Roots grown in stagnant solution formed a barrier to ROL, but variation existed amongst accessions in apparent barrier ‘strength’. Leaf Na⁺ concentration was 142–692 µmol g⁻¹ d. wt for plants in saline solution (aerated), and only increased to 247–748 µmol g⁻¹ d. wt in the stagnant plus saline treatment. Leaf Cl⁻ also showed only small effects of stagnant plus saline treatment, compared with saline alone. In comparison with H. marinum, wheat was more adversely affected by each stress alone, and particularly when combined; growth reductions were greater, adventitious root porosity was 21 %, it lacked a barrier to ROL, leaf K⁺ declined to lower levels, and leaf Na⁺ and Cl⁻ concentrations were 3·1–9-fold and 2·8–6-fold higher, respectively, in wheat.

Conclusions

Stagnant treatment plus salinity reduced growth more than salinity alone, or stagnant alone, but some accessions of H. marinum were still relatively tolerant of these combined stresses, maintaining Na⁺ and Cl⁻ ‘exclusion’ even in an O₂-deficient, saline rooting medium.Key words: Aerenchyma, combined salinity and waterlogging, leaf Cl⁻, leaf K⁺, leaf Na⁺, radial O₂ loss, salt tolerance, salinity–waterlogging interaction, sea barleygrass, waterlogging tolerance, wheat, wild Triticeae     

Tolerance of combined submergence and salinity in the halophytic stem-succulent Tecticornia pergranulata

Background and Aims

Habitats occupied by many halophytes are not only saline, but are also prone to flooding. Few studies have evaluated submergence tolerance in halophytes.

Methods

Responses to submergence, at a range of salinity levels, were studied for the halophytic stem-succulent Tecticornia pergranulata subsp. pergranulata (syn. Halosarcia pergranulata subsp. pergranulata). Growth and total sugars in succulent stems were assessed as a function of time after submergence. Underwater net photosynthesis, dark respiration, total sugars, glycinebetaine, Na⁺, Cl⁻ and K⁺, in succulent stems, were assessed in a NaCl dose-response experiment.

Key Results

Submerged plants ceased to grow, and tissue sugars declined. Photosynthesis by succulent stems was reduced markedly when underwater, as compared with in air. Capacity for underwater net photosynthesis (P_N) was not affected by 10–400 mm NaCl, but it was reduced by 30 % at 800 mm. Dark respiration, underwater, increased in succulent stems at 200–800 mm NaCl, as compared with those at 10 mm NaCl. On an ethanol-insoluble dry mass basis, K⁺ concentration in succulent stems of submerged plants was equal to that in drained controls, across all NaCl treatments. Na⁺ and Cl⁻ concentrations, however, were elevated in stems of submerged plants, but so was glycinebetaine. Submerged stems increased in succulence, so solutes would have been ‘diluted’ on a tissue-water basis.

Conclusions

Tecticornia pergranulata tolerates complete submergence, even in waters of high salinity. A ‘quiescence response’, i.e. no shoot growth, would conserve carbohydrates, but tissue sugars still declined with time. A low K⁺ : Na⁺ ratio, typical for tissues of succulent halophytes, was tolerated even during prolonged submergence, as evidenced by maintenance of underwater P_N at up to 400 mm NaCl. Underwater P_N provides O₂ and sugars, and thus should enhance survival of submerged plants.Key words: Flooding, halophyte, Halosarcia pergranulata, inundation, inland salt marsh, respiration, Salicornioideae, salt lake, submergence–salinity interaction, tissue solutes, underwater net photosynthesis     

Salt accumulation and depletion in the root-zone of the halophyte Atriplex nummularia Lindl.: influence of salinity,leaf area and plant water use

Background and aims

Salt is known to accumulate in the root-zone of Na⁺ excluding glycophytes under saline conditions. We examined the effect of soil salinity on Na⁺ and Cl^? depletion or accumulation in the root-zone of the halophyte (Atriplex nummularia Lindl).

Methods

A pot experiment was conducted in soil to examine Na⁺ and Cl^? concentrations adjacent to roots at four initial NaCl treatments (20, 50, 200 or 400 mM NaCl in the soil solution). Plant water use was manipulated by leaving plants with all leaves intact, removing approximately 50 % of leaves, or removing all leaves. Daily evapotranspiration was replaced by watering undrained pots to weight with deionised water. After 35-38 days, samples were taken of the bulk soil and of soil loosely- and closely-adhering to the roots.

