Evidence of fast serotonin transmission in frog slowly adapting type 1 responses

The Merkel cell–neurite (MCN) complex generates slowly adapting type 1 (SA1) response when mechanically stimulated. Both serotonin (5-HT) and glutamate have been implicated in the generation of normal SA1 responses, but previous studies have been inconclusive as to what their roles are or how synaptic transmission occurs. In this study, excised dorsal skin patches from common water frogs (Rana ridibunda) were stimulated by von Frey hairs during perfusion in a tissue bath, and single-unit spike activity was recorded from SA1 fibres. Serotonin had no significant effect on the SA1 response at low (10?µM) concentration, significantly increased activity in a force-independent manner at 100?µM, but decreased activity with reduced responsiveness to force at 1?mM. Glutamate showed no effect on the responsiveness to force at 100?µM. MDL 72222 (100?µM), an ionotropic 5-HT₃ receptor antagonist, completely abolished the responsiveness to force, suggesting that serotonin is released from Merkel cells as a result of mechanical stimulation, and activated 5-HT₃ receptors on the neurite. The metabotropic 5-HT₂ receptor antagonist, ketanserin, greatly reduced the SA1 fibre's responsiveness to force, as did the non-specific glutamate receptor antagonist, kynurenic acid. This supports a role for serotonin and glutamate as neuromodulators in the MCN complex, possibly by activation and/or inhibition of signalling cascades in the Merkel cell associated with vesicle release. Additionally, it was observed that SA1 responses contained a force-independent component, similar to a dynamic response observed during mechanical vibrations.     

Salicylic Acid Application Mitigates Oxidative Damage and Improves the Growth Performance of Barley under Drought Stress

Drought is a severe environmental constraint, causing a significant reduction in crop productivity across the world. Salicylic acid (SA) is an important plant growth regulator that helps plants cope with the adverse effects induced by various abiotic stresses. The current study investigated the potential effects of SA on drought tolerance efficacy in two barley (Hordeum vulgare) genotypes, namely BARI barley 5 and BARI barley 7. Ten-day-old barley seedlings were exposed to drought stress by maintaining 7.5% soil moisture content in the absence or presence of 0.5, 1.0 and 1.5 mM SA. Drought exposure led to severe damage to both genotypes, as indicated by phenotypic aberrations and reduction of dry biomass. On the other hand, the application of SA to drought-stressed plants protected both barley genotypes from the adverse effects of drought, which was reflected in the improvement of phenotypes and biomass production. SA supplementation improved relative water content and proline levels in drought-stressed barley genotypes, indicating the osmotic adjustment functions of SA under water-deficit conditions. Drought stress induced the accumulation of reactive oxygen species (ROS), such as hydrogen peroxide (H₂O₂) and superoxide (O₂ ^•− ), and the lipid peroxidation product malondialdehyde (MDA) in the leaves of barley plants. Exogenous supply of SA reduced oxidative damage by restricting the accumulation of ROS through the stimulation of the activities of key antioxidant enzymes, including superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), ascorbate peroxidase (APX) and glutathione peroxidase (GPX). Among the three-applied concentrations of SA, 0.5 mM SA exhibited better mitigating effects against drought stress considering the phenotypic performance and biochemical data. Furthermore, BARI barley 5 showed better performance under drought stress than BARI barley 7 in the presence of SA application. Collectively, our results suggest that SA played a crucial role in improving water status and antioxidant defense strategy to protect barley plants from the deleterious effects of water deficiency.     

Salicylic acid mitigates salinity stress by improving antioxidant defence system and enhances vincristine and vinblastine alkaloids production in periwinkle [<Emphasis Type="Italic">Catharanthus roseus</Emphasis> (L.) G. Don]

