Comparative studies on the Arabidopsis aldehyde oxidase (AAO) gene family revealed a major role of AAO3 in ABA biosynthesis in seeds

The Arabidopsis aldehyde oxidase 3 (AAO3) gene encodes an enzyme that catalyzes the final step of ABA biosynthesis. AAO3 has been shown to be the major AAO involved in ABA biosynthesis in leaves under stress conditions. On the other hand, less severe phenotypes of the aao3 seeds suggested that other AAO(s) might also be involved in ABA biosynthesis in seeds. Among four AAOs (AAO1-AAO4), AAO1 and AAO4 were the AAO expressed most abundantly in dry seeds and developing siliques, respectively. Unlike aao3, single loss-of-function mutants for AAO1 and AAO4 (aao1 and aao4), failed to show significant changes in endogenous ABA levels in seeds when compared with wild type. While aao3 seed germination was resistant to the gibberellin biosynthesis inhibitor, uniconazole, aao1 and aao4 showed no resistance and were similar to wild type. These results indicate that AAO3, but not AAO1 or AAO4, plays an important role in ABA biosynthesis in seeds. Mutations of AAO1 or AAO4 in the aao3 mutant background enhanced ABA deficiency in seeds, demonstrating that both gene products contribute partially to ABA biosynthesis in the aao3 mutant background. However, considering the enzymatic characters of AAO1 and AAO4, their involvement in ABA biosynthesis in wild-type seeds may be negligible. We have concluded that AAO3 is the AAO that plays a major role in ABA biosynthesis in Arabidopsis seeds as well as in leaves.     

Phytohormones Related to Host Plant Manipulation by a Gall-Inducing Leafhopper

The maize orange leafhopper Cicadulina bipunctata (Hemiptera: Cicadellidae) induces galls characterized by growth stunting and severe swelling of leaf veins on various plants of Poaceae. Previous studies revealed that galls are induced not on feeding site but on distant, newly extended leaves during the feeding, and strongly suggested that some chemicals injected by the leafhopper affect at the leaf primordia. To approach the mechanism underlying gall induction by C. bipunctata, we examined physiological response of plants to feeding by the leafhopper. We performed high-throughput and comprehensive plant hormone analyses using LC-ESI-MS/MS. Galled maize leaves contained higher contents of abscisic acid (ABA) and trans-Zeatin (tZ) and lower contents of gibberellins (GA₁ and GA₄) than ungalled maize leaves. Leafhopper treatment significantly increased ABA and tZ contents and decreased GA₁ and GA₄ contents in extending leaves. After the removal of leafhoppers, contents of tZ and gibberellins in extending leaves soon became similar to the control values. ABA content was gradually decreased after the removal of leafhoppers. Such hormonal changes were not observed in leafhopper treatment on leaves of resistant maize variety. Water contents of galled leaves were significantly lower than control leaves, suggesting water stress of galled leaves and possible reason of the increase in ABA content. These results imply that ABA, tZ, and gibberellins are related to gall induction by the leafhopper on susceptible variety of maize.     

Movement of Abscisic Acid into the Apoplast in Response to Water Stress in Xanthium strumarium L

The effect of water stress on the redistribution of abcisic acid (ABA) in mature leaves of Xanthium strumarium L. was investigated using a pressure dehydration technique. In both turgid and stressed leaves, the ABA in the xylem exudate, the `apoplastic' ABA, increased before `bulk leaf' stress-induced ABA accumulation began. In the initially turgid leaves, the ABA level remained constant in both the apoplast and the leaf as a whole until wilting symptoms appeared. Following turgor loss, sufficient quantities of ABA moved into the apoplast to stimulate stomatal closure. Thus, the initial increase of apoplastic ABA may be relevant to the rapid stomatal closure seen in stressed leaves before their bulk leaf ABA levels rise.

Following recovery from water stress, elevated levels of ABA remained in the apoplast after the bulk leaf contents had returned to their prestress values. This apoplastic ABA may retard stomatal reopening during the initial recovery period.