Results

In plants with leaves intact grown with 200 and 400 mM NaCl, average Na⁺ and Cl^? concentrations in the closely adhering soil were about twice the concentrations of the bulk soil. Ion accumulation increased with final leaf area and with cumulative transpiration over the duration of the trial. By contrast, in plants grown with the lowest salinity treatment (20 mM NaCl), Na⁺ and Cl^? concentrations decreased in the closely adhering soil with increasing leaf area and increasing cumulative water use.

Conclusions

Our data show that Na⁺ and Cl^? are depleted from the root-zone of A. nummularia at low salinity but accumulate in the root-zone at moderate to high salinity, and that the ions are drawn towards the plant in the transpiration stream.     

Extracellular Chloride Regulates the Epithelial Sodium Channel

The extracellular domain of the epithelial sodium channel ENaC is exposed to a wide range of Cl⁻ concentrations in the kidney and in other epithelia. We tested whether Cl⁻ alters ENaC activity. In Xenopus oocytes expressing human ENaC, replacement of Cl⁻ with SO₄²⁻, H₂PO₄⁻, or SCN⁻ produced a large increase in ENaC current, indicating that extracellular Cl⁻ inhibits ENaC. Extracellular Cl⁻ also inhibited ENaC in Na⁺-transporting epithelia. The anion selectivity sequence was SCN⁻ < SO₄²⁻ < H₂PO₄⁻ < F⁻ < I⁻ < Cl⁻ < Br⁻. Crystallization of ASIC1a revealed a Cl⁻ binding site in the extracellular domain. We found that mutation of corresponding residues in ENaC (α_H418A and β_R388A) disrupted the response to Cl⁻, suggesting that Cl⁻ might regulate ENaC through an analogous binding site. Maneuvers that lock ENaC in an open state (a DEG mutation and trypsin) abolished ENaC regulation by Cl⁻. The response to Cl⁻ was also modulated by changes in extracellular pH; acidic pH increased and alkaline pH reduced ENaC inhibition by Cl⁻. Cl⁻ regulated ENaC activity in part through enhanced Na⁺ self-inhibition, a process by which extracellular Na⁺ inhibits ENaC. Together, the data indicate that extracellular Cl⁻ regulates ENaC activity, providing a potential mechanism by which changes in extracellular Cl⁻ might modulate epithelial Na⁺ absorption.The epithelial Na⁺ channel ENaC² is a heterotrimer of homologous α, β, and γ subunits (1, 2). ENaC functions as a pathway for Na⁺ absorption across epithelial cells in the kidney collecting duct, lung, distal colon, and sweat duct (reviewed in Refs. 3 and 4). Na⁺ transport is critical for the maintenance of Na⁺ homeostasis and for the control of the composition and quantity of the fluid on the apical membrane of these epithelia. ENaC mutations and defects in its regulation cause inherited forms of hypertension and hypotension (5) and may contribute to the pathogenesis of lung disease in cystic fibrosis (6).ENaC is a member of the DEG/ENaC family of ion channels. A common structural feature of these channels is a large extracellular domain that plays a critical role in channel gating. For example, in ASICs, the extracellular domain functions as a receptor for protons, which transiently activate the channel by titrating residues that form an acidic pocket (7). FaNaCh is a ligand-gated family member in Helix aspersa, activated by the peptide FMRFamide (8). In Caenorhabditis elegans MEC family members, the extracellular domain is thought to respond to mechanical signals (9).ENaC differs from other family members because it is constitutively active in the absence of a ligand/stimulus. However, a convergence of data indicate that ENaC gating is modulated by a variety of molecules that bind to or modify its extracellular domains, including proteases (10–12), Na⁺ (13–15), protons (16), and the divalent cations Zn²⁺ and Ni²⁺ (17, 18). These findings suggest that the ENaC extracellular domain might regulate epithelial Na⁺ transport by sensing and integrating diverse signals in the extracellular environment.In the current study, we tested the hypothesis that ENaC activity is regulated by changes in the extracellular Cl⁻ concentration. Several observations suggested that Cl⁻ might be a strong candidate to regulate the channel. First, transport of Na⁺ and Cl⁻ are often coupled to maintain electroneutrality. Second, ENaC is exposed to large changes in extracellular Cl⁻ concentration. For example, in the kidney collecting duct, the urine Cl⁻ concentration varies widely (19). As the predominant anion, its concentration parallels that of Na⁺ in most clinical states. However, under conditions of metabolic alkalosis and metabolic acidosis, the Na⁺ and Cl⁻ concentrations can become dissociated as a result of increased urinary bicarbonate (alkalosis) or ammonium (acidosis) (19). Thus, ENaC is well positioned to respond to changes in Cl⁻ concentration. Third, crystallization of ASIC1a revealed a binding site for a Cl⁻ ion at the base of the thumb domain (7). The Cl⁻ is coordinated by Arg-310 and Glu-314 from one subunit and Lys-212 from an adjacent subunit. Although the functional role of Cl⁻ binding to ASIC1a is unknown, it supports the hypothesis that extracellular Cl⁻ might regulate the activity of DEG/ENaC ion channels.     