A pot experiment was conducted to find out whether the foliar spray of salicylic acid (SA) could successfully ameliorate the adverse effects of salinity stress on periwinkle. Thirty-day-old plants were supplied with Control; 0 mM NaCl + 10⁻⁵ M SA (T₁); 50 mM NaCl + 0 SA (T₂); 100 mM NaCl + 0 SA (T₃); 150 mM NaCl + 0 SA (T₄); 50 mM NaCl + 10⁻⁵ M SA (T₅); 100 mM NaCl + 10⁻⁵ M SA (T₆); 150 mM NaCl + 10⁻⁵ M SA (T₇). The plants were sampled 90 days after sowing to assess the effect of SA on stressed and unstressed plants. Salt stress significantly reduced the growth attributes including plant height, leaf-area index, shoot and root fresh weights, shoot and root dry weights. Increasing NaCl concentrations led to a gradual decrease in photosynthetic parameters and activities of nitrate reductase and carbonic anhydrase. Ascorbic acid, total alkaloids and antioxidants enzymes superoxide dismutase, catalase and peroxidase also declined in NaCl-treated plants. The plants, undergoing NaCl stress, exhibited a significant increase in electrolyte leakage and proline content. Foliar application of SA (10⁻⁵ M) reduced the damaging effect of salinity on plant growth and accelerated the restoration of growth processes. It not only improved the growth parameters but also reversed the effects of salinity. Total alkaloid content was improved by SA application both in unstressed and stressed plants. The highest level of total alkaloid content recorded in leaves of SA-treated stressed plants was 11.1%. Foliar spray of SA overcame the adverse effect of salinity by improving the content of vincristine (14.0%) and vinblastine (14.6%) in plants treated with 100 M NaCl.     

Abiotic Elicitors Influence Antioxidative Enzyme Activities and Shelf Life of Carrot During Storage Under Refrigerated Conditions

The objective of the present study was to investigate the effectiveness of the post-harvest treatments of abiotic elicitors, that is, calcium chloride (CaCl₂) and salicylic acid (SA) on physicochemical and biochemical parameters in relation to activities of antioxidative enzymes in carrot to enhance shelf life. Carrot of variety Punjab Carrot Red was harvested, washed, surface dried and treated with CaCl₂ (1, 1.5 and 2%) or SA (1, 1.5 and 2 mM) for 5 min, while distilled water was used as the control. Treated as well as untreated carrots were placed in open trays and stored under refrigerated (5 ± 1 °C, 90% RH) conditions for 63 days. Treatment of carrots with CaCl₂ and SA showed a reduction in changes in physiological weight, color, total soluble solids, ascorbic acid, titratable acidity, total phenolics, carotenoids, antioxidant activity and TBA reactive compound as compared to untreated samples. Higher activities of antioxidative enzymes, that is, catalase (CAT), superoxide dismutase (SOD), glutathione reductase (GR), peroxidase (POD), dehydro-ascorbate-reductase (DHAR) and monodehydro-ascorbate-reductase (MDHAR), were found in treated carrots as compared to untreated carrots during the whole storage period. SA treatment exhibited more usefulness in maintaining the quality of carrot than CaCl₂ treatment. Among all the treatments, 1.5 mM SA exhibited the highest antioxidative enzyme activities and slowest changes in biochemical quality of carrot during storage. Thus, 1.5 mM SA can be used to extend the shelf life of carrot during refrigerated storage.

     

The dual effects of salicylic acid on dehydrin accumulation in water-stressed barley seedlings

Dehydrins are a group of plant proteins that usually accumulate in response to environmental stresses. They are proposed to play specific protective roles in plant cells. Present study showed that the accumulation of dehydrins in water-stressed barley (Hordeum vulgare L.) seedlings was influenced by their treatment with salicylic acid (SA). The level of dehydrin proteins was increased by 0.20 mM SA, but decreased by 0.50 mM SA treatment. Both mRNA expression and protein accumulation of a typical barley dehydrin, DHN5, were enhanced by SA treatment when SA concentrations were lower than 0.25 mM. However, the higher SA concentrations significantly decreased the protein level of DHN5 despite of a stable mRNA level. Our results also showed that low SA concentrations (less than 0.25 mM) decreased the electrolyte leakage and malondialdehyde (MDA) and H₂O₂ contents in water-stressed barley seedlings. But high SA concentrations (more than 0.25 mM) enhanced H₂O₂ accumulation, tended to cause more electrolyte leakage, and increase MDA content. These data indicated that SA could up-regulate the dehydrin gene expression and protein accumulation. Since the protective role of dehydrins in plant cells, such effect could be an important reason for the SA-mediated alleviation on water stress injury. But excessive SA could suppress the accumulation of dehydrin proteins and aggravate the oxidative damage. Published in Russian in Fiziologiya Rastenii, 2009, Vol. 56, No. 3, pp. 388–394. This text was submitted by the authors in English.     