     

花椰菜幼苗抗黑腐病的生理机制研究

以对黑腐病抗性不同的2个花椰菜(B rassica oleracea var.botry tis)品种为材料,研究了花椰菜苗期抵抗黑腐病的生理机制。结果表明:接种7 d后,黑腐病菌(X anthom onas camp estris pv.C amp estris)的侵入导致花椰菜幼苗干物质积累下降,抗病品种‘雪峰’的下降幅度明显低于感病品种‘2003X-106’,这与接种3 d后雪峰的净光合速率、气孔导度、蒸腾速率和叶绿素含量等下降较慢有关。病原菌侵染后,2个品种叶片的可溶性糖含量均有增加,但是抗病品种‘雪峰’的增幅较感病品种‘2003X-106’低;‘雪峰’叶片的可溶性蛋白质含量在接种后逐渐降低,而‘2003X-106’却逐渐增加;接种后,‘雪峰’叶片的IAA和M e-JA含量均上升,ABA则显著降低,而‘2003X-106’的IAA含量降低,ABA显著增加,M e-JA则在接种后的不同时期有增有减。     

干旱胁迫下不同抗旱性小麦叶片内源激素含量的变化与抗旱力强弱的关系

以抗旱性强的小麦品种昌乐5号和抗旱性弱的鲁麦5号的幼苗为材料,研究了随着干旱胁迫的加剧,小麦叶片相对含水量、气孔导度和内源激素水平的变化．结果表明,叶片的气孔导度和细胞激动素与脱落酸含量的比值（CTKs/ABA）呈较强的线性正相关关系而与叶片RWC的相关关系较弱,说明气孔导度受CTKs/ABA调控,而不是受叶片RWC的调控．在不同强度的干旱胁迫下我们可以利用CTKs/ABA的变化判断品种抗旱性大小：①抗旱力强的小麦品种叶片的平均CTKs/ABA值较高,而弱者较低,说明强者内源促进型激素含量相对较高,从而在干旱胁迫下保持较高的生活力．②抗旱力强的小麦品种叶片CTKs/ABA与干旱处理天数呈二次负相关关系,而弱者是线性负相关关系,说明强者具有较强的抗逆缓冲能力．③抗旱力强的小麦品种成熟叶（第二叶）CTKs／ABA下降快,新叶（第三叶）下降慢,而弱者反之,说明抗旱性强者可能存在着较强的从成熟叶向新叶的物质运输从而具有自我保护性调节机制．     

Dormancy and Germination of Abscisic Acid-Deficient Tomato Seeds : Studies with the sitiens Mutant

The role of abscisic acid (ABA) in the dormancy induction of tomato (Lycopersicon esculentum) seeds was studied by comparison of the germination behavior of the ABA-deficient sitiens mutant with that of the isogenic wild-type genotype. Freshly harvested mutant seeds, in contrast to wild-type seeds, always readily germinate and even exhibit viviparous germination in overripe fruits. Crosses between mutant and wild-type and self-pollination of heterozygous plants show that in particular the ABA fraction of embryo and endosperm is decisive for the induction of dormancy. After-ripened wild-type seeds fully germinate in water but are more sensitive toward osmotic inhibition than mutant seeds. Germination of both wild-type and mutant seeds is equally sensitive toward inhibition by exogenous ABA. ABA content of mature wild-type seeds is about 10-fold the level found in mutant seeds. Nevertheless, it is argued that the differences in dormancy between the seeds of both genotypes are not a result of actual ABA levels in the mature seeds or fruits but a result of differences in ABA levels during seed development. It is hypothesized that the high levels of ABA that occur during seed development in wild-type seeds induce an inhibition of cell elongation of the radicle that can still be observed after long periods of dry storage.     

自然干旱胁迫下长春花叶片和根部的激素和可溶性糖代谢变化

植物在离开生长环境较短时间内（1~6 h）会导致缓慢的表面水分散失,引起自然的干旱胁迫。本文以耐旱植物长春花（Catharanthus roseus）为材料,研究其在离土干旱胁迫中的脱落酸（ABA）及可溶性糖含量变化。结果表明,长春花根部ABA含量在正常条件下低于叶片中的含量,干旱胁迫促进了ABA在根部的积累,6 h时增加至最高值。蔗糖酸性转化酶活性可能受到ABA的诱导在胁迫6 h时最高,比对照高出30%左右。长春花叶片中总可溶性糖含量在对照条件下非常稳定,但在干旱胁迫过程中,其随着时间的延长呈现线性增加的趋势（r²=0.964）,蔗糖和已糖含量在胁迫过程中也呈增加的趋势,可能发挥着渗透调控节功能。     