The K+ channel opener 1-EBIO potentiates residual function of mutant CFTR in rectal biopsies from cystic fibrosis patients

Background

The identification of strategies to improve mutant CFTR function remains a key priority in the development of new treatments for cystic fibrosis (CF). Previous studies demonstrated that the K⁺ channel opener 1-ethyl-2-benzimidazolone (1-EBIO) potentiates CFTR-mediated Cl⁻ secretion in cultured cells and mouse colon. However, the effects of 1-EBIO on wild-type and mutant CFTR function in native human colonic tissues remain unknown.

Methods

We studied the effects of 1-EBIO on CFTR-mediated Cl⁻ secretion in rectal biopsies from 47 CF patients carrying a wide spectrum of CFTR mutations and 57 age-matched controls. Rectal tissues were mounted in perfused micro-Ussing chambers and the effects of 1-EBIO were compared in control tissues, CF tissues expressing residual CFTR function and CF tissues with no detectable Cl⁻ secretion.

Results

Studies in control tissues demonstrate that 1-EBIO activated CFTR-mediated Cl⁻ secretion in the absence of cAMP-mediated stimulation and potentiated cAMP-induced Cl⁻ secretion by 39.2±6.7% (P<0.001) via activation of basolateral Ca²⁺-activated and clotrimazole-sensitive KCNN4 K⁺ channels. In CF specimens, 1-EBIO potentiated cAMP-induced Cl⁻ secretion in tissues with residual CFTR function by 44.4±11.5% (P<0.001), but had no effect on tissues lacking CFTR-mediated Cl⁻conductance.

Conclusions

We conclude that 1-EBIO potentiates Cl⁻secretion in native CF tissues expressing CFTR mutants with residual Cl⁻ channel function by activation of basolateral KCNN4 K⁺ channels that increase the driving force for luminal Cl⁻ exit. This mechanism may augment effects of CFTR correctors and potentiators that increase the number and/or activity of mutant CFTR channels at the cell surface and suggests KCNN4 as a therapeutic target for CF.     

Response to non-uniform salinity in the root zone of the halophyte Atriplex nummularia: growth, photosynthesis, water relations and tissue ion concentrations

Background and Aims

Soil salinity is often heterogeneous, yet the physiology of halophytes has typically been studied with uniform salinity treatments. An evaluation was made of the growth, net photosynthesis, water use, water relations and tissue ions in the halophytic shrub Atriplex nummularia in response to non-uniform NaCl concentrations in a split-root system.

Methods

Atriplex nummularia was grown in a split-root system for 21 d, with either the same or two different NaCl concentrations (ranging from 10 to 670 mm), in aerated nutrient solution bathing each root half.

Key Results

Non-uniform salinity, with high NaCl in one root half (up to 670 mm) and 10 mm in the other half, had no effect on shoot ethanol-insoluble dry mass, net photosynthesis or shoot pre-dawn water potential. In contrast, a modest effect occurred for leaf osmotic potential (up to 30 % more solutes compared with uniform 10 mm NaCl treatment). With non-uniform NaCl concentrations (10/670 mm), 90 % of water was absorbed from the low salinity side, and the reduction in water use from the high salinity side caused whole-plant water use to decrease by about 30 %; there was no compensatory water uptake from the low salinity side. Leaf Na⁺ and Cl⁻ concentrations were 1·9- to 2·3-fold higher in the uniform 670 mm treatment than in the 10/670 mm treatment, whereas leaf K⁺ concentrations were 1·2- to 2·0-fold higher in the non-uniform treatment.

Conclusions

Atriplex nummularia with one root half in 10 mm NaCl maintained net photosynthesis, shoot growth and shoot water potential even when the other root half was exposed to 670 mm NaCl, a concentration that inhibits growth by 65 % when uniform in the root zone. Given the likelihood of non-uniform salinity in many field situations, this situation would presumably benefit halophyte growth and physiology in saline environments.Key words: Split-root system, salinity heterogeneity, root zone heterogeneity, water potential, water use, stomatal conductance, saltbush, leaf ions, photosynthesis, NaCl     

The uptake and flow of C, N and ions between roots and shoots in Ricinus communis L. III. Long-distance transport of abscisic acid depending on nitrogen nutrition and salt stress

