Biochemical changes during seed development in <Emphasis Type="Italic">Pinus taeda</Emphasis> L.

Amino acids, polyamines, 3-indoleacetic acid (IAA), abscisic acid (ABA), buffer-soluble protein and starch contents and dry matter accumulation were analyzed in megagametophytes containing developing embryos during seed development in Pinus taeda. The highest total amino acids and polyamine contents occurred at the cotyledonary stage, followed by a significant decrease in the mature seed. Free polyamines exhibited higher levels than conjugated ones, with putrescine being the predominant type until the cotyledonary stage, and spermidine at the mature seed stage. IAA content increased continually from the globular stage reaching the maximum at the cotyledonary stage, followed by a decrease in the mature seed. The highest ABA level occurred at the globular stage, followed by a continuous reduction until stabilization at the pre-cotyledonary stage. Buffer-soluble protein and starch contents, and dry matter increased progressively during development, reaching their maximum values at the mature stage.     

Free Amino Acid, Protein and Water Content Changes Associated with Seed Development in Araucaria angustifolia

The free amino acid, protein, water and dry matter contents were determined during the seed development of Araucaria angustifolia. Soluble and insoluble proteins in the mature seed represent 4.2 % of the fresh matter. The embryonic axis stored the greatest amount of soluble proteins, while cotyledons both with the embryonic axis showed the largest quantities of insoluble proteins in the mature seed. The greatest concentration of free amino acids was detected during the stage when cotyledons start to develop. Glutamic acid, aspartic acid, alanine and serine were predominant in the whole seed while arginine, lysine and γ-aminobutyric acid were present in great amounts only in cotyledons and embryonic axis. Although megagametophyte was important as a source of free amino acids, it was not the major protein storage organ in the mature seed. In the embryogenetic process, the rise of cotyledons is closely related to physiological and biochemical changes. This revised version was published online in July 2006 with corrections to the Cover Date.     

Effect of exogenous ABA on embryo maturation and quantification of endogenous levels of ABA and IAA in <Emphasis Type="Italic">Quercus suber</Emphasis> somatic embryos

Knowledge of the relationship between indole-3-acetic acid (IAA) and abscisic acid (ABA) is relevant to control the development and the maturation of cork oak (Quercus suber L.) somatic embryos. The addition of 1 M ABA to the culture medium significantly promoted somatic embryo maturation and increased both fresh and dry matter without affecting the relative water content. This effect was parallel to the pattern of variation observed in the endogenous ABA level, which increased from the immature to the mature stage. Endogenous ABA content during the occurrence of secondary embryogenesis was similar to that of the immature stage, showing that embryos with lower ABA levels produced secondary embryos. In contrast, IAA showed the highest concentration during early embryo development and decreased afterwards. Only in somatic embryos subjected to 1-week desiccation followed by stratification at 4 °C for 2 weeks, was a moderate increment of endogenous IAA content observed. IAA and ABA showed opposite levels during the development and maturation of cork oak somatic embryos and characterised specific stages of the embryonic development.     

Seed dormancy in Acer: The role of abscisic acid in the regulation of seed development in Acer platanoides L.

Germinability of isolated embryos from developing fruits of Acer platanoides was high at the earliest developmental stage assessed (90 dpa), but fell subsequently and at seed maturity was very low. These observations showed an inverse correlation with changes in endogenous free abscisic acid (ABA) levels in the embryo, which were low during early ontogeny, but reached maximum levels late in development (150–160 dpa). These observations suggest the possibility that dormancy may be induced during development as a result of ABA accumulation in the embryo, an argument strengthened by the obvious inhibitory effect of added ABA on the germinability of isolated embryos. The cotyledons appear to exert an inhibitory influence on embryo germinability that may result from their free ABA content although the embryonic axis itself possesses an innate dormancy that may reflect its own free ABA content. The increased germinability of isolated embryos resulting form added kinetin serves only to emphasise the complexity of the system and the dangers of simplistic interpretation.The correlation between germinability and ABA content is not complete, however, since much of the reduction in germinability had occurred before any appreciable increase in free ABA levels in the embryo was observed. Indeed the failure of the intact seed to respond to endogenous changes in embryonal ABA levels suggests that even though free ABA in the embryo may influence embryo germinability, it has little effect in the intact seed, where the presence of an intact testa may be a more important factor.The absence of a desiccation phase in the embryo during the late stages of development suggests that the large increases in endogenous free ABA did not cause dormancy by inhibiting water uptake, nor did they result from water stress in the embryonal tissues.     