Dynamic distribution and the role of abscisic acid during seed development of a lady’s slipper orchid,Cypripedium formosanum

Background and Aims Although abscisic acid (ABA) is commonly recognized as a primary cause of seed dormancy, there is a lack of information on the role of ABA during orchid seed development. In order to address this issue, the localization and quantification of ABA were determined in developing seeds of Cypripedium formosanum.Methods The endogenous ABA profile of seeds was measured by enzyme-linked immunosorbent assay (ELISA). Temporal and spatial distributions of ABA in developing seeds were visualized by immunohistochemical staining with monoclonal ABA antibodies. Fluoridone was applied to test the causal relationship between ABA content and seed germinability.Key Results ABA content was low at the proembryo stage, then increased rapidly from 120 to 150 days after pollination (DAP), accompanied by a progressive decrease in water content and seed germination. Immunofluorescence signals indicated an increase in fluorescence over time from the proembryo stage to seed maturation. From immunogold labelling, gold particles could be seen within the cytoplasm of embryo-proper cells during the early stages of seed development. As seeds approached maturity, increased localization of gold particles was observed in the periplasmic space, the plasmalemma between embryo-proper cells, the surface wall of the embryo proper, and the inner walls of inner seed-coat cells. At maturity, gold particles were found mainly in the apoplast, such as the surface wall of the embryo proper, and the shrivelled inner and outer seed coats. Injection of fluoridone into capsules resulted in enhanced germination of mature seeds.Conclusions The results indicate that ABA is the key inhibitor of germination in C. formosanum. The distinct accumulation pattern of ABA suggests that it is synthesized in the cytosol of embryo cells during the early stages of seed development, and then exported to the apoplastic region of the cells for subsequent regulatory processes as seeds approach maturity.     

Relationship between Stress-Induced ABA and Proline Accumulations and ABA-Induced Proline Accumulation in Excised Barley Leaves

When excised second leaves from 2-week-old barley (Hordeum vulgare var Larker) plants were incubated in a wilted condition, abscisic acid (ABA) levels increased to 0.6 nanomole per gram fresh weight at 4 hours then declined to about 0.3 nanomole per gram fresh weight and remained at that level until rehydrated. Proline levels began to increase at about 4 hours and continued to increase as long as the ABA levels were 0.3 nanomole per gram fresh weight or greater. Upon rehydration, proline levels declined when the ABA levels fell below 0.3 nanomole per gram fresh weight.

Proline accumulation was induced in turgid barley leaves by ABA addition. When the amount of ABA added to leaves was varied, it was observed that a level of 0.3 nanomole ABA per gram fresh weight for a period of about 2 hours was required before proline accumulation was induced. However, the rate of proline accumulation was slower in ABA-treated leaves than in wilted leaves at comparable ABA levels. Thus, the threshold level of ABA for proline accumulation appeared to be similar for wilted leaves where ABA increased endogenously and for turgid leaves where ABA was added exogenously. However, the rate of proline accumulation was more dependent on ABA levels in turgid leaves to which ABA was added exogenously than in wilted leaves.

Salt-induced proline accumulation was not preceded by increases in ABA levels comparable to those observed in wilted leaves. Levels of less than 0.2 nanomole ABA per gram fresh weight were measured 1 hour after exposure to salt and they declined rapidly to the control level by 3 hours. Proline accumulation commenced at about 9 hours. Thus, ABA accumulation did not appear to be involved in salt-induced proline accumulation.

     

In Vivo Inhibition of Seed Development and Reserve Protein Accumulation in Recombinants of Abscisic Acid Biosynthesis and Responsiveness Mutants in Arabidopsis thaliana

In Arabidopsis thaliana, seed development in recombinants of the ABA-deficient aba mutant with the ABA response mutants abi1 or abi3 is compared to wild type and the monogenic parents. Aberrant seed development occurred in the aba,abi3 recombinant and was normal in aba,abi1, abi3 and aba,abi1 seeds. Embryos of the recombinant aba,abi3 seeds maintained the green color until maturity, the seeds kept a high water content, did not form the late abundant 2S and 12S storage proteins, were desiccation intolerant, and often showed viviparous germination. Application of ABA, and particularly of an ABA analog, to the roots of plants during seed development partially alleviated the aberrant phenotype. Seeds of aba,abi3 were normal when they developed on a mother plant heterozygous for Aba. In contrast to seed development, the induction of dormancy was blocked in all monogenic mutants and recombinants. Dormancy was only induced by embryonic ABA; it could not be increased by maternal ABA or ABA applied to the mother plant. It is concluded that endogenous ABA has at least two different effects in developing seeds. The nature of these responses and of the ABA response system is discussed.     

