Localized Wounding by Heat Initiates the Accumulation of Proteinase Inhibitor II in Abscisic Acid-Deficient Plants by Triggering Jasmonic Acid Biosynthesis

To test whether the response to electrical current and heat treatment is due to the same signaling pathway that mediates mechanical wounding, we analyzed the effect of electric-current application and localized burning on proteinase inhibitor II (Pin2) gene expression in both wild-type and abscisic acid (ABA)-deficient tomato (Lycopersicon esculentum Mill.) and potato (Solanum phureja) plants. Electric-current application and localized burning led to the accumulation of Pin2 mRNA in potato and tomato wild-type plants. Among the treatments tested, only localized burning of the leaves led to an accumulation of Pin2 mRNA in the ABA-deficient plants. Electric-current application, like mechanical injury, was able to initiate ABA and jasmonic acid (JA) accumulation in wild-type but not in ABA-deficient plants. In contrast, heat treatment led to an accumulation of JA in both wild-type and ABA-deficient plants. Inhibition of JA biosynthesis by aspirin blocked the heat-induced Pin2 gene expression in tomato wild-type leaves. These results suggest that electric current, similar to mechanical wounding, requires the presence of ABA to induce Pin2 gene expression. Conversely, burning of the leaves activates Pin2 gene expression by directly triggering the biosynthesis of JA by an alternative pathway that is independent of endogenous ABA levels.     

Characterization of a wilty sunflower (Helianthus annuus L.) mutant: III. Phenotypic interaction in reciprocal grafts from wilty mutant and wild-type plants

The present study was conducted to evaluate phenotypic interactionin reciprocal grafts between wilty (w-1) sunflower mutant andnormal (W-1) plants. The w-1 genotype is a ‘leaky’ABA-deficient mutant, characterized by high stomatal conductance,in both light and dark conditions, and high transpiration rate. In well-watered conditions, mutant scions grafted on to normalrootstock (w-1/W-1) showed higher leaf relative water content,leaf water potential and ABA levels than those of control grafts(w-1/w-1). In addition, detached leaves of w-1/W-1 exhibitedlower water loss than w-1/w-1 grafts, while mutant rootstockdid not affect the transpiration rate of detached W-1 leaves.When drought stress was imposed to potted plants by withholdingwater, the mutant scions grafted on to normal roots showed apartial phenotypic reversion. A rapid stomatal closure and arise in ABA levels in response to a small decrease in leaf waterpotential was observed. By contrast, in w-1/w-1 grafts significantreductions in stomatal conductance and ABA accumulation weredetected only in conjunction with a severe water deficit. W-1scions on mutant stocks (W-1/w-1) maintained the normal phenotypeof control wild-type grafts (W1/W-1). Key words: ABA, grafting, Helianthus annuus, stomatal conductance, water relations, wilty mutant     

Abscisic acid-deficient plants do not accumulate proteinase inhibitor II following systemin treatment

The role of systemin inPin2 gene expression was analyzed in wild-type plants of potato (Solanum tuberosum L.) and tomato (Lycopersicon esculentum Mill.), as well as in abscisic acid (ABA)-deficient tomato (sitiens) and potato (droopy) plants. The results showed that systemin initiates Pin2 mRNA accumulation only in wildtype tomato and potato plants. As in the situation after mechanical wounding,Pin2 gene expression in ABA-deficient plants was not activated by systemin. Increased endogenous levels of jasmonic acid (JA) and accumulation of Pin2 mRNA were observed following treatment with α-linolenic acid, the precursor of JA biosynthesis, suggesting that these ABA mutants still have the capability to synthesize de novo JA. Measurement of endogenous levels of ABA and JA showed that systemin leads to an increase of both phytohormones (ABA and JA) only in wild-type but not in ABA-deficient plants.     