Embryology in Norway spruce (Picea abies). An analysis of the composition of seed storage proteins and deposition of storage reserves during seed development and somatic embryogenesis

Seed cones were collected from open-pollinated trees of Norway spruce ( Picea abies ) in a seed orchard from pollination until maturation of the seeds. Immature embryos were isolated for embryogenic tissue cultures that were maintained either on solidified medium or as liquid cultures. By transferring young somatic embryos to medium containing 90 m M sucrose and 7. 6 μ M ABA growth continued to mature embryos that accumulated storage reserves in both the hypocotyl-shoot axis and the cotyledons. Both zygotic and somatic embryos at different developmental stages were processed for microscopy as were the megagametophytes. Total protein was extracted from the seed material at intervals during development and analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. These analyses revealed that storage protein started to accumulate in the megagametophytes at the time when embryos were growing into the gametophytic tissue, while it occurred a few weeks later in the embryos at rapid embryo growth and organ differentiation. Lipid bodies also became abundant in the mature plant material. Although plastids with prominent starch grains were very frequent in both megagametophytes and embryos during development they were not observed in the desiccated tissues. Zygotic and somatic embryos displayed a similar developmental pattern.
By sequential salt-extraction and dilution two fractions highly enriched in storage protein were obtained. One fraction (G-1), requiring higher salt concentration for protein solubilization, was dominated by a protein migrating to around M, 55000–60000 when separated under non-reduced condition. After exposure to reducing agent this protein was replaced by two new ones with M, 33000 and 22000 giving evidence of disulfide bonded polypeptides. The other fraction (G-2), was dominated by polypeptides around M, 42000 and low molecular mass polypeptides (<14000).     

Maturation and germination of Phaseolus vulgaris embryonic axes in culture

In this study of embryo development in Phaseolus vulgaris L., we found that immature embryonic axes placed in culture show a growth lag before germinating. The length of this lag phase varies according to axis age at excision, but is not affected by transfer to fresh medium, alteration of sucrose concentration between 0.5 and 2%, or whether the culture medium is liquid or agar-solidified. The lag phase was shortened by both actinomycin D and cordycepin treatment, and by treatment with 10^-5 to 10^-6 M benzyladenine. The effect of abscisic acid (ABA) varied with concentration: below a certain level, it had no effect on the lag phase, but above that level it inhibited, germination. This threshold concentration was 10^-7 M for 20-d-old axes but increased to 10^-5 M by the time the axes were 32 to 34 d old. To determine whether the axes were continuing their embryonic development during the lag phase, we tested them for desiccation-tolerance and for synthesis of phaseolin, a seed storage protein which is specific for embryos of P. vulgaris. The ability to germinate after rapid desiccation was acquired by axes at 26 d past anthesis; when axes younger than this were placed in culture, they developed desiccation-tolerance during the lag phase of growth, indicating that they were continuing embryonic maturation. Phaseolin was present in isolated axes, although at lower levels than in cotyledons. It accumulated during axis development in parallel with total protein, staying at about 1% of total protein content. When isolated immature axes were pulsed with ³H-or ¹⁴C-amino acids, they incorporated label into phaseolin, shown by precipitation with anti-phaseolin antibody. Isolated axes from mature seeds, however, did not synthesize detectable amounts of phaseolin. Immature axes cultured in vitro for a period of one to several days continued synthesizing phaseolin until the day prior to visible germination. Treatment of cultured axes with ABA increased the amount of precursor amino acids incorporated into protein, but had a small or no effect on the relative proportion of phaseolin synthesized. We conclude that P. vulgaris axes in culture continue to develop embryonically for a period of time which seems to be under intrinisc control by the axis. This contrasts with precocious germanation, a pattern of embryo behavior seen in many other species. When such embryos are excised from seeds while immature and placed in culture, they switch promptly from embryo development into germination. If ABA or water stress is responsible for preventing precocious germination, it may be that a high level of ABA is maintained or synthesized internally by embryonic axes of Phaseolus, while in other embryos the maternal environment supplies ABA and/or causes water stress.Abbreviations ABA Abscisic acid - BA benzyladenine     

Temperature effects on embryonic abscisic acid levels during development of wheat grain dormancy

Abscisic acid (ABA) levels were determined in both the embryo and remaining grain remnant during development of wheat caryopses under temperature conditions which produced either high or low levels of dormancy in mature grain. Higher levels of grain dormancy were produced in grain from plants grown at 15°C as compared to 25°C. In grain grown at 15°C, embryonic ABA levels steadily increased during development, reaching a maximum at stage IV, just before grain desiccation. At 25°C, ABA levels were very high at the earliest stages of embryonic development, but dropped rapidly during maturation. Only small cultivar differences in ABA levels were observed during development at either temperature. In general, higher levels of dormancy in mature grain correlated with prolonged elevation of ABA levels during grain maturation.     