Seedlings of Ricinus communis L. were cultivated in quartz sandand supplied with media which contained either different concentrationsof nitrate or ammonium nitrogen and were treated with a lowsalt stress. The concentration of ABA was determined in tissuesand in xylem and phloem saps. Between 41 and 51 day after sowing,abscisic acid (ABA) flows between roots and shoots were modelled.Long-distance transport of ABA was not stimulated under conditionsof nitrate deficiency (0.2 mol m^–3). However, when ammoniumwas given as the only N source (1.0 mol m^–3), ABA transportin both xylem and phloem was increased significantly. Mild saltstress (40 mol m^–3 NaCl) increased ABA transport in nitrate-fedplants, but not in ammonium-fed plants. The leaf conductancewas lowered by salt treatment with both nitrogen sources, butit was always lower in ammonium-fed compared to nitrate-fedplants. A negative correlation of leaf conductance to ABA levelsin leaves or flow in xylem was found only in comparison of ammonium-fedto nitrate-fed plants. Key words: Abscisic acid, ammonium, Ricinus communis, phloem, xylem, transport, nitrate, nitrogen nutrition     

Ammonium nutrition in the halophyte Spartina alterniflora under salt stress: evidence for a priming effect of ammonium?

Background and aims

The effects of salt stress on the salt marsh halophyte Spartina alterniflora have been well documented. However, plant responses to combined salinity and ammonium toxicity and the underlying mechanisms are relatively unknown. The aim of the present investigation was to study the effects of both salinity (0, 200 and 500 mM NaCl) and nitrogen form (NO₃ ^?, NH₄ ⁺ or NH₄NO₃) on S. alterniflora.

Methods

Plants were cultivated in sandy soil under greenhouse conditions for 3 months. At harvest, growth parameters were measured and leaf samples were analysed for oxidative stress parameters (malondialdehyde, MDA; electrolyte leakage, EL; and hydrogen peroxide, H₂O₂ concentration) and the activity of antioxidant enzymes (glutathione reductase, GR; superoxide dismutase, SOD; catalase, CAT; ascorbate peroxidase, APX and Guaiacol peroxidase, GPX).

Results

In the absence of NaCl, plant growth rate was the highest in the medium containing both nitrogen forms, and the lowest in the medium containing only nitrate. Irrespective of the nitrogen form, plant growth was generally higher at 200 mM NaCl than without salinity. Ammonium-fed plants showed better growth than nitrate-fed plants under high salinity. In the absence of salinity, ammonium-fed plants showed higher SOD, APX, GR, CAT, and GPX activities than nitrate-fed ones. The antioxidant enzymes exhibited higher activity in saline-treated plants. The considerable advantage of NH₄ ⁺ nutrition to S. alterniflora under saline conditions was associated with high antioxidant enzyme activities, together with low MDA content, EL, and H₂O₂ concentration.

Conclusion

These data clearly demonstrate that NH₄ ⁺ is more favourable for the growth of S. alterniflora under high salinity than NO₃ ^?. It is suggested that NH₄ ⁺ nutrition improves the plant’s capacity to limit oxidative damage by stimulating the activities of the major antioxidant enzymes.     

Cation and Chloride Partitioning through Xylem and Phloem within the Whole Plant of Ricinus communis L. under Conditions of Salt Stress

Uptake and partitioning through the xylem and phloem of K⁺,Na⁺, Mg²⁺ , Ca²⁺ and Cl^– were studied over a 9 d intervalduring late vegetative growth of castor bean (Ricinus communisL.) plants exposed to a mean salinity stress of 128 mol m^–3NaCl. Empirically based models of flow and utilization of eachion within the whole plant were constructed using informationon ion increments of plant parts, molar ratios of ions to carbonin phloem sap sampled from petioles and stem internodes andpreviously derived information on carbon flow between plantsparts in xylem and phloem in identical plant material. Salientfeatures of the plant budget for K⁺ were prominent depositionin leaves, high mobility of K⁺ in phloem, high rates of cyclingthrough leaves and downward translocation of K⁺ providing theroot with a large excess of K⁺ . Corresponding data for Na⁺showed marked retention in the root, lateral uptake from xylemby hypocotyl, stem internodes and petioles leading to low intakeby young leaf laminae and substantial cycling from older leavesback to the root. The partitioning of the anionic componentof NaCl salinity, Cl^–, contrasted to that of Na⁺ in thatit was not substantially retained in the root, but depositedmore or less uniformly in stem, petiole and leaf lamina tissues.The flow pattern for Mg²⁺ showed relatively even depositionthrough the plant but some preferential uptake by young leaves,generally lesser export than import by leaf laminae, and a returnflow of Mg²⁺ from shoot to root considerably less than the recordedincrement of the root. Ca²⁺ partitioning contrasted with thatof the other ions in showing extremely poor phloem mobility,leading to progressive preferential accumulation in leaf laminaeand negligible cycling of the element through leaves or root.Features of the response of Ricinus to salinity shown in thepresent study were discussed with data from similar modellingstudies on white lupin (Lupinus albus L.) and barley (Hordeumvulgare L.) Key words: Ricinus communis L, potassium, sodium, chloride, calcium, magnesium, phloem, xylem, transport, partitioning, salinity     