A cotton mitogen-activated protein kinase （GhMPK6） is involved in ABA-induced CAT1 expression and H2O2 production

The mitogen-activated protein kinase (MAPK) cascade is one of the major and evolutionally conserved signaling pathways and plays a pivotal role in the regulation of stress and developmental signals in plants.Here,we identified one gene,GhMPK6,encoding an MAPK protein in cotton.GFP fluorescence assay demonstrated that GhMAPK6 is a cytoplasm localized protein.Quantitative RT-PCR analysis revealed that mRNA accumulation of GhMPK6 was significantly promoted by abscisic acid (ABA).Overexpression of GhMPK6 gene in the T-DNA insertion mutant atmkkl (SALK_015914) conferred a wild-type phenotype to the transgenic plants in response to ABA.Under ABA treatment,cotyledon greening/expansion in GhMPK6 transgenic lines and wild type was significantly inhibited,whereas the atmkkl mutant showed a relatively high cotyledon greening/expansion ratio.Furthermore,CAT1 expression and H2O2 levels in leaves of GhMPK6 transgenic lines and wild type were remarkably higher than those of atmkkl mutant with ABA treatment.Collectively,our results suggested that GhMPK6 may play an important role in ABA-induced CAT1 expression and H2O2 production.     

NaCl胁迫下抗盐突变体和野生型小麦Na~ 、K~ 累积的差异分析

对抗盐突变体和野生型小麦（ＴｒｉｔｉｃｕｍａｅｓｔｉｖｕｍＬ．）在盐胁迫下Ｎａ＋、Ｋ＋的累积状况进行了比较研究。结果表明,ＮａＣｌ胁迫下突变体根和叶中Ｎａ＋的相对累积较野生型显著地少,同时Ｎａ＋的净累积速率显著地低。这种Ｎａ＋相对累积少的状况在叶中表现尤为明显。两者叶中Ｋ＋含量随盐浓度的增加显著下降,但突变体的含量均高于野生型。突变体根中Ｋ＋的含量也显著高于野生型,且这种差异随盐浓度的提高而增大。分别统计突变体和野生型根和叶中Ｎａ＋的含量以及每株幼苗的Ｎａ＋总量以分析二者Ｎａ＋在根和叶中的分布差异,结果表明３００ｍｍｏｌ／ＬＮａＣｌ胁迫９６ｈ后,突变体根中Ｎａ＋的累积量占其整株幼苗Ｎａ＋累积总量的４４４％,而野生型根中含量占其总量的２４３％。相对于野生型而言,在盐胁迫下突变体根中Ｎａ＋分布比例的提高可有效地减少根中Ｎａ＋向地上部分转运。     

Abscisic acid deficiency leads to rapid activation of tomato defence responses upon infection with Erwinia chrysanthemi

In addition to the important role of abscisic acid (ABA) in abiotic stress signalling, basal and high ABA levels appear to have a negative effect on disease resistance. Using the ABA-deficient sitiens tomato ( Solanum lycopersicum ) mutant and different application methods of exogenous ABA, we demonstrated the influence of this plant hormone on disease progression of Erwinia chrysanthemi . This necrotrophic plant pathogenic bacterium is responsible for soft rot disease on many plant species, causing maceration symptoms mainly due to the production and secretion of pectinolytic enzymes. On wild-type (WT) tomato cv. Moneymaker E. chrysanthemi leaf inoculation resulted in maceration both within and beyond the infiltrated zone of the leaf, but sitiens showed a very low occurrence of tissue maceration, which never extended the infiltrated zone. A single ABA treatment prior to infection eliminated the effect of pathogen restriction in sitiens , while repeated ABA spraying during plant development rendered both WT and sitiens very susceptible. Quantification of E. chrysanthemi populations inside the leaf did not reveal differences in bacterial growth between sitiens and WT. Sitiens was not more resistant to pectinolytic cell-wall degradation, but upon infection it showed a faster and stronger activation of defence responses than WT, such as hydrogen peroxide accumulation, peroxidase activation and cell-wall fortifications. Moreover, the rapid activation of sitiens peroxidases was also observed after application of bacteria-free culture filtrate containing E. chrysanthemi cell-wall-degrading enzymes and was absent during infection with an out E. chrysanthemi mutant impaired in secretion of these extracellular enzymes.     