离体发育和萌发中花生种子不同部份内源ABA含量的变化

离体发育和萌发中花生种子不同部份的内源ＡＢＡ含量变化存在明显的差异．种皮和子叶内存在ＡＢＡ的Ｃ（４０）生物合成途径,胚轴内恻为Ｃ（１５）生物合成途径．种子内源ＡＢＡ含量变化和种子活力有密切关系．     

Dehydrin-like proteins in castor bean seeds and seedlings are differentially produced in response to ABA and water-deficit-related stresses

The stress inducibility of dehydrin protein production in seedlingsof castor bean was analysed by subjecting them to ABA and variouswater-deficit-related treatments including desiccation, waterstress, high salt, high osmolarity, and low temperature. A furthergoal was to determine whether the immature seed (at stages priorto major dehydrin synthesis) would respond in a similar mannerto these stresses. A number of dehydrin-like proteins increasedin seedlings subjected to the various stress treatments. Inthe endosperm, these appear to be different from the dehydrin-relatedpolypeptides that are induced during late seed development andwhich persist following germination/growth of mature seeds.In the endosperm of seedlings, ABA, water stress and desiccationinduced the same dehydrin polypeptides, while high osmolarity,high salt and low temperature induced a different set. Stress-specificdifferences in dehydrin synthesis were also found in the cotyledonsand radicle of castor bean seedlings; however, dehydrins indu-cibleby exogenous ABA were consistently produced. Immature seedstreated with ABA or subjected to stress responded by producingdehydrin-like proteins associated with late development; however,the same proteins were induced following detachment of immatureseeds from the parent plant and maintenance on water. When seedlingswere exposed simultaneously to GA and either ABA, high salt,or low temperature, dehydrin production was suppressed. It isconcluded that dehydrin production in castor bean is tissue-specificand is dependent upon the physiological stage of the seed. Inthe endosperm, the response to different stresses may rely uponmore than one signal trans-duction pathway. Key words: Dehydrin, castor bean, ABA, desiccation     

Concentrations of Abscisic Acid and Indole-3-Acetic Acid in Soybean Seeds during Development

Concentrations of abscisic acid (ABA) and indole-3-acetic acid (IAA) in seed parts were determined during reproductive development of soybean plants (Glycine max [L.] Merr. cv `Chippewa 64'). The concentration of ABA and IAA changed independently in individual seed parts with time. Measurement of the level of ABA and IAA in whole seeds masked the changes which occurred in individual seed tissues. The concentration of ABA was generally highest and that of IAA was generally lowest in the embryonic axis of soybean seeds. In the testa, the IAA concentration was generally highest while the ABA concentration was generally the lowest compared to other parts of the seed.     

Changes in abscisic acid and its β-D-glucopyranosyl ester levels during tomato (Lycopersicon esculentum Mill.) seed development

Summary The role of abscisic acid (ABA) in tomato (Lycopersicon esculentum Mill.) zygotic embryogenesis was analysed. ABA and ABA ß-D-glucopyranosyl ester (ABA-GE) changes were determined in seeds and fruit tissues — placenta and mesocarp — during seed development, which was defined with eight embryo stages: from globular (stage 1) to mature embryo (stage 8). In whole seeds, ABA changes paralleled fresh and dry weight pattern curves and could be characterized by a high increase during embryo growth followed by a decrease as the seed matured and dehydrated. Moreover this dehydration phase led, at stage 8, to a new ABA distribution within the seed, preferentially into integument and embryo. Fruit tissue analyses provided new information about the ABA origin in seeds. ABA-GE levels were also measured and the results suggested different ABA metabolism in seed and fruit tissues.Abbreviations ABA abscisic acid - ABA-GE abscisic acid ß-D-glucopyranosyl ester - ABTS 2,2 — azino — bis (3 — ethylben-zthiazoline — 6 — sulfonic acid) - BHT butylhydroxytoluene - DW dry weight - ELISA Ezyme linked immunosorbent assay - HPLC high performance liquid chromatography     

Temperature effects on embryonic abscisic acid levels during development of wheat grain dormancy