Preparation and Evaluation of Microemulsion Systems Containing Salicylic Acid

Microemulsions (MEs) are clear, thermodynamically stable systems. They were used to solubilize drugs and to improve topical drug availability. Salicylic acid (SA) is a keratolytic agent used in topical products with antimicrobial actions. The objective of this work was to prepare and evaluate SA ME systems. Different concentrations of SA were incorporated in an ME base composed of isopropyl myristate, water, and Tween 80: propylene glycol in the ratio of 15:1. Three ME systems were prepared: S_2%, S_5%, and S_10% which contain 2%, 5%, and 10% of SA, respectively. Evaluation by examination under cross-polarizing microscope, measuring of percent transmittance, pH measurement, determination of the specific gravity, assessment of rheological properties, and accelerated stability study were carried out. The data showed that the addition of SA markedly affected the physical properties of the base. All systems were not affected by accelerated stability tests. Stability study for 6 months under ambient conditions was carried out for S_10%. No remarkable changes were recorded except a decrease in the viscosity value after 1 month. The results suggested that ME could be a suitable vehicle for topical application of different concentrations of SA.     

闽江口互花米草淤积作用对其自身和短叶茳芏残体分解及硫养分释放的影响

2016—2017年,以闽江口鳝鱼滩西北部互花米草(SA)入侵初期与短叶茳芏(CM)形成的典型交错带植物残体为研究对象,基于野外原位分解试验,通过设定无淤积强度(S_0,0 cm/a)、当前淤积强度(S_5,5 cm/a)和未来淤积增强(S_(10),10 cm/a)3种处理,模拟互花米草入侵初期导致的淤积作用对其自身以及短叶茳芏残体分解及硫养分释放的影响。结果表明,随着互花米草入侵导致的淤积强度的增加,互花米草和短叶茳芏残体的分解速率均明显降低;与S_0相比,二者在S_5与S_(10)处理下的分解速率分别降低49.09%(SA)、35.14%(CM)和56.36%(SA)、44.59%(CM)。随着淤积强度的增加,互花米草和短叶茳芏残体分解过程中的TS含量整体均呈增加趋势,且其对短叶茳芏TS含量变化的影响较为明显;互花米草和短叶茳芏残体在分解过程中均表现为不同程度的硫释放,但随淤积强度的增加,二者硫释放量均呈降低趋势,且在相同淤积强度下,前者的硫释放量要高于后者。不同淤积强度下残体分解速率及硫养分释放强度的差异不仅与分解环境中的EC密切相关,且与残体残留率、初始基质质量(C/N和C/S)以及淤积导致养分条件改变而对分解过程中残体基质质量的影响有关。研究发现,随着淤积强度的增加,两种残体的分解速率及硫释放强度均降低;但在相同淤积强度下,短叶茳芏残体的分解速率和硫释放量均大于互花米草。     

The mechanism of stomatal closing by salicylic acid inCommelina communis L.

The mechanism of stomatal closing by salicylic acid (SA) has been investigated. The addition of 1 mM SA to fully opened stomata resulted in a significant reduction of 75% in stomatal aperture. Stomata in the treatment of SA with EGTA closed as observed in the treatment of SA. However, the addition of catalase with SA completely inhibited stomatal closing. Stomatal closing induced by SA was also reduced by Ca²⁺. To understand the relation bewteen stomatal closing by SA and catalase activity, the effect of SA on catalse activity and the effect of AT (catalase inhibitor) on stomatal closing was investigated. SA inhibited 32% of catalase activity. Stomata in isolated epidermis floated on an incubation solution containing 0.1 mM AT closed from 9.6 μm to 3.2 μm after 1 hour. SA stimulated K⁺ efflux as much as the twice of the control in isolated strips. SA inhibited 53% of photosynthetic activity at the light intensity of 1000 μmole m² s¹ on SA infiltrated leaves. A similar result was found on stomatal conductance in SA infiltrated leaves. These results indicate that SA inhibit catalase activity and increase the concentration of H₂O₂ in guard cell cytoplasm. H₂O₂ oxidize the plasma membrane and increase the membrane permeability of K⁺. The mass efflux of K⁺ induce the loss of turgor pressure and lead to stomatal closing. The inhibition of photosynthetic activity by SA suggests that stomatal closing by SA is also related with the decrease of photosynthetic activity.     