Oryzalin-Induced Changes in Water Status and Cytoskeleton Proteins of Winter Wheat Seedlings upon Cold Acclimation and ABA Treatment

The effect of oryzalin (a specific inhibitor of tubulin polymerization in plant cells) on water retention by the leaves and roots of winter wheat (Triticum aestivum L.) seedlings was studied. The cultivars differing in their frost resistance were compared after their acclimation to low temperature (3°C for 3 or 7 days) and after treatment with ABA. In control untreated plants, oryzalin reduced the water-retaining capacity (WRC) of leaves and roots. Both hardening and ABA lowered the effect of the inhibitor on WRC in leaves, whereas their effects on water retention by roots were opposite, i.e., hardening weakened and ABA intensified the effect of oryzalin. Oryzalin-induced reduction of WRC decreased in the following sequence of cultivars: weakly frost resistant moderately frost resistant highly frost resistant. It was more pronounced in the leaves than in the roots, the latter being characterized by the lower WRC and lower frost resistance. After three-day-long hardening of plants, an additive effect of hypothermia and ABA on oryzalin-induced decrease in WRC of leaves and the lack of such effect in the roots were observed. The immunochemical analysis of the composition and content of cytoskeletal proteins with Western blotting showed that in the leaves the actin/tubulin ratio was higher than in the roots. The treatment of nonacclimated plants with ABA lowered the content of - and -tubulins and actin in roots but did not affect the level of actin in leaves. Hardening negated the effects of ABA on cytoskeletal proteins. Oryzalin produced the greatest inhibitory effect on WRC and an increase in frost resistance in ABA-treated plants in the experiments with leaves of the weakly frost resistant cultivar before and after hardening. Organ- and cultivar-specific and ABA-mediated dependence of WRC on cytoskeletal proteins and microtubules and microfilaments formed by them is supposed to result from their effect on the state of intracellular water and water permeability of the plasma membrane. In the course of cold acclimation of plants and upon their treatment with ABA, this dependence was more distinctly expressed in leaves than in roots, and especially in the plants of the weakly frost resistant cultivar.     

Effects of pod removal on the transport and accumulation of abscisic Acid and indole-3-acetic Acid in soybean leaves

Concentrations of abscisic acid (ABA) and indole-3-acetic acid (IAA) in the second most recently expanded trifoliolate leaf were determined during reproductive development of soybean (Glycine max [L.] Merr cv `Chippewa 64'). The concentration of ABA in leaves was constant during most of the seed filling period until the seeds began to dry. The concentration of IAA in the leaves decreased throughout development. Removal of pods 36 hours prior to sampling resulted in increased concentrations of ABA in leaves during the period of rapid pod filling but had little effect on the concentration of IAA in leaves. ABA appears to accumulate in leaves after fruit removal only when fruits represent the major sink for photosynthate.

ABA and IAA moving acropetally and basipetally in petioles of soybean were estimated using a phloem exudation technique. ABA was found to move mostly in the basipetal direction in petioles (away from laminae). IAA, primarily in the form of ester conjugate(s), was found to be moving acropetally (toward laminae) in petioles. The highest amount of IAA ester(s) was found in petiole exudate during the mid and late stages of seed filling. Removal of fruits 36 hours prior to exudation reduced the amount of IAA ester recovered in exudate, suggesting that fruits were a source of the IAA conjugate in petiole exudate.

     