Abscisic acid (ABA) levels were determined in both the embryo and remaining grain remnant during development of wheat caryopses under temperature conditions which produced either high or low levels of dormancy in mature grain. Higher levels of grain dormancy were produced in grain from plants grown at 15°C as compared to 25°C. In grain grown at 15°C, embryonic ABA levels steadily increased during development, reaching a maximum at stage IV, just before grain desiccation. At 25°C, ABA levels were very high at the earliest stages of embryonic development, but dropped rapidly during maturation. Only small cultivar differences in ABA levels were observed during development at either temperature. In general, higher levels of dormancy in mature grain correlated with prolonged elevation of ABA levels during grain maturation.Contribution from USDA-ARS and the College of Agriculture and Home Economics Research Center, Washington State University, scientific paper no. 8901-13.Mention of a specific product name by the United States Department of Agriculture does not constitute an endorsement and does not imply a recommendation over other suitable products.     

Hormonal Complex and Ultrastructure of Maturing Aesculus hippocastanum Seeds

The content and temporal changes in the endogenous IAA, cytokinins, gibberellin-like compounds (GLC), and ABA were determined during horse chestnut (Aesculus hippocastanum L.) seed development (the stages of embryo axis development, its active growth, and storage compound deposition). The active growth of the embryo was characterized by the highest amounts of free phytohormones. Later, by the end of seed maturation, we observed the accumulation of the bound forms of IAA and ABA and a trend to a decrease in the content of free IAA, zeatin, and GLC (butanol fraction). The electron-microscopic examination of the embryo from the mature seed demonstrated that some structural components of the cytoplasm were similar in the cells of embryo axes and cotyledons. During the entire period of maturation, the embryo cells preserved native vacuoles and protein bodies were not formed. Thus, the structure of cotyledonary and axial cells and the distribution of free and bound phytohormones in the horse-chestnut seeds are similar to those in maturing seeds characterized by exogenous dormancy.     

Heat-stable proteins and abscisic Acid action in barley aleurone cells

[³⁵S]Methionine labeling experiments showed that abscisic acid (ABA) induced the synthesis of at least 25 polypeptides in mature barley (Hordeum vulgare) aleurone cells. The polypeptides were not secreted. Whereas most of the proteins extracted from aleurone cells were coagulated by heating to 100°C for 10 minutes, most of the ABA-induced polypeptides remained in solution (heat-stable). ABA had little effect on the spectrum of polypeptides that were synthesized and secreted by aleurone cells, and most of these secreted polypeptides were also heatstable. Coomassie blue staining of sodium dodecyl sulfate polyacrylamide gels indicated that ABA-induced polypeptides already occurred in high amounts in mature aleurone layers having accumulated during grain development. About 60% of the total protein extracted from mature aleurone was heat stable. Amino acid analyses of total preparations of heat-stable and heat-labile proteins showed that, compared to heat-labile proteins, heat-stable intracellular proteins were characterized by higher glutamic acid/glutamine (Glx) and glycine levels and lower levels of neutral amino acids. Secreted heat-stable proteins were rich in Glx and proline. The possibilities that the accumulation of the heat-stable polypeptides during grain development is controlled by ABA and that the function of these polypeptides is related to their abundance and extraordinary heat stability are considered.     

Control of rice pre‐harvest sprouting by glutaredoxin‐mediated abscisic acid signaling

Pre‐harvest sprouting (PHS) is one of the major problems in cereal production worldwide, which causes significant losses of both yield and quality; however, the molecular mechanism underlying PHS remains largely unknown. Here, we identified a dominant PHS mutant phs9‐D. The corresponding gene PHS9 encodes a higher plant unique CC‐type glutaredoxin and is specifically expressed in the embryo at the late embryogenesis stage, implying that PHS9 plays some roles in the late stage of seed development. Yeast two‐hybrid screening showed that PHS9 could interact with OsGAP, which is an interaction partner of the abscicic acid (ABA) receptor OsRCAR1. PHS9‐ or OsGAP overexpression plants showed reduced ABA sensitivity in seed germination, whereas PHS9 or OsGAP knock‐out mutant plants showed increased ABA sensitivity in seed germination, suggesting that PHS9 and OsGAP acted as negative regulators in ABA signaling during seed germination. Interestingly, the germination of PHS9 and OsGAP overexpression or knock‐out plant seeds was weakly promoted by H₂O₂, implying that PHS9 and OsGAP could affect reactive oxygen species (ROS) signaling during seed germination. These results indicate that PHS9 plays an important role in the regulation of rice PHS through the integration of ROS signaling and ABA signaling.     