The Postnatal Functional Development of Muscle Stretch Receptors in the Rat

The response to a 5-sec stretch of the triceps muscle was studied in dorsal root filaments L₅ of 72 infant rats (1-19 days old) under urethane anesthesia.

More than 50% of all units in 1-day-old rats responded by repetitive firing until the end of the 5-sec stretch (slowly adapting or SA receptors), while the rest ceased to fire earlier (relatively rapidly adapting or l/2 SA receptors), or gave an “on” response only. The number of units exhibiting an SA response increased with age and attained 80% in 5-day-old rats. By the 10th day of life, almost 90% of endings behaved as SA receptors. During development, the maximal discharge frequencies at the peak of stretch increased markedly, and their values in 18-day-old rats were comparable to those in adult rats. The phasic component of the response to stretch, although less well defined in the younger animals, was already present even in 1 -day-old rats. Adaptation of the static response during maintained stretch was relatively steep in all the age groups studied.

The results indicate that, in the rat, large numbers of muscle stretch receptors are capable of responding to sustained stretch as SA receptors, even at an age when their morphological and ultra-structural maturation is not yet fully accomplished.     

Role of salicylic acid and oxidative burst in expression of resistance to Alternaria blight

The accumulation of salicylic acid and H₂O₂ during pathogenic infection of mustard plants with Alternaria brassicae spores was investigated to understand the role of these two defense compounds in the expression of resistance. Comparisons were made between a susceptible Brassica juncea variety RH30 and a Brassica carinata variety HC1, which is known to be resistant. An oxidative burst was detected as in situ accumulation of H₂O₂, in both the Brassica spp. after pathogen application. However, H₂O₂ generation was extracellular in the resistant variety and both extra- and intracellular in the susceptible variety. Endogenous levels of SA increased over 2.5-fold in the resistant variety HC1 in response to pathogen application and this increase was observed only in conjugated SA levels. Pathogen application also led to an increase in the antioxidant enzymes, guaiacol-dependent peroxidase (GDP) and superoxide dismutase (SOD) in HC1. Exogenous SA application to leaves led to over threefold increase in the free and conjugated SA levels in both varieties. Pathogen application to the SA pretreated plants led to over 10-fold increase in endogenous SA levels in both varieties as compared to the levels in controls and this correlated with a decrease in disease symptoms in both species. SA appeared to regulate defense responses in Brassica spp. in a concentration-dependent manner. While 2.7-fold increase in endogenous SA levels (as seen in HC1) led to an induction of antioxidant enzymes, over 10-fold increases in endogenous SA levels (as seen after exogenous SA application in both varieties) brought about no induction of the antioxidant enzymes, probably because SA itself served as an antioxidant.     

Partial purification and properties of an inducible uridine 5'-diphosphate-glucose-salicylic Acid glucosyltransferase from oat roots

A salicylic acid (SA)-inducible uridine 5′-diphosphate (UDP)-glucose:SA 3-O-glucosyltransferase was extracted from oat (Avena sativa L. cv Dal) roots. Reverse phase high-performance liquid chromatography or anion exchange chromatography was used to separate SA from the product, β-O-d-glucosylsalicylic acid. The soluble enzyme was purified 176-fold with 5% recovery using a combination of pH fractionation, anion exchange, gel filtration, and chromatofocusing chromatography. The partially purified protein had a native molecular weight of about 50,000, an apparent isoelectric point at pH 5.0, and maximum activity at pH 5.5. The enzyme had a K_m of 0.28 mm for UDP-glucose and was highly specific for this sugar donor. More than 20 hydroxybenzoic and hydroxycinnamic acid derivatives were assayed as potential glucose acceptors. UDP-glucose:SA 3-O-glucosyltransferase activity was highly specific toward SA (K_m = 0.16 mm). The enzyme was inhibited by UDP and uridine 5′-triphosphate but not by up to 7.5 mm uridine 5′-monophosphate.     