Abscisic acid accumulation and cadmium tolerance in rice seedlings

Rice ( Oryza sativa L.) seeds were soaked for 18 h in distilled water in the absence (–PBZ) or presence (+PBZ, a triazole) of 100 mg l⁻¹ paclobutrazol and then air dried. These air-dried seeds were germinated in the dark and then cultivated in a Phytotron. Twelve-day-old –PBZ and +PBZ seedlings were treated or not with CdCl₂. Cd toxicity was judged by the decrease in biomass production, decrease in chlorophyll and protein content, increase in NH₄⁺ content and induction of oxidative stress. The results indicated that PBZ applied to seeds was able to protect rice seedlings from Cd toxicity. On treatment with CdCl₂, the abscisic acid (ABA) content increased in +PBZ leaves, but not in –PBZ leaves. The decrease in the transpiration rate of –PBZ seedlings by CdCl₂ was less than that of +PBZ seedlings. Exogenous application of the ABA biosynthesis inhibitor, fluridone (Flu), reduced ABA accumulation, increased the transpiration rate and Cd content, and decreased the Cd tolerance of +PBZ seedlings. The effects of Flu on the Cd toxicity, transpiration rate and Cd content were reversed by the application of ABA. It seems that the PBZ-induced Cd tolerance of rice seedlings is mediated through an accumulation of ABA.     

Dormancy of Medicago marina (L.) seed

Medicago marina (L.) is a Mediterranean species whose seeds show strong dormancy that prevents germination. We used an integrated approach of physiological analyses and proteomics to investigate the mechanisms that control M. marina dormancy/germination and that underlie stress tolerance. First, we evaluated the effects on dormancy breaking of the following treatments: mechanical scarification, freezing at −20 °C, storage for 4 months and heating at 100 °C for 1 h. Mechanical scarification and freezing were the most effective treatments in overcoming dormancy. The role of abscisic acid (ABA) in M. marina dormancy was studied by ELISA immuno-enzymatic assay. The ABA content of germinated and non-germinated mature (control) and treated seeds was determined. The level of ABA was higher in treated seeds than in control seeds; the most significant increase occurred in the heated seeds. A comparison of the ABA level in the germinated, control and treated seeds suggests that different mechanisms modulate ABA content in response to different stresses, and that a specific ABA-signalling pathway regulates germination. Proteomic analysis revealed 46 proteins differentially expressed between treated and untreated seeds; 14 of these proteins were subsequently identified by mass spectrometry. Several of the proteins identified are important factors in the stress response, and are involved in such diverse functions as lipid metabolism, protein folding and chromatin protection. Lastly, an analysis of the phosphoproteome maps showed that the function of many proteins in seeds subjected to temperature treatment is modulated through post-translational modifications.     

Regulation and manipulation of ABA biosynthesis in roots

Overexpression of 9-cis-epoxycarotenoid dioxygenase (NCED) is known to cause abscisic acid (ABA) accumulation in leaves, seeds and whole plants. Here we investigated the manipulation of ABA biosynthesis in roots. Roots from whole tomato plants that constitutively overexpress LeNCED1 had a higher ABA content than wild-type (WT) roots. This could be explained by enhanced in situ ABA biosynthesis, rather than import of ABA from the shoot, because root cultures also had higher ABA content, and because tetracycline (Tc)-induced LeNCED1 expression caused ABA accumulation in isolated tobacco roots. However, the Tc-induced expression led to greater accumulation of ABA in leaves than in roots. This demonstrates for the first time that NCED is rate-limiting in root tissues, but suggests that other steps were also restrictive to pathway flux, more so in roots than in leaves. Dehydration and NCED overexpression acted synergistically in enhancing ABA accumulation in tomato root cultures. One explanation is that xanthophyll synthesis was increased during root dehydration, and, in support of this, dehydration treatments increased beta-carotene hydroxylase mRNA levels. Whole plants overexpressing LeNCED1 exhibited greatly reduced stomatal conductance and grafting experiments from this study demonstrated that this was predominantly due to increased ABA biosynthesis in leaves rather than in roots. Genetic manipulation of both xanthophyll supply and epoxycarotenoid cleavage may be needed to enhance root ABA biosynthesis sufficiently to signal stomatal closure in the shoot.     

ABI4 Regulates Primary Seed Dormancy by Regulating the Biogenesis of Abscisic Acid and Gibberellins in Arabidopsis

