Role of TaALMT1 malate-GABA transporter in alkaline pH tolerance of wheat.

Malate exudation through wheat (Triticum aestivum L) aluminium-activated malate transporter 1 (TaALMT1) confers Al3+ tolerance at low pH, but is also activated by alkaline pH, and is regulated by and facilitates significant transport of gamma-aminobutyric acid (GABA, a zwitterionic buffer). Therefore, TaALMT1 may facilitate acidification of an alkaline rhizosphere by promoting exudation of both malate and GABA. Here, the performance of wheat near isogenic lines ET8 (Al+3 -tolerant, high TaALMT1 expression) and ES8 (Al+3 -sensitive, low TaALMT1 expression) are compared. Root growth (at 5 weeks) was higher for ET8 than ES8 at pH 9. ET8 roots exuded more malate and GABA at high pH and acidified the rhizosphere more rapidly. GABA and malate exudation was enhanced at high pH by the addition of aluminate in both ET8 and transgenic barley expressing TaALMT1. Xenopus laevis oocytes expressing TaALMT1 acidified an alkaline media more rapidly than controls corresponding to higher GABA efflux. TaALMT1 expression did not change under alkaline conditions but key genes involved in GABA turnover changed in accordance with a high rate of GABA synthesis. We propose that TaALMT1 plays a role in alkaline tolerance by exuding malate and GABA, possibly coupled to proton efflux.

Chemical Profile and Biological Activities of Essential Oil from Artemisia vulgaris L. Cultivated in Brazil.

Essential oil from the leaves of Artemisia vulgaris L. (Compositae) cultivated in Brazil was investigated for its chemical composition and biological activities including antibacterial, antifungal, and anthelmintic. The constituents of essential oils isolated by hydro-distillation were examined by GC-MS and a total of 18 components were identified. The essential oil was dominated by oxygenated sesquiterpenes (44.4%), sesquiterpene hydrocarbons (33.3%), and oxygenated monoterpenes (16.6%). Caryophyllene (37.45%), germacrene D (16.17%), and humulene (13.66%) were the major components. The essential oils from A. vulgaris showed bactericidal and fungicidal properties against Staphylococcus aureus and Candida albicans, respectively. Anthelmintic activity against Haemonchus contortus was absent in this essential oil. Altogether above results indicate that essential oils from A. vulgaris can be used for various medicinal purposes.

Cell-Type-Specific H+-ATPase Activity in Root Tissues Enables K+ Retention and Mediates Acclimation of Barley (Hordeum vulgare) to Salinity Stress.

While the importance of cell type specificity in plant adaptive responses is widely accepted, only a limited number of studies have addressed this issue at the functional level. We have combined electrophysiological, imaging, and biochemical techniques to reveal the physiological mechanisms conferring higher sensitivity of apical root cells to salinity in barley (Hordeum vulgare). We show that salinity application to the root apex arrests root growth in a highly tissue- and treatment-specific manner. Although salinity-induced transient net Na+ uptake was about 4-fold higher in the root apex compared with the mature zone, mature root cells accumulated more cytosolic and vacuolar Na+, suggesting that the higher sensitivity of apical cells to salt is not related to either enhanced Na+ exclusion or sequestration inside the root. Rather, the above differential sensitivity between the two zones originates from a 10-fold difference in K+ efflux between the mature zone and the apical region (much poorer in the root apex) of the root. Major factors contributing to this poor K+ retention ability are (1) an intrinsically lower H+-ATPase activity in the root apex, (2) greater salt-induced membrane depolarization, and (3) a higher reactive oxygen species production under NaCl and a larger density of reactive oxygen species-activated cation currents in the apex. Salinity treatment increased (2- to 5-fold) the content of 10 (out of 25 detected) amino acids in the root apex but not in the mature zone and changed the organic acid and sugar contents. The causal link between the observed changes in the root metabolic profile and the regulation of transporter activity is discussed.

Difference in root K+ retention ability and reduced sensitivity of K+-permeable channels to reactive oxygen species confer differential salt tolerance in three Brassica species.