Uptake and Metabolism of [C]Salicylic Acid in Lemna gibba G3

When the long-day plant Lemna gibba L., strain G3 is grown under continuous light on ammonium-free half-strength Hutner's medium (NH₄⁺-free 0.5 H medium) there is virtually no flowering, but addition of 10 micromolar salicylic acid (SA) to the medium results in substantial flowering. Using this system, the uptake and metabolism of [¹⁴C]SA in L. gibba G3 has been examined. SA uptake is rapid and linear for at least the first 24 hours. After 30 minutes, nearly 90% of the radioactivity in the plants is present as free SA. Part of this is rapidly converted to one or more bound forms of SA that appear either in the acidic butanol fraction or in the aqueous residue, and after 12 hours an equilibrium is reached between the free and bound forms of SA. When plants receive SA for 6 days and then are switched to control medium, both the free and bound SA remain nearly constant for at least 5 days. However, there is virtually no transfer of SA from mother fronds to daughter fronds, indicating that the SA is apparently sequestered within the cell. Cell fractionation studies show that nearly 95% of the SA remains in the supernatant even after a 2-hour centrifugation at 300,000 g. Thus, it is unlikely that SA is confined within a specific organelle, but rather is probably secreted into the vacuole.     

Salicylic acid and H2O2 function by independent pathways in the induction of freezing tolerance in potato

The effects of salicylic acid (SA) and hydrogen peroxide (H₂O₂) on freezing tolerance were studied in two potato (Solanum tuberosum) cultivars (Alpha and Atlantic) that differ in cold sensitivity, Alpha being more tolerant to freezing than Atlantic. Lowest freezing survival rates were observed in 4-week-old plants. Freezing treatments consisting of exposure to 6° C for 4 h in the dark were applied 24 h after plants had been transferred from in vitro culture to soil. Catalase activity and H₂O₂ were estimated at the following harvest points: stage (a) 4-week-old in vitro plants treated with either 0.1 mM SA or 5 mM H₂O₂; stage (b) as in (a) but 24 h following transfer to soil prior to freezing treatment; stage (c) as in (b) but measured 15 days after a 4-h freezing treatment. The results show that (1) SA induced freezing tolerance in both cultivars; (2) SA inhibited ascorbate peroxidase activities in both cultivars at all harvest points but inhibited catalase activities in only at stage (a); (3) SA induced H₂O₂ accumulation only in Atlantic at stage (a); (4) H₂O₂ enhanced shoot catalase activities in Atlantic at stages (a) and (b) whereas this treatment had no effect on shoot catalase activities in Alpha; (5) H₂O₂ treatment induced freezing tolerance in Atlantic, even though shoot catalase activities were lower than those of the controls following exposure to freezing temperatures. We conclude that SA does not always lead to H₂O₂ accumulation even though catalase and ascorbate peroxidase activities are decreased as a result of the treatment. Moreover, H₂O₂ accumulation is not always associated with the induction of freezing tolerance, for example at stage (a) where SA-induced tolerance in Alpha was not accompanied by H₂O₂ accumulation. H₂O₂ was able to induce freezing tolerance only in Atlantic, even though H₂O₂ accumulated in both cultivars following this treatment.     

Effect of Gibberellic Acid and Some Phenols on Flowering of Impatiens balsamina, a Qualitative Short-day Plant

GA₃ as well as SA (salicylic acid) and β-N (β-naphthol) induce floral buds in Impatiens balsamina under strictly non-inductive photoperiods. The floral bud initiation is accelerated when 1 mg/1 SA is used in combination with 100 mg/1 GA₃. 100 mg/1 GA₃+ 1 mg/1 SA and 100 mg/1 GA₃+ 100 mg/1 β-N increase the number of floral buds as compared with 100 mg/1 GA₃ alone.     

Signaling Effects of Nitric Oxide, Salicylic Acid, and Reactive Oxygen Species on Isoeuphpekinensin Accumulation in Euphorbia pekinensis Suspension Cells Induced by an Endophytic Fungal Elicitor

Nitric oxide (NO), salicylic acid (SA), and reactive oxygen species (ROS) are important signal molecules that mediate plant resistance reactions and play important roles in secondary metabolism. To research the signal transduction pathway of the endophytic fungal elicitor from Fusarium sp. E5 promoting secondary metabolism in Euphorbia pekinensis suspension cells, the changes in NO, SA, ROS, and isoeuphpekinensin contents in the cells were investigated after elicitor addition to the cell suspension culture. The elicitor did not change H₂O₂ or O₂ ^? contents notably, whereas NO and SA contents were enhanced. Both the NO donator sodium nitroprusside (SNP) and SA enhanced isoeuphpekinensin content in the absence of the fungal elicitor, whereas the NO scavenger cPTIO and SA biosynthesis inhibitor cinnamic acid (CA) inhibited isoeuphpekinensin accumulation in the presence of the elicitor. In addition, cPTIO inhibited SA production induced by the fungal elicitor. CA did not inhibit NO production, but it significantly inhibited isoeuphpekinensin accumulation. The results demonstrated that in Euphorbia pekinensis suspension cells the endophytic fungal elicitor induced increased NO content and SA production, which promoted isoeuphpekinensin accumulation. ROS are clearly not involved in the endophytic fungus–host interaction signaling pathway.     