Seed dormancy is an important economic trait for agricultural production. Abscisic acid (ABA) and Gibberellins (GA) are the primary factors that regulate the transition from dormancy to germination, and they regulate this process antagonistically. The detailed regulatory mechanism involving crosstalk between ABA and GA, which underlies seed dormancy, requires further elucidation. Here, we report that ABI4 positively regulates primary seed dormancy, while negatively regulating cotyledon greening, by mediating the biogenesis of ABA and GA. Seeds of the Arabidopsis abi4 mutant that were subjected to short-term storage (one or two weeks) germinated significantly more quickly than Wild-Type (WT), and abi4 cotyledons greened markedly more quickly than WT, while the rates of germination and greening were comparable when the seeds were subjected to longer-term storage (six months). The ABA content of dry abi4 seeds was remarkably lower than that of WT, but the amounts were comparable after stratification. Consistently, the GA level of abi4 seeds was increased compared to WT. Further analysis showed that abi4 was resistant to treatment with paclobutrazol (PAC), a GA biosynthesis inhibitor, during germination, while OE-ABI4 was sensitive to PAC, and exogenous GA rescued the delayed germination phenotype of OE-ABI4. Analysis by qRT-PCR showed that the expression of genes involved in ABA and GA metabolism in dry and germinating seeds corresponded to hormonal measurements. Moreover, chromatin immunoprecipitation qPCR (ChIP-qPCR) and transient expression analysis showed that ABI4 repressed CYP707A1 and CYP707A2 expression by directly binding to those promoters, and the ABI4 binding elements are essential for this repression. Accordingly, further genetic analysis showed that abi4 recovered the delayed germination phenotype of cyp707a1 and cyp707a2 and further, rescued the non-germinating phenotype of ga1-t. Taken together, this study suggests that ABI4 is a key factor that regulates primary seed dormancy by mediating the balance between ABA and GA biogenesis.     

Overexpression of wheat <Emphasis Type="Italic">TaNCED</Emphasis> gene in <Emphasis Type="Italic">Arabidopsis</Emphasis> enhances tolerance to drought stress and delays seed germination

Abscisic acid (ABA) regulates various plant physiological processes, especially participates in the plant responses to harsh environments. The 9-cis-epoxycarotenoid dioxygenase (NCED) is a key enzyme in ABA biosynthesis pathway. Here, a TaNCED with an 1 887-bp open reading frame was cloned from wheat, which encodes a peptide of 628 amino acids. A chloroplast transit peptide sequence was found at the N-terminus of the TaNCED protein. Multiple sequence alignments indicate that the TaNCED protein shared high similarities with other NCEDs from different species. Real-time quantitative PCR analysis shows that expression of TaNCED was strongly up-regulated by treatments with ABA, polyethylene glycol, and drought stress, and it was down-regulated during germination of the wheat seeds. Ectopic overexpression of the TaNCED gene in Arabidopsis resulted in an increase of endogenous ABA and free proline content. A lower water loss rate and stomatal conductance of leaves were found in the transgenic plants in comparison with the wild type. Subsequently, the transgenic plants displayed an enhanced tolerance to drought stress but delayed seed germination. These data provide evidence that the TaNCED might play a primary role in regulation of ABA content during water stress and seed dormancy.     

Abscisic Acid Metabolism in Relation to Water Stress and Leaf Age in Xanthium strumarium

Intact plants of Xanthium strumarium L. were subjected to a water stress-recovery cycle. As the stress took effect, leaf growth ceased and stomatal resistance increased. The mature leaves then wilted, followed by the half expanded ones. Water, solute, and pressure potentials fell steadily in all leaves during the rest of the stress period. After 3 days, the young leaves lost turgor and the plants were rewatered. All the leaves rapidly regained turgor and the younger ones recommenced elongation. Stomatal resistance declined, but several days elapsed before pre-stress values were attained.

Abscisic acid (ABA) and phaseic acid (PA) levels rose in all the leaves after the mature ones wilted. ABA-glucose ester (ABA-GE) levels increased to a lesser extent, and the young leaves contained little of this conjugate. PA leveled off in the older leaves during the last 24 hours of stress, and ABA levels declined slightly. The young leaves accumulated ABA and PA throughout the stress period and during the 14-hour period immediately following rewatering. The ABA and PA contents, expressed per unit dry weight, were highest in the young leaves. Upon rewatering, large quantities of PA appeared in the mature leaves as ABA levels fell to the pre-stress level within 14 hours. In the half expanded and young leaves, it took several days to reach pre-stress ABA values. ABA-GE synthesis ceased in the mature leaves, once the stress was relieved, but continued in the half expanded and young leaves for 2 days.