Saline-induced changes of epicuticular waxy layer on the Puccinellia tenuiflora and Oryza sativa leave surfaces

Background

The epicuticular waxy layer of plant leaves enhances the extreme environmental stress tolerance. However, the relationship between waxy layer and saline tolerance was not established well. The epicuticular waxy layer of rice (Oryza sativa L.) was studied under the NaHCO₃ stresses. In addition, strong saline tolerance Puccinellia tenuiflora was chosen for comparative studies.

Results

Scanning electron microscope (SEM) images showed that there were significant changes in waxy morphologies of the rice epicuticular surfaces, while no remarkable changes in those of P. tenuiflora epicuticular surfaces. The NaHCO₃-induced morphological changes of the rice epicuticular surfaces appeared as enlarged silica cells, swollen corns-shapes and leaked salt columns under high stress. Energy dispersive X-ray (EDX) spectroscopic profiles supported that the changes were caused by significant increment and localization of [Na⁺] and [Cl⁻] in the shoot. Atomic absorption spectra showed that [Na⁺]_shoot/[Na⁺]_root for P. tenuiflora maintained stable as the saline stress increased, but that for rice increased significantly.

Conclusion

In rice, NaHCO₃ stress induced localization and accumulation of [Na⁺] and [Cl⁻] appeared as the enlarged silica cells (MSC), the swollen corns (S-C), and the leaked columns (C), while no significant changes in P. tenuiflora.     

Anti-diarrheal mechanism of the traditional remedy Uzara via reduction of active chloride secretion

Background and Purpose

The root extract of the African Uzara plant is used in traditional medicine as anti-diarrheal drug. It is known to act via inhibition of intestinal motility, but malabsorptive or antisecretory mechanisms are unknown yet.

Experimental Approach

HT-29/B6 cells and human colonic biopsies were studied in Ussing experiments in vitro. Uzara was tested on basal as well as on forskolin- or cholera toxin-induced Cl⁻ secretion by measuring short-circuit current (I_SC) and tracer fluxes of ²²Na⁺ and ³⁶Cl⁻. Para- and transcellular resistances were determined by two-path impedance spectroscopy. Enzymatic activity of the Na⁺/K⁺-ATPase and intracellular cAMP levels (ELISA) were measured.

Key Results

In HT-29/B6 cells, Uzara inhibited forskolin- as well as cholera toxin-induced I_SC within 60 minutes indicating reduced active chloride secretion. Similar results were obtained in human colonic biopsies pre-stimulated with forskolin. In HT-29/B6, the effect of Uzara on the forskolin-induced I_SC was time- and dose-dependent. Analyses of the cellular mechanisms of this Uzara effect revealed inhibition of the Na⁺/K⁺-ATPase, a decrease in forskolin-induced cAMP production and a decrease in paracellular resistance. Tracer flux experiments indicate that the dominant effect is the inhibition of the Na⁺/K⁺-ATPase.

Conclusion and Implications

Uzara exerts anti-diarrheal effects via inhibition of active chloride secretion. This inhibition is mainly due to an inhibition of the Na⁺/K⁺-ATPase and to a lesser extent to a decrease in intracellular cAMP responses and paracellular resistance. The results imply that Uzara is suitable for treating acute secretory diarrhea.     

Biogenic volatile organic compound and respiratory CO2 emissions after 13C-labeling: online tracing of C translocation dynamics in poplar plants

Background

Globally plants are the primary sink of atmospheric CO₂, but are also the major contributor of a large spectrum of atmospheric reactive hydrocarbons such as terpenes (e.g. isoprene) and other biogenic volatile organic compounds (BVOC). The prediction of plant carbon (C) uptake and atmospheric oxidation capacity are crucial to define the trajectory and consequences of global environmental changes. To achieve this, the biosynthesis of BVOC and the dynamics of C allocation and translocation in both plants and ecosystems are important.

Methodology

We combined tunable diode laser absorption spectrometry (TDLAS) and proton transfer reaction mass spectrometry (PTR-MS) for studying isoprene biosynthesis and following C fluxes within grey poplar (Populus x canescens) saplings. This was achieved by feeding either ¹³CO₂ to leaves or ¹³C-glucose to shoots via xylem uptake. The translocation of ¹³CO₂ from the source to other plant parts could be traced by ¹³C-labeled isoprene and respiratory ¹³CO₂ emission.