Hydrogen Peroxide Is a Second Messenger in the Salicylic Acid-Triggered Adventitious Rooting Process in Mung Bean Seedlings

In plants, salicylic acid (SA) is a signaling molecule that regulates disease resistance responses, such as systemic acquired resistance (SAR) and hypertensive response (HR). SA has been implicated as participating in various biotic and abiotic stresses. This study was conducted to investigate the role of SA in adventitious root formation (ARF) in mung bean (Phaseolus radiatus L) hypocotyl cuttings. We observed that hypocotyl treatment with SA could significantly promote the adventitious root formation, and its effects were dose and time dependent. Explants treated with SA displayed a 130% increase in adventitious root number compared with control seedlings. The role of SA in mung bean hypocotyl ARF as well as its interaction with hydrogen peroxide (H₂O₂) were also elucidated. Pretreatment of mung bean explants with N, N’-dimethylthiourea (DMTU), a scavenger for H₂O₂, resulted in a significant reduction of SA-induced ARF. Diphenyleneiodonium (DPI), a specific inhibitor of membrane-linked NADPH oxidase, also inhibited the effect of adventitious rooting triggered by SA treatment. The determination of the endogenous H₂O₂ level indicated that the seedlings treated with SA could induce H₂O₂ accumulation compared with the control treatment. Our results revealed a distinctive role of SA in the promotion of adventitious rooting via the process of H₂O₂ accumulation. This conclusion was further supported by antioxidant enzyme activity assays. Based on these results, we conclude that the accumulation of free H₂O₂ might be a downstream event in response to SA-triggered adventitious root formation in mung bean seedlings.     

Additive, dominant, and epistatic effects for maize grain yield in acid and non-acid soils

Acid soils severely reduce maize (Zea mays L.) yield in the tropics. Breeding for tolerance to soil acidity provides a permanent, environmentally friendly, and inexpensive solution to the problem. This study was carried out to determine the relative importance of additive, dominant, and epistatic effects on maize grain-yields in different tropical genotypes. Divergent selection in three populations (SA4, SA5, and SA7) provided inbred lines tolerant or sensitive to acid soils. The tolerant and sensitive lines from each population were used to obtain the F₁, F₂, F₃, back-crosses, second back-crosses, and selfed back-cross generations. In addition, the tolerant lines from SA4 and SA5 were crossed with a sensitive line from the Tuxpeño Sequía population, from which the same generations were also derived. All generations from each of the five sets of crosses were evaluated in three acid-soil environments and one non-acid-soil environment. A generation-mean analysis was performed on each set for yield. The sequential sum of squares associated with additive, dominance, and digenic epistatic effects were used to estimate the relative importance of each genetic effect. Epistasis was not important in any set in the non-acid-soil environment, with dominance accounting for 80.76% of the total variation among generation means across sets. In acid-soil environments, epistasis was more important. The relative importance of digenic epistasis was greater in those evaluations with large experimental errors. The tolerant line from population SA5 was prone to severe root lodging, suggesting a very poor root system. Apparently, the tolerance to soil acidity in this line is not associated with a large root system.     

Salicylic acid-induced antioxidant protection against low temperature in cold-hardy winter wheat