Brassica species are known to possess significant inter and intraspecies variability in salinity stress tolerance, but the cell-specific mechanisms conferring this difference remain elusive. In this work, the role and relative contribution of several key plasma membrane transporters to salinity stress tolerance were evaluated in three Brassica species (B. napus, B. juncea, and B. oleracea) using a range of electrophysiological assays. Initial root growth assay and viability staining revealed that B. napus was most tolerant amongst the three species, followed by B. juncea and B. oleracea At the mechanistic level, this difference was conferred by at least three complementary physiological mechanisms: (i) higher Na(+) extrusion ability from roots resulting from increased expression and activity of plasma membrane SOS1-like Na(+)/H(+) exchangers; (ii) better root K(+) retention ability resulting from stress-inducible activation of H(+)-ATPase and ability to maintain more negative membrane potential under saline conditions; and (iii) reduced sensitivity of B. napus root K(+)-permeable channels to reactive oxygen species (ROS). The last two mechanisms played the dominant role and conferred most of the differential salt sensitivity between species. Brassica napus plants were also more efficient in preventing the stress-induced increase in GORK transcript levels and up-regulation of expression of AKT1, HAK5, and HKT1 transporter genes. Taken together, our data provide the mechanistic explanation for differential salt stress sensitivity amongst these species and shed light on transcriptional and post-translational regulation of key ion transport systems involved in the maintenance of the root plasma membrane potential and cytosolic K/Na ratio as a key attribute for salt tolerance in Brassica species.

GABA signalling modulates plant growth by directly regulating the activity of plant-specific anion transporters.

The non-protein amino acid, gamma-aminobutyric acid (GABA) rapidly accumulates in plant tissues in response to biotic and abiotic stress, and regulates plant growth. Until now it was not known whether GABA exerts its effects in plants through the regulation of carbon metabolism or via an unidentified signalling pathway. Here, we demonstrate that anion flux through plant aluminium-activated malate transporter (ALMT) proteins is activated by anions and negatively regulated by GABA. Site-directed mutagenesis of selected amino acids within ALMT proteins abolishes GABA efficacy but does not alter other transport properties. GABA modulation of ALMT activity results in altered root growth and altered root tolerance to alkaline pH, acid pH and aluminium ions. We propose that GABA exerts its multiple physiological effects in plants via ALMT, including the regulation of pollen tube and root growth, and that GABA can finally be considered a legitimate signalling molecule in both the plant and animal kingdoms.

Rapid regulation of the plasma membrane H⁺-ATPase activity is essential to salinity tolerance in two halophyte species, Atriplex lentiformis and Chenopodium quinoa.

BACKGROUND AND AIMS: The activity of H(+)-ATPase is essential for energizing the plasma membrane. It provides the driving force for potassium retention and uptake through voltage-gated channels and for Na(+) exclusion via Na(+)/H(+) exchangers. Both of these traits are central to plant salinity tolerance; however, whether the increased activity of H(+)-ATPase is a constitutive trait in halophyte species and whether this activity is upregulated at either the transcriptional or post-translation level remain disputed. METHODS: The kinetics of salt-induced net H(+), Na(+) and K(+) fluxes, membrane potential and AHA1/2/3 expression changes in the roots of two halophyte species, Atriplex lentiformis (saltbush) and Chenopodium quinoa (quinoa), were compared with data obtained from Arabidopsis thaliana roots. KEY RESULTS: Intrinsic (steady-state) membrane potential values were more negative in A. lentiformis and C. quinoa compared with arabidopsis (-144 ± 3·3, -138 ± 5·4 and -128 ± 3·3 mV, respectively). Treatment with 100 mm NaCl depolarized the root plasma membrane, an effect that was much stronger in arabidopsis. The extent of plasma membrane depolarization positively correlated with NaCl-induced stimulation of vanadate-sensitive H(+) efflux, Na(+) efflux and K(+) retention in roots (quinoa > saltbush > arabidopsis). NaCl-induced stimulation of H(+) efflux was most pronounced in the root elongation zone. In contrast, H(+)-ATPase AHA transcript levels were much higher in arabidopsis compared with quinoa plants, and 100 mm NaCl treatment led to a further 3-fold increase in AHA1 and AHA2 transcripts in arabidopsis but not in quinoa. CONCLUSIONS: Enhanced salinity tolerance in the halophyte species studied here is not related to the constitutively higher AHA transcript levels in the root epidermis, but to the plant's ability to rapidly upregulate plasma membrane H(+)-ATPase upon salinity treatment. This is necessary for assisting plants to maintain highly negative membrane potential values and to exclude Na(+), or enable better K(+) retention in the cytosol under saline conditions.

Ion transport in broad bean leaf mesophyll under saline conditions.