Principal Finding

In intact plants, assimilated ¹³CO₂ was rapidly translocated via the phloem to the roots within 1 hour, with an average phloem transport velocity of 20.3±2.5 cm h⁻¹. ¹³C label was stored in the roots and partially reallocated to the plants'' apical part one day after labeling, particularly in the absence of photosynthesis. The daily C loss as BVOC ranged between 1.6% in mature leaves and 7.0% in young leaves. Non-isoprene BVOC accounted under light conditions for half of the BVOC C loss in young leaves and one-third in mature leaves. The C loss as isoprene originated mainly (76–78%) from recently fixed CO₂, to a minor extent from xylem-transported sugars (7–11%) and from photosynthetic intermediates with slower turnover rates (8–11%).

Conclusion

We quantified the plants'' C loss as respiratory CO₂ and BVOC emissions, allowing in tandem with metabolic analysis to deepen our understanding of ecosystem C flux.     

Ionic components of electric current at rat corneal wounds

Background

Endogenous electric fields and currents occur naturally at wounds and are a strong signal guiding cell migration into the wound to promote healing. Many cells involved in wound healing respond to small physiological electric fields in vitro. It has long been assumed that wound electric fields are produced by passive ion leakage from damaged tissue. Could these fields be actively maintained and regulated as an active wound response? What are the molecular, ionic and cellular mechanisms underlying the wound electric currents?

Methodology/Principal Findings

Using rat cornea wounds as a model, we measured the dynamic timecourses of individual ion fluxes with ion-selective probes. We also examined chloride channel expression before and after wounding. After wounding, Ca²⁺ efflux increased steadily whereas K⁺ showed an initial large efflux which rapidly decreased. Surprisingly, Na⁺ flux at wounds was inward. A most significant observation was a persistent large influx of Cl⁻, which had a time course similar to the net wound electric currents we have measured previously. Fixation of the tissues abolished ion fluxes. Pharmacological agents which stimulate ion transport significantly increased flux of Cl⁻, Na⁺ and K⁺. Injury to the cornea caused significant changes in distribution and expression of Cl⁻ channel CLC2.

Conclusions/Significance

These data suggest that the outward electric currents occurring naturally at corneal wounds are carried mainly by a large influx of chloride ions, and in part by effluxes of calcium and potassium ions. Ca²⁺ and Cl⁻ fluxes appear to be mainly actively regulated, while K⁺ flux appears to be largely due to leakage. The dynamic changes of electric currents and specific ion fluxes after wounding suggest that electrical signaling is an active response to injury and offers potential novel approaches to modulate wound healing, for example eye-drops targeting ion transport to aid in the challenging management of non-healing corneal ulcers.     

Sodium transport in basil

In order to diversify the production of plants with pharmacological interest, it is important to understand the mechanisms involved in their tolerance to environmental constraints, such as salinity. Basil (Ocimum basilicum), known for its therapeutic uses, has been claimed to be salt tolerant, but physiological aspects of this behavior remain unknown. Since salt tolerance is known to be associated with several characteristics concerning Na⁺ transport to leaves, we studied this function in hydroponically grown basil. We analyzed the response of 30-day-old seedlings to 25–50 mM NaCl applied for 15 days. Growth was poorly affected, indicating that these concentrations corresponded to the tolerated salinity range. Leaves accumulated Na⁺ at relatively high concentration, without dehydrating. Potassium concentration in leaf tissues was maintained close to control level, indicating that K⁺ was 15- to 25-fold preferred over Na⁺ for ion transport and deposition. Collection of xylem sap on detopped plants revealed that this preference was only 10-fold for ion introduction into root xylem sap. Short-term (24 h) changes in Na⁺ distribution between organs after stem (steam) girdling suggested that Na⁺ downward recirculation by phloem occurred. Although modest, this transport might have augmented K⁺ selectivity of ion deposition in leaves.     

Learning from halophytes: physiological basis and strategies to improve abiotic stress tolerance in crops

Background

Global annual losses in agricultural production from salt-affected land are in excess of US$12 billion and rising. At the same time, a significant amount of arable land is becoming lost to urban sprawl, forcing agricultural production into marginal areas. Consequently, there is a need for a major breakthrough in crop breeding for salinity tolerance. Given the limited range of genetic diversity in this trait within traditional crops, stress tolerance genes and mechanisms must be identified in extremophiles and then introduced into traditional crops.