“Dongnongdongmai 1” is a cultivated winter wheat which can endure cold temperature as low as ?30 °C with a reviving rate of 85 %. We aimed to explore the involvement of antioxidant protection system in salicylic acid (SA)-enhanced cold resistance of winter wheat. Seedlings were prayed with 0.1 mM SA at three-leaf stage, followed by cold acclimation at tillering stage (4 °C for 5 days) prior to cold treatment at 4, 0, ?10 or ?20 °C for 2 days. Under low temperature, the relative electrical conductivity (REC) of rhizomes and H₂O₂ content in rhizomes were lower compared with leaves, while in the reactive oxygen species (ROS) removal system, only the POD activity was higher. Foliar spray with SA significantly inhibited the cold-increased REC of rhizomes at ?20 °C and REC of leaves at ?10 and ?20 °C. In addition, application of SA prior to ?10 or ?20 °C treatment suppressed the increase in H₂O₂ content both in rhizomes and leaves. SA enhanced the activities of SOD, POD, and CAT in wheat following low-temperature treatment, especially at ?10 and ?20 °C. In addition, spray with SA resulted in 1.1-to-4.9-fold enhanced activities of the key enzymes in AsA–GSH cycle, including APX, DHAR, and MDHAR. Our results suggested that SA could improve the resistance of winter wheat against extreme low temperature by enhancing the activities of antioxidases to eliminate ROS and maintain the redox homeostasis. In addition, the less damage to rhizomes in comparison with leaves may be attributed to enhanced POD activity.     

Parallel Changes in H2O2 and Catalase during Thermotolerance Induced by Salicylic Acid or Heat Acclimation in Mustard Seedlings

Spraying mustard (Sinapis alba L.) seedlings with salicylic acid (SA) solutions between 10 and 500 μm significantly improved their tolerance to a subsequent heat shock at 55°C for 1.5 h. The effects of SA were concentration dependent, with higher concentrations failing to induce thermotolerance. The time course of thermotolerance induced by 100 μm SA was similar to that obtained with seedlings acclimated at 45°C for 1 h. We examined the hypothesis that induced thermotolerance involved H₂O₂. Heat shock at 55°C caused a significant increase in endogenous H₂O₂ and reduced catalase activity. A peak in H₂O₂ content was observed within 5 min of either SA treatment or transfer to the 45°C acclimation temperature. Between 2 and 3 h after SA treatment or heat acclimation, both H₂O₂ and catalase activity significantly decreased below control levels. The lowered H₂O₂ content and catalase activity occurred in the period of maximum thermoprotection. It is suggested that thermoprotection obtained either by spraying SA or by heat acclimation may be achieved by a common signal transduction pathway involving an early increase in H₂O₂.     

Altered Kinetic Properties of Tyrosine-183 to Cysteine Mutation in Glutamine Synthetase of <Emphasis Type="Italic">Anabaena variabilis</Emphasis> Strain SA1 Is Responsible for Excretion of Ammonium Ion Produced by Nitrogenase

A L-methionine-D,L-sulfoximine-resistant mutant of the cyanobacterium Anabaena variabilis, strain SA1, excreted the ammonium ion generated from N₂ reduction. In order to determine the biochemical basis for the NH₄ ⁺-excretion phenotype, glutamine synthetase (GS) was purified from both the parent strain SA0 and from the mutant. GS from strain SA0 (SA0-GS) had a pH optimum of 7.5, while the pH optimum for GS from strain SA1 (SA1-GS) was 6.8. SA1-GS required Mn⁺² for optimum activity, while SA0-GS was Mg⁺² dependent. SA0-GS had the following apparent K _m values at pH 7.5: glutamate, 1.7 mM; NH₄ ⁺, 0.015 mM; ATP, 0.13 mM. The apparent K _m for substrates was significantly higher for SA1-GS at its optimum pH (glutamate, 9.2 mM; NH₄ ⁺, 12.4 mM; ATP, 0.17 mM). The amino acids alanine, aspartate, cystine, glycine, and serine inhibited SA1-GS less severely than the SA0-GS. The nucleotide sequences of glnA (encoding glutamine synthetase) from strains SA0 and SA1 were identical except for a single nucleotide substitution that resulted in a Y183C mutation in SA1-GS. The kinetic properties of SA1-GS isolated from E. coli or Klebsiella oxytoca glnA mutants carrying the A. variabilis SA1 glnA gene were also similar to SA1-GS isolated from A. variabilis strain SA1. These results show that the NH₄ ⁺-excretion phenotype of A. variabilis strain SA1 is a direct consequence of structural changes in SA1-GS induced by the Y183C mutation, which elevated the K _m values for NH₄ ⁺ and glutamate, and thus limited the assimilation of NH₄ ⁺ generated by N₂ reduction. These properties and the altered divalent cation-mediated stability of A. variabilis SA1-GS demonstrate the importance of Y183 for NH₄ ⁺ binding and metal ion coordination. Received: 3 July 2002 / Accepted: 29 July 2002