Effect of Salicylhydroxamic Acid on Endosperm Strength and Embryo Growth of Lactuca sativa L. cv Waldmann''s Green Seeds

Salicylhydroxamic acid (SHAM) stimulated germination of photosensitive lettuce (Lactuca sativa L. cv Waldmann's Green) seeds in darkness. To determine whether SHAM acts on the embryo or the endosperm, we investigated separately effects of SHAM on growth potential of isolated embryos as well as on endosperm strength. Embryo growth potential was quantified by incubating decoated embryos in various concentrations of osmoticum and measuring subsequent radicle elongation. Growth potential of embryos isolated from seeds pretreated with 4 millimolar SHAM was equal to that of untreated controls. Rupture strength of endosperm tissue excised from seeds pretreated with SHAM was 33% less than that of controls in the micropylar region. To determine if the embryo must be in contact with the endosperm for SHAM to weaken the endosperm, some endosperms were incubated with SHAM only after dissection from seeds. Rupture strength of SHAM-treated, isolated endosperms in the micropylar region was 25% less than that of untreated controls. There was no difference in rupture strength in the cotyledonary region of endosperm isolated from seeds treated with SHAM in buffer or buffer alone. SHAM therefore stimulates germination not by enhancing embryo growth potential, but by weakening the micropylar region of the endosperm enclosing the embryo.     

Ketose reductase activity in developing maize endosperm

Ketose reductase (NAD-dependent polyol dehydrogenase EC 1.1.1.14) activity, which catalyzes the NADH-dependent reduction of fructose to sorbitol (d-glucitol), was detected in developing maize (Zea mays L.) endosperm, purified 104-fold from this tissue, and partially characterized. Product analysis by high performance liquid chromatography confirmed that the enzyme-catalyzed reaction was freely reversible. In maize endosperm, 15 days after pollination, ketose reductase activity was of the same order of magnitude as sucrose synthase activity, which produces fructose during sucrose degradation. Other enzymes of hexose metabolism detected in maize endosperm were present in activities of only 1 to 3% of the sucrose synthase activity. CaCl₂, MgCl₂, and MnCl₂ stimulated ketose reductase activity 7-, 6-, and 2-fold, respectively, but had little effect on NAD-dependent polyol dehydrogenation (the reverse reaction). The pH optimums for ketose reductase and polyol dehydrogenase reactions were 6.0 and 9.0, respectively. K_m values were 136 millimolar fructose and 8.4 millimolar sorbitol. The molecular mass of ketose reductase was estimated to be 78 kilodaltons by gel filtration. It is postulated that ketose reductase may function to metabolize some of the fructose produced during sucrose degradation in maize endosperm, but the metabolic fate of sorbitol produced by this reaction is not known.     

A germination-specific endo-beta-mannanase gene is expressed in the micropylar endosperm cap of tomato seeds

Endo-beta-mannanase (EC 3.2.1.78) is involved in hydrolysis of the mannan-rich cell walls of the tomato (Lycopersicon esculentum Mill.) endosperm during germination and post-germinative seedling growth. Different electrophoretic isoforms of endo-beta-mannanase are expressed sequentially in different parts of the endosperm, initially in the micropylar endosperm cap covering the radicle tip and subsequently in the remaining lateral endosperm surrounding the rest of the embryo. We have isolated a cDNA from imbibed tomato seeds (LeMAN2) that shares 77% deduced amino acid sequence similarity with a post-germinative tomato mannanase (LeMAN1). When expressed in Escherichia coli, the protein encoded by LeMAN2 cDNA was recognized by anti-mannanase antibody and exhibited endo-beta-mannanase activity, confirming the identity of the gene. LeMAN2 was expressed exclusively in the endosperm cap tissue of tomato seeds prior to radicle emergence, whereas LeMAN1 was expressed only in the lateral endosperm after radicle emergence. LeMAN2 mRNA accumulation and mannanase activity were induced by gibberellin in gibberellin-deficient gib-1 mutant seeds but were not inhibited by abscisic acid in wild-type seeds. Distinct mannanases are involved in germination and post-germinative growth, with LeMAN2 being associated with endosperm cap weakening prior to radicle emergence, whereas LeMAN1 mobilizes galactomannan reserves in the lateral endosperm.     

Relationship of Endo-[beta]-D-Mannanase Activity and Cell Wall Hydrolysis in Tomato Endosperm to Germination Rates

The endosperm tissue enclosing the radicle tip (endosperm cap) governs radicle emergence in tomato (Lycopersicon esculentum Mill.) seeds. Weakening of the endosperm cap has been attributed to hydrolysis of its mannan-rich cell walls by endo-[beta]-D-mannanase. To test this hypothesis, we measured mannanase activity in tomato endosperm caps from seeds allowed to imbibe under conditions of varying germination rates. Over a range of suboptimal temperatures, mannanase activity prior to radicle emergence increased in accordance with accumulated thermal time. Reduced water potential delayed or prevented radicle emergence but enhanced mannanase activity in the endosperm caps. Abscisic acid did not prevent the initial increase in mannanase activity, although radicle emergence was markedly delayed. Sugar composition and percent mannose (Man) content of endosperm cap cell walls did not change prior to radicle emergence under any condition. Man, glucose, and other sugars were released into the incubation solution by endosperm caps isolated from intact seeds during imbibition. Pregerminative release of Man was suppressed and the release of glucose was enhanced when seeds were incubated in osmoticum or abscisic acid; the opposite occurred in the presence of gibberellin. Thus, whereas sugar release patterns were sensitive to environmental and hormonal factors affecting germination, neither assayable endo-[beta]-D-mannanase activity nor changes in cell wall sugar composition of endosperm caps correlated well with tomato seed germination rates under all conditions.     

Callose Deposition