Abscisic acid is a key inducer of hydrogen peroxide production in leaves of maize plants exposed to water stress

The histochemical and cytochemical localization of water stress-induced H(2)O(2) production in the leaves of ABA-deficient vp5 mutant and wild-type maize (Zea mays L.) plants were examined, using 3,3-diaminobenzidine and CeCl(3) staining, respectively, and the roles of endogenous ABA in the production of H(2)O(2) induced by water stress were assessed. Water stress induced by polyethylene glycol resulted in the accumulation of H(2)O(2) in mesophyll cells, bundle-sheath cells and vascular bundles of wild-type maize leaves, and the accumulation was substantially blocked in the mutant maize leaves exposed to water stress. Pre-treatments with several apoplastic H(2)O(2) manipulators abolished the majority of H(2)O(2) accumulation induced by water stress in the wild-type leaves. The subcellular localization of H(2)O(2) production was demonstrated in the cell walls, xylem vessels, chloroplasts, mitochondria and peroxisomes in the leaves of wild-type maize plants exposed to water stress, and the accumulation of H(2)O(2) induced by water stress in the cell walls and xylem vessels, but not in the chloroplasts, mitochondria and peroxisomes, was arrested in the leaves of the ABA mutant or the ABA biosynthesis inhibitor (tungstate)-pre-treated maize plants. Pre-treatments with the apoplastic H(2)O(2) manipulators also blocked the apoplastic but not the intracellular H(2)O(2) accumulation induced by water stress in the leaves of wild-type plants. These data indicate that under water stress, the apoplast is the major source of H(2)O(2) production and ABA is a key inducer of apoplastic H(2)O(2) production. These data also suggest that H(2)O(2) generated in the apoplast could not diffuse freely into subcellular compartments.     

Proteinase Inhibitor II Gene Expression Induced by Electrical Stimulation and Control of Photosynthetic Activity in Tomato Plants

Mechanical damage and heat stimulation were used to activateproteinase inhibitor II (Pin2) gene expression in tomato plantsin both treated (local induction) and non-treated tissues (systemicinduction). Both stimuli have been shown to generate electricalsignals, leading to a systemic activation of gene expression.Treatment of tomato leaves with electrical current resultedin the accumulation of Pin2 mRNA in the local and systemic leaves.Additionally, all treatments inducing Pin2 gene activity gaverise to a significant alteration of stomatal aperture. However,heat stimulation provoked a different response in the stomatalparameters than mechanical wounding or electric treatment. Bothmechanical damage and electrical stimulation activated two characteristictime constants in the gas exchange relaxation kinetics. Conversely,heat stimulation resulted in only one major time constant. Theresults clearly show that direct current application to tomatoleaves initiates Pin2 mRNA accumulation locally and systemically.In addition, they suggest the participation of a second slowelectrical/hydraulic component in the wound response mechanismof tomato plants and a possible alternative pathway regulatingheat-induced Pin2 gene expression. (Received February 13, 1995; Accepted April 14, 1995)     

Difference in Na+,K+ Accumulation in the Salt tolerant Mutant and the Wild Type of Wheat During Exposure to NaCl Stress

The difference in Na+, K+ accumulation between the mutant and the wild type of wheat ( Triticum aestivum L. ) has been investigated. The authors report here that the mutant accumulated less Na + in the root and leaf than the wild type in response to NaCI stress. This difference in Na + accumulation in leaf was more significant than that in the root. The mutant kept a lower net accumulation rate of Na + than that in the wild type during the stress. K+ content in the leaves and roots of beth species reduced severely when exposed to NaC1, but the contents in the leaf and root of the mutant was higher than those of the wild type. The Na + dis- tribution in the seedlings of the mutant and the wild type was significantly different. When exposed to salt stress for 96 h, the quantity of the accumulated Na + in root was 44.3 % of the total Na + per seedling of the mutant, whereas it was 24.3% in the wild type, which was likely resulted from the reduction of Na+ transfer from roots to shoots in the mutant.     