Quantification of ABA and its metabolites in sweet cherries usingdeuterium-labeled internal standards

Abscisic acid (ABA), phaseic acid (PA), dihydrophaseic acid (DPA), and epi-dihydrophaseic acid (epi-DPA) were quantified in developing fruit and seeds of sweet cherry using each deuterium-labeled internal standard. ABA concentrations in the pulp were low at the early stage of fruit development, reached to the maximum before maturation, and subsequently declined during maturation. The significant increase of ABA after 29 days after full bloom (DAFB) coincides with the softening suggests that ABA may play a role to induce fruit maturation in sweet cherries. ABA metabolite levels were high at the immature stage and decreased with fruit maturation. This fact suggests that fruit may not need ABA in the early stage of fruit development. It was considered that DPA may be the major metabolite of ABA since the concentrations were higher than PA and epi-DPA at all stages of fruit development. ABA concentrations increased at the beginning of seed maturation and then decreased toward harvest. This decrease may be necessary to end seed dormancy. DPA in seeds changed similarly with ABA but its concentrations were always higher than those of ABA.     

Maternal synthesis of abscisic acid controls seed development and yield in<Emphasis Type="Italic"> Nicotiana plumbaginifolia</Emphasis>

The role of maternally derived abscisic acid (ABA) during seed development has been studied using ABA-deficient mutants of Nicotiana plumbaginifolia Viviani. ABA deficiency induced seed abortion, resulting in reduced seed yield, and delayed growth of the remaining embryos. Mutant grafting onto wild-type stocks and reciprocal crosses indicated that maternal ABA, synthesized in maternal vegetative tissues and translocated to the seed, promoted early seed development and growth. Moreover ABA deficiency delayed both seed coat pigmentation and capsule dehiscence. Mutant grafting did not restore these phenotypes, indicating that ABA synthesized in the seed coat and capsule envelope may have a positive effect on capsule and testa maturation. Together these results shed light on the positive role of maternal ABA during N. plumbaginifolia seed development.Abbreviations ABA Abscisic acid - DAP Days after pollination - g Grafted - Wt Wild-type     

IAA, ABA, polyamines and free amino acids associated with zygotic embryo development of Ocotea catharinensis

The aim of this work was to study morphological and biochemical aspects during zygotic embryogenesis in O. catharinensis, by measuring changes in the endogenous concentrations of proteins, amino acids, polyamines (PAs), indole-3-acetic acid (IAA) and abscisic acid (ABA). Buffer-soluble and insoluble protein contents were determined by spectrometry, and amino acids, PAs, IAA and ABA concentrations were determined by high performance liquid chromatography. Total amino acid accumulation, predominantly asparagine, occurred when the embryo showed completely developed cotyledons, with posterior reduction in the mature embryo. This decrease in total amino acid concentration in the mature embryo may result from their use in storage␣as well as for LEA protein synthesis. Free putrescine (Put) concentration decreased, while free spermine (Spm) increased during embryo development. This suggest a role for Put in the initial phases of embryogenesis when high rates of cell division occur, while elevated concentration of Spm are essential from the middle to the end of embryo development, when growth is mainly due to cell elongation. An IAA peak in zygotic embryos occurred during initial development, suggesting a link between growth and cellular division as well as with the establishment of bilateral symmetry. ABA concentration declined during initial stages of development then increased at the mature embryo stage, suggesting a possible relationship with dormancy and recalcitrance characteristics. Our results show that changes in the phytohormones (IAA, ABA and PAs) concentrations in combination with amino acids are likely important factors determining the developmental stages of O.␣catharinensis zygotic embryos.     

Abscisic Acid Represses Growth of the Arabidopsis Embryonic Axis after Germination by Enhancing Auxin Signaling

Under unfavorable environmental conditions, the stress phytohormone ABA inhibits the developmental transition from an embryo in a dry seed into a young seedling. We developed a genetic screen to isolate Arabidopsis thaliana mutants whose early seedling development is resistant to ABA. Here, we report the identification of a recessive mutation in AUXIN RESISTANT1 (AUX1), encoding a cellular auxin influx carrier. Although auxin is a major morphogenesis hormone in plants, little is known about ABA–auxin interactions during early seedling growth. We show that aux1 and pin2 mutants are insensitive to ABA-dependent repression of embryonic axis (hypocotyl and radicle) elongation. Genetic and physiological experiments show that this involves auxin transport to the embryonic axis elongation zone, where ABA enhances the activity of an auxin-responsive promoter. We propose that ABA represses embryonic axis elongation by potentiating auxin signaling in its elongation zone. This involves repression of the AUXIN INDUCIBLE (Aux/IAA) gene AXR2/IAA7, encoding a key component of ABA- and auxin-dependent responses during postgerminative growth.