MAIN CONCLUSION: Salt stress reduces the ability of mesophyll tissue to respond to light. Potassium outward rectifying channels are responsible for 84 % of Na (+) induced potassium efflux from mesophyll cells. Modulation in ion transport of broad bean (Vicia faba L.) mesophyll to light under increased apoplastic salinity stress was investigated using vibrating ion-selective microelectrodes (the MIFE technique). Increased apoplastic Na(+) significantly affected mesophyll cells ability to respond to light by modulating ion transport across their membranes. Elevated apoplastic Na(+) also induced a significant K(+) efflux from mesophyll tissue. This efflux was mediated predominately by potassium outward rectifying channels (84 %) and the remainder of the efflux was through non-selective cation channels. NaCl treatment resulted in a reduction in photosystem II efficiency in a dose- and time-dependent manner. In particular, reductions in Fv'/Fm' were linked to K(+) homeostasis in the mesophyll tissue. Increased apoplastic Na(+) concentrations induced vanadate-sensitive net H(+) efflux, presumably mediated by the plasma membrane H(+)-ATPase. It is concluded that the observed pump's activation is essential for the maintenance of membrane potential and ion homeostasis in the cytoplasm of mesophyll under salt stress.

Polyamines cause plasma membrane depolarization, activate Ca2+-, and modulate H+-ATPase pump activity in pea roots.

Polyamines regulate a variety of cation and K(+) channels, but their potential effects on cation-transporting ATPases are underexplored. In this work, noninvasive microelectrode ion flux estimation and conventional microelectrode techniques were applied to study the effects of polyamines on Ca(2+) and H(+) transport and membrane potential in pea roots. Externally applied spermine or putrescine (1mM) equally activated eosin yellow (EY)-sensitive Ca(2+) pumping across the root epidermis and caused net H(+) influx or efflux. Proton influx induced by spermine was suppressed by EY, supporting the mechanism in which Ca(2+) pump imports 2 H(+) per each exported Ca(2+). Suppression of the Ca(2+) pump by EY diminished putrescine-induced net H(+) efflux instead of increasing it. Thus, activities of Ca(2+) and H(+) pumps were coupled, likely due to the H(+)-pump inhibition by intracellular Ca(2+). Additionally, spermine but not putrescine caused a direct inhibition of H(+) pumping in isolated plasma membrane vesicles. Spermine, spermidine, and putrescine (1mM) induced membrane depolarization by 70, 50, and 35 mV, respectively. Spermine-induced depolarization was abolished by cation transport blocker Gd(3+), was insensitive to anion channels' blocker niflumate, and was dependent on external Ca(2+). Further analysis showed that uptake of polyamines but not polyamine-induced cationic (K(+)+Ca(2+)+H(+)) fluxes were a main cause of membrane depolarization. Polyamine increase is a common component of plant stress responses. Activation of Ca(2+) efflux by polyamines and contrasting effects of polyamines on net H(+) fluxes and membrane potential can contribute to Ca(2+) signalling and modulate a variety of transport processes across the plasma membrane under stress.

Kinetics of xylem loading, membrane potential maintenance, and sensitivity of K(+) -permeable channels to reactive oxygen species: physiological traits that differentiate salinity tolerance between pea and barley.

Salt sensitive (pea) and salt tolerant (barley) species were used to understand the physiological basis of differential salinity tolerance in crops. Pea plants were much more efficient in restoring otherwise depolarized membrane potential thereby effectively decreasing K(+) efflux through depolarization-activated outward rectifying potassium channels. At the same time, pea root apex was 10-fold more sensitive to physiologically relevant H2 O2 concentration and accumulated larger amounts of H2 O2 under saline conditions. This resulted in a rapid loss of cell viability in the pea root apex. Barley plants rapidly loaded Na(+) into the xylem; this increase was only transient, and xylem and leaf Na(+) concentration remained at a steady level for weeks. On the contrary, pea plants restricted xylem Na(+) loading during the first few days of treatment but failed to prevent shoot Na(+) elevation in the long term. It is concluded that superior salinity tolerance of barley plants compared with pea is conferred by at least three different mechanisms: (1) efficient control of xylem Na(+) loading; (2) efficient control of H2 O2 accumulation and reduced sensitivity of non-selective cation channels to H2 O2 in the root apex; and (3) higher energy saving efficiency, with less ATP spent to maintain membrane potential under saline conditions.

Low-pH and aluminum resistance in arabidopsis correlates with high cytosolic magnesium content and increased magnesium uptake by plant roots.