Stomatal control in tomato with ABA-deficient roots: response of grafted plants to soil drying

The hypothesis that ABA produced by roots in drying soil is responsible for stomatal closure was tested with grafted plants constructed from the ABA-deficient tomato mutants, sitiens and flacca and their near-isogenic wild-type parent. Three types of experiments were conducted. In the first type, reciprocal grafts were made between the wild type and sitiens or flacca. Stomatal conductance accorded with the genotype of the shoot, not the root. Stomates closed in all of the grafted plants in response to soil drying, regardless of the root genotype, i.e. regardless of the ability of the roots to produce ABA. In the second type of experiment, wild-type shoots were grafted onto a split-root system consisting of one wild-type root grafted to one mutant (flacca or sitiens) root. Water was withheld from one root system, while the other was watered well so that the shoots did not experience any decline in water potential or loss of turgor. Stomates closed to a similar extent when water was withheld from the mutant roots or the wild-type roots. In the third type of experiment, grafted plants with wild-type shoots and either wild-type or sitiens roots were established in pots that could be placed inside a pressure chamber, and the pressure increased as the soil dried so that the shoots remained fully turgid throughout. Stomates closed as the soil dried, regardless of whether the roots were wild type or sitiens. These experiments demonstrate that stomatal closure in response to soil drying can occur in the absence of leaf water deficit, and does not require ABA production by roots. A chemical signal from roots leading to a change in apoplastic ABA levels in leaves may be responsible for the stomatal closure.     

Wild-type levels of abscisic Acid are not required for heat shock protein accumulation in tomato

Levels of endogenous abscisic acid (ABA) in wild type were not required for the synthesis of heat shock proteins in detached leaves of tomato (Lycopersicon esculentum Mill., cv Ailsa Craig). Heat-induced alterations in gene expression were the same in the ABA-deficient mutant of tomato, flacca, and the wild type. Heat tolerance of the mutant was marginally less that the wild type, and in contrast, ABA applications significantly reduced the heat tolerance of wild-type leaves. It was concluded that elevated levels of endogenous ABA are not involved in the tomato heat shock response.     

The Ascorbate-deficient vtc-1 Arabidopsis Mutant Shows Altered ABA Accumulation in Leaves and Chloroplasts

Abscisic acid (ABA) accumulation has been analyzed in irrigated and water-stressed wild-type and the vtc-1 mutant of Arabidopsis thaliana, which shows an ascorbate deficiency in leaves of approximately 60%. The amounts of ABA increased progressively up to 2.3-fold in water-stressed wild-type plants, whereas levels were kept at low levels in the irrigated plants. In contrast, initial increases followed by a sharp decrease of abscisic acid levels were observed in water-stressed vtc-1 mutants. Furthermore, the levels of this phytohormone increased up to fivefold in irrigated mutants. This differential accumulation of ABA in the mutant strongly correlated with the ascorbate redox state, but not with ascorbate levels. Changes in ABA levels in leaves paralleled those of chloroplasts. Immunolocalization studies showed a differential ABA accumulation in chloroplasts of vtc-1 mutants, which displayed the highest ABA labeling in irrigated plants. Our results indicate an altered pattern of ABA accumulation in the vtc-1 mutant compared to the wild type, under both irrigated conditions and water-stress conditions, which is strongly dependent on the ascorbate redox state.     

Effect of salinity and abscisic acid on accumulation of glycinebetaine and betaine aldehyde dehydrogenase mRNA in Sorghum leaves (Sorghum bicolor)

The effects of salt stress and abscisic acid (ABA) on the expression of betaine aldehyde dehydrogenase (BADH) were determined in sorghum (Sorghum bicolor L.) plants. BADH mRNA expression was induced by salinity, and the timing coincided with the observed glycinebetaine (betaine) accumulation. The leaf water potential in the leaves of the sorghum plants was significantly affected by salinity. In response to salinity, betaine, ABA, Na and Cl accumulations increased 6-, 16-, 90-, and 3-fold, respectively. In the leaf disks from unsalinized plants incubated on NaCl, or ABA solution, the BADH mRNA level was lower than in the ABA-treated disks. Exogenous application of the ABA biosynthetic inhibitor fluridone to the NaCl-treated disks reduced the ABA accumulation and BADH mRNA levels compared with NaCl-treated leaves. The results indicate that the salt-induced accumulation of betaine and BADH mRNA coincides with the presence of ABA.     