Scope and Conclusions

This review argues that learning from halophytes may be a promising way of achieving this goal. The paper is focused around two central questions: what are the key physiological mechanisms conferring salinity tolerance in halophytes that can be introduced into non-halophyte crop species to improve their performance under saline conditions and what specific genes need to be targeted to achieve this goal? The specific traits that are discussed and advocated include: manipulation of trichome shape, size and density to enable their use for external Na⁺ sequestration; increasing the efficiency of internal Na⁺ sequestration in vacuoles by the orchestrated regulation of tonoplast NHX exchangers and slow and fast vacuolar channels, combined with greater cytosolic K⁺ retention; controlling stomata aperture and optimizing water use efficiency by reducing stomatal density; and efficient control of xylem ion loading, enabling rapid shoot osmotic adjustment while preventing prolonged Na⁺ transport to the shoot.     

High phenotypic plasticity of Suaeda maritima observed under hypoxic conditions in relation to its physiological basis

Background and Aims

Phenotypic plasticity, the potential of specific traits of a genotype to respond to different environmental conditions, is an important adaptive mechanism for minimizing potentially adverse effects of environmental fluctuations in space and time. Suaeda maritima shows morphologically different forms on high and low areas of the same salt marsh. Our aims were to examine whether these phenotypic differences occurred as a result of plastic responses to the environment. Soil redox state, indicative of oxygen supply, was examined as a factor causing the observed morphological and physiological differences.

Methods

Reciprocal transplantation of seedlings was carried out between high and low marsh sites on a salt marsh and in simulated tidal-flow tanks in a glasshouse. Plants from the same seed source were grown in aerated or hypoxic solution, and roots were assayed for lactate dehydrogenase (LDH) and alcohol dehydrogenase, and changes in their proteome.

Key Results

Transplanted (away) seedlings and those that remained in their home position developed the morphology characteristic of the home or away site. Shoot Na⁺, Cl⁻ and K⁺ concentrations were significantly different in plants in the high and low marsh sites, but with no significant difference between home and away plants at each site. High LDH activity in roots of plants grown in aeration and in hypoxia indicated pre-adaptation to fluctuating root aeration and could be a factor in the phenotypic plasticity and growth of S. maritima over the full tidal range of the salt marsh environment. Twenty-six proteins were upregulated under hypoxic conditions.

Conclusions

Plasticity of morphological traits for growth form at extremes of the soil oxygenation spectrum of the tidal salt marsh did not correlate with the lack of physiological plasticity in the constitutively high LDH found in the roots.     

Control of xylem Na+ loading and transport to the shoot in rice and barley as a determinant of differential salinity stress tolerance

Control of xylem Na⁺ loading has often been named as the essential component of salinity tolerance mechanism. However, it is less clear to what extent the difference in this trait may determine differential salinity tolerance between species. In this study, barley (Hordeum vulgare L. cv. CM72) and rice (Oryza sativa L. cv. Dongjin) plants were grown under two levels of salinity. Na⁺ and K⁺ concentrations in the xylem sap, and shoot and root tissues were measured at different time points after stress onset. Salt‐exposed rice plants prevented xylem Na⁺ loading for several days, but failed to control this process in the longer term, ultimately resulting in a massive Na⁺ shoot loading. Barley plants quickly increased xylem Na⁺ concentration and its delivery to the shoot (most likely for the purpose of osmotic adjustment) but were able to reduce this process later on, keeping most of accumulated Na⁺ in the root, thus maintaining non‐toxic shoot Na⁺ level. Rice plants increased shoot K⁺ concentration, while barley plants maintained higher root K⁺ concentration. Control of xylem Na⁺ loading is remarkably different between rice and barley; this difference may differentiate the extent of the salinity tolerance between species. This trait should be investigated in more detail to be used in the breeding programs aimed to improve salinity tolerance in crops.     

Salt sensitivity in chickpea is determined by sodium toxicity

Main conclusion

Salt sensitivity in chickpea is determined by Na⁺ toxicity, whereas relatively high leaf tissue concentrations of Cl^? were tolerated, and the osmotic component of 60-mM NaCl was not detrimental.Chickpea (Cicer arietinum L.) is sensitive to salinity. This study dissected the responses of chickpea to osmotic and ionic components (Na⁺ and/or Cl^?) of salt stress. Two genotypes with contrasting salt tolerances were exposed to osmotic treatments (?0.16 and ?0.29 MPa), Na⁺-salts, Cl^?-salts, or NaCl at 0, 30, or 60 mM for 42 days and growth, tissue ion concentrations and leaf gas-exchange were assessed. The osmotic treatments and Cl^?-salts did not affect growth, whereas Na⁺-salts and NaCl treatments equally impaired growth in either genotype. Shoot Na⁺ and Cl^? concentrations had markedly increased, whereas shoot K⁺ had declined in the NaCl treatments, but both genotypes had similar shoot concentrations of each of these individual ions after 14 and 28 days of treatments. Genesis836 achieved higher net photosynthetic rate (64–84 % of control) compared with Rupali (35–56 % of control) at equivalent leaf Na⁺ concentrations. We conclude that (1) salt sensitivity in chickpea is determined by Na⁺ toxicity, and (2) the two contrasting genotypes appear to differ in ‘tissue tolerance’ of high Na⁺. This study provides a basis for focus on Na⁺ tolerance traits for future varietal improvement programs for salinity tolerance in chickpea.