Low-pH stress and Al(3+) toxicity affect root growth in acid soils. It was hypothesized that the capacity of genotypes to maintain Mg(2+) uptake in acidic environments may contribute to low-pH and Al resistance, but explicit evidence is lacking. In this work, an Al-resistant alr104 mutant and two Al-sensitive mutants (als5 and als3) of Arabidopsis thaliana were compared with the wild type (Col-0) for Mg(2+) uptake and intracellular Mg(2+) concentration under low-pH and combined low-pH/Al stresses. Magnesium accumulation in roots was measured in long-term (7 d) experiments. The Mg(2+) fluxes were measured using ion-sensitive microelectrodes at the distal elongation and the mature root zones in short-term (0-60 min) experiments. Intracellular Mg(2+) concentrations were measured in intact root cells at the distal elongation zone using magnesium-specific fluorescent dye and fluorescent lifetime imaging (FLIM) analysis. Under low-pH stress, Arabidopsis mutants als5 and alr104 maintained a higher Mg concentration in roots, and had greater Mg(2+) influx than the wild type and the als3 mutant. Under combined low-pH/Al treatment, Al-resistant genotypes (wild type and alr104) maintained a higher Mg(2+) accumulation, and had a higher Mg(2+) influx and higher intracellular Mg(2+) concentration than Al-sensitive genotypes (als3 and als5). Overall, these results show that increased Mg(2+) uptake correlates with an enhanced capacity of Arabidopsis genotypes to cope with low-pH and combined low-pH/Al stresses.

Haem oxygenase modifies salinity tolerance in Arabidopsis by controlling K⁺ retention via regulation of the plasma membrane H⁺-ATPase and by altering SOS1 transcript levels in roots.

Reactive oxygen species (ROS) production is a common denominator in a variety of biotic and abiotic stresses, including salinity. In recent years, haem oxygenase (HO; EC 1.14.99.3) has been described as an important component of the antioxidant defence system in both mammalian and plant systems. Moreover, a recent report on Arabidopsis demonstrated that HO overexpression resulted in an enhanced salinity tolerance in this species. However, physiological mechanisms and downstream targets responsible for the observed salinity tolerance in these HO mutants remain elusive. To address this gap, ion transport characteristics (K(+) and H(+) fluxes and membrane potentials) and gene expression profiles in the roots of Arabidopsis thaliana HO-overexpressing (35S:HY1-1/2/3/4) and loss-of-function (hy-100, ho2, ho3, and ho4) mutants were compared during salinity stress. Upon acute salt stress, HO-overexpressing mutants retained more K(+) (less efflux), and exhibited better membrane potential regulation (maintained more negative potential) and higher H(+) efflux activity in root epidermis, compared with loss-of-function mutants. Pharmacological experiments suggested that high activity of the plasma membrane H(+)-ATPase in HO overexpressor mutants provided the proton-motive force required for membrane potential maintenance and, hence, better K(+) retention. The gene expression analysis after 12h and 24h of salt stress revealed high expression levels of H(+)-ATPases (AHA1/2/3) and Na(+)/H(+) antiporter [salt overly sensitive1 (SOS1)] transcripts in the plasma membrane of HO overexpressors. It is concluded that HO modifies salinity tolerance in Arabidopsis by controlling K(+) retention via regulation of the plasma membrane H(+)-ATPase and by altering SOS1 transcript levels in roots.

Aluminum-dependent dynamics of ion transport in Arabidopsis: specificity of low pH and aluminum responses.

Low-pH and Al(3+) stresses are the major causes of poor plant growth in acidic soils. However, there is still a poor understanding of plant responses to low-pH and Al(3+) toxicity. Low-pH or combined low-pH and Al(3+) stress was imposed in order to measure rhizosphere pH, ion fluxes, plasma membrane potential and intracellular H(+) concentration in distal elongation and mature zones (MZs) along the longitudinal axis of Arabidopsis thaliana roots. Low-pH stress facilitated H(+) influx into root tissues and caused cytoplasmic acidification; by contrast, combined low-pH/Al(3+) treatment either decreased H(+) influx in the distal elongation zone (DEZ) or induced H(+) efflux in the MZ, leading to cytoplasmic alkalinization in both zones. Low-pH stress induced an increase in rhizosphere pH in the DEZ, whereas combined low-pH/Al(3+) stress resulted in lower rhizosphere pH in both root zones compared with the low-pH treatment alone. Low-pH stress facilitated K(+) efflux; the presence of Al(3+) diminished K(+) efflux or favored K(+) influx into root tissues. In both zones, low-pH treatment induced plasma membrane (PM) depolarization, which was significantly diminished (P