Abscisic Acid accumulation is not required for proline accumulation in wilted leaves

Leaves from dark-grown barley (Hordeum vulgare L. var Larker) seedlings grown in the presence and absence of fluridone were used to determine whether or not abscisic acid (ABA) accumulation was necessary for proline to accumulate in wilted tissue. Wilted tissue (polyethylene glycol-treated) leaves from fluridone-grown seedlings did not accumulate ABA but did accumulate proline at a rate that was not different from the non-fluridone-treated leaves. Thus ABA accumulation is not required for wilting-induced proline accumulation in barley leaves. Proline accumulation in wilted leaves from the wilty tomato (Lycopersicon esculentum) mutant, flacca, was compared to that in the wild type, Rheinlands Ruhm. Proline accumulated in wilted leaves from flacca. The rate of accumulation was faster in flacca compared to the rate in the wild type because the wilty mutant wilted faster. ABA accumulated in wilted leaves from the wild type but not in the wilty mutant. This result is a further confirmation that ABA accumulation is not required for wilting-induced proline accumulation. These results are significant in that proline accumulation in barley leaves can be induced independently by any one of three treatments: wilting, ABA, or salt.     

ABA and polyamines act independently in primary leaves of cold-stressed tomato (Lycopersicon esculentum)

The effects of ABA and putrescine, a polyamine, on cold-induced membrane leakage were investigated using primary leaves of wild-type and an ABA-deficient mutant, flacca , of tomato ( Lycopersicon esculentum Mill.). The amount of chilling-induced electrolyte leakage from flacca leaves was much higher than that from the wild-type leaves. When applied exogenously ABA reduced cold-induced electrolyte leakage from leaves of both wild-type and the flacca mutant. However, the cold-induced electrolyte leakage from flacca leaves was not as pronounced as in the wild-type indicating that ABA is an important mediator in response to cold stress in the leaves. Putrescine reduced cold-induced electrolyte leakage from both wild-type and flacca leaves. Synthesis of putrescine in the leaves was increased by cold treatment. DFMO, a biosynthetic inhibitor of the polyamine, increased electrolyte leakage from cold-treated leaves, and exogenously applied putrescine decreased the enhanced leakage to the control level. Therefore, this polyamine is thought also to be involved in the response to cold stress of tomato leaves. Both ABA and putrescine were protective against cold stress, but exogenously applied ABA decreased the endogenous level of putrescine in the leaves. Furthermore, the DMFO-increased electrolyte leakage in cold-stressed leaves was completely abolished by the application of ABA. These results suggest that ABA is a major regulator in the response to cold stress in tomato leaves and that it does not exert its role via putrescine in the response to cold stress.     

ABA content in shoots and roots of pea mutants af and tl as related to their growth and morphogenesis

The comparative study of shoot and root growth was carried out, and the level of ABA therein determined in the mutant af and tl and wild-type isogenic lines of pea. The recessive af mutation transformed the leaflets into tendrils, and the tl mutation transformed the tendrils into leaflets. These mutations did not affect the length and number of internodes. In all plants, the level of ABA in the leaves was 3–10 times greater than in the roots, and in the course of vegetative growth it rose in both organs. An increase in the shoot area of tl mutant did not change the dry weight of underground and above-ground parts; therefore, the ratio shoot/root in the mutant was identical to that in the wild-type plants. The maintenance of shoot dry weight in the tl mutant at the level of wild-type plant while its area considerably increased was accounted for by a decrease in the thickness of the leaflet and stipule blades. The level of ABA in the stipules of mutant plants was greater than in the wild-type plants. A decrease in the shoot area in the af mutant brought about a decline in its dry weight; however, the ratio root/shoot was maintained at the wild-type level due to a reduced accumulation of dry weight by the root. The level of ABA in the roots of the af mutant was twice greater than in the leafy forms. ABA was assumed to participate in the control over the root growth exerted by the shoot. The absence of leaflets in the af plants was partially compensated for by expanding stipules. The level of ABA therein was three times higher than in the plants of wild type and comparable with the level in the leaflets of the tl mutant and in the wild-type plants. The role of ABA in the growth and final size of leaf blades is discussed.