     

Induced terpene accumulation in Norway spruce inhibits bark beetle colonization in a dose-dependent manner

Background

Tree-killing bark beetles (Coleoptera, Scolytinae) are among the most economically and ecologically important forest pests in the northern hemisphere. Induction of terpenoid-based oleoresin has long been considered important in conifer defense against bark beetles, but it has been difficult to demonstrate a direct correlation between terpene levels and resistance to bark beetle colonization.

Methods

To test for inhibitory effects of induced terpenes on colonization by the spruce bark beetle Ips typographus (L.) we inoculated 20 mature Norway spruce Picea abies (L.) Karsten trees with a virulent fungus associated with the beetle, Ceratocystis polonica (Siem.) C. Moreau, and investigated induced terpene levels and beetle colonization in the bark.

Results

Fungal inoculation induced very strong and highly variable terpene accumulation 35 days after inoculation. Trees with high induced terpene levels (n = 7) had only 4.9% as many beetle attacks (5.1 vs. 103.5 attacks m⁻²) and 2.6% as much gallery length (0.029 m m⁻² vs. 1.11 m m⁻²) as trees with low terpene levels (n = 6). There was a highly significant rank correlation between terpene levels at day 35 and beetle colonization in individual trees. The relationship between induced terpene levels and beetle colonization was not linear but thresholded: above a low threshold concentration of ∼100 mg terpene g⁻¹ dry phloem trees suffered only moderate beetle colonization, and above a high threshold of ∼200 mg terpene g⁻¹ dry phloem trees were virtually unattacked.

Conclusion/Significance

This is the first study demonstrating a dose-dependent relationship between induced terpenes and tree resistance to bark beetle colonization under field conditions, indicating that terpene induction may be instrumental in tree resistance. This knowledge could be useful for developing management strategies that decrease the impact of tree-killing bark beetles.     

Ecophysiological response of Crambe maritima to airborne and soil-borne salinity

Background and Aims

There is a need to evaluate the salt tolerance of plant species that can be cultivated as crops under saline conditions. Crambe maritima is a coastal plant, usually occurring on the driftline, with potential use as a vegetable crop. The aim of this experiment was to determine the growth response of Crambe maritima to various levels of airborne and soil-borne salinity and the ecophysiological mechanisms underlying these responses.

Methods

In the greenhouse, plants were exposed to salt spray (400 mm NaCl) as well as to various levels of root-zone salinity (RZS) of 0, 50, 100, 200 and 300 mm NaCl during 40 d. The salt tolerance of Crambe maritima was assessed by the relative growth rate (RGR) and its components. To study possible salinity effects on the tissue and cellular level, the leaf succulence, tissue Na⁺ concentrations, Na⁺ : K⁺ ratio, net K⁺/Na⁺ selectivity, N, P, K⁺, Ca²⁺, Mg²⁺, proline, soluble sugar concentrations, osmotic potential, total phenolics and antioxidant capacity were measured.

Key Results

Salt spray did not affect the RGR of Crambe maritima. However, leaf thickness and leaf succulence increased with salt spray. Root zone salinities up to 100 mm NaCl did not affect growth. However, at 200 mm NaCl RZS the RGR was reduced by 41 % compared with the control and by 56 % at 300 mm NaCl RZS. The reduced RGR with increasing RZS was largely due to the reduced specific leaf area, which was caused by increased leaf succulence as well as by increased leaf dry matter content. No changes in unit leaf rate were observed but increased RZS resulted in increased Na⁺ and proline concentrations, reduced K⁺, Ca²⁺ and Mg²⁺ concentrations, lower osmotic potential and increased antioxidant capacity. Proline concentrations of the leaves correlated strongly (r = 0·95) with RZS concentrations and not with plant growth.

Conclusions

Based on its growth response, Crambe maritima can be classified as a salt spray tolerant plant that is sensitive to root zone salinities exceeding 100 mm NaCl.