Mature leaves, when detached and stressed, accumulated an amount of ABA similar to that in leaves on the intact plant. In contrast, detached and stressed young leaves produced little ABA. Detached mature leaves, and to a lesser extent the half expanded ones, rapidly catabolized ABA to PA and ABA-GE, but the young leaves did not. Studies with radioactive (±)-ABA indicated that in young leaves the conversion of ABA to PA took place at a much lower rate than in mature ones. Leaves of all ages rapidly conjugated PA to PA-glucose ester. Furthermore, when half expanded leaves were stressed on the intact plant, their rate of ABA catabolism was enhanced, an effect not observed in the young leaves.

In conclusion, young leaves on intact Xanthium plants produce little stress-induced ABA themselves, but due to import and a low rate of catabolism accumulate more ABA and PA than mature leaves.

     

A monoclonal antibody to (S)-abscisic acid: its characterisation and use in a radioimmunoassay for measuring abscisic acid in crude extracts of cereal and lupin leaves

A monoclonal antibody produced to abscisic acid (ABA) has been characterised and the development of a radioimmunoassay (RIA) for ABA using the antibody is described. The antibody had a high selectivity for the free acid of (S)-cis, trans-ABA. Using the antibody, ABA could be assayed reliably in the RIA over a range from 100 to 4000 pg (0.4 to 15 pmol) ABA per assay vial. As methanol and acetone affected ABA-antibody binding, water was used to extract ABA from leaves. Water was as effective as aqueous methanol and acetone in extracting the ABA present. Crude aqueous extracts of wheat, maize and lupin leaves could be analysed without serious interference from other immunoreactive material. This was shown by measuring the distribution of immunoreactivity in crude extracts separated by thin-layer chromatography (TLC) and high-performance liquid chromatography (HPLC), or by comparing the assay with physicochemical methods of analysis. Analysis of crude extracts by RIA and either, after TLC purification, by gas chromatography using an electron-capture detector or, after HPLC purification, by combined gas chromatography-mass spectrometry (GC-MS) gave very similar ABA concentrations in the initial leaf samples. However, RIA analysis of crude aqueous extracts of pea seeds resulted in considerable overestimation of the amount of ABA present. Determinations of ABA content by GC-MS and RIA were similar after pea seed extracts had been purified by HPLC. Although the RIA could not be used to analyse ABA in crude extracts of pea seeds, it is likely that crude extracts of leaves of several other species may be assayed successfully.Abbreviations ABA abscisic acid - DW dry weight - FW fresh weight - GC-ECD gas chromatography using an electron capture detector - GC-MS combined gas chromatographymass spectrometry - HPLC high-performance liquid chromatography - McAb monoclonal antibody - PVP soluble polyvinylpyrrolidone - RIA radioimmunoassay - TLC thin-layer chromatography     

Differences in growth and physiological traits of Populus cathayana populations as affected by enhanced UV-B radiation and exogenous ABA

During one growing season, the effects of enhanced ultraviolet-B (UV-B) radiation, exogenous abscisic acid (ABA) and their combination on biomass accumulation, gas exchange, endogenous ABA, the concentration of UV-absorbing compounds, antioxidant system and on the carbon (C) and nitrogen (N) content and C/N ratio were investigated in two contrasting Populus cathayana Rehd. populations, originating from high and low altitudes in south-west China. Exogenous ABA was sprayed to the leaves, and enhanced UV-B treatments were applied using a square-wave system to expose the seedlings to ambient (1×) or twice ambient (2×) doses of biologically effective UV-B radiation (UV-B_BE). Enhanced UV-B radiation significantly decreased height, basal diameter, total leaf area, total biomass, net CO₂ assimilation rate (A), stomatal conductance (g_s), transpiration rate (E) and carbon (C) content in leaves, and significantly increased the activities of superoxide dismutase (SOD) and guaiacol peroxidase (GPx), and the contents of hydrogen peroxide (H₂O₂) and malonaldehyde (MDA), as well as the accumulation of UV-absorbing compounds and endogenous ABA concentrations among different organs in both populations. In contrast, exogenous ABA induced a significant decrease in A and significant increases in the activities of SOD and GPx, in the content of H₂O₂ and MDA, and in the endogenous ABA concentrations. Compared with the low altitude population, the high altitude population was more tolerant to enhanced UV-B and exogenous ABA. Significant interactions between UV-B and ABA were observed in A, E, and in the activities of SOD and GPx, as well as in endogenous ABA in the leaves and roots of both populations. Across all treatments, the C and N contents of leaves were strongly correlated with their contents in stems and roots. Additionally, the N content of leaves and stems were significantly correlated with the C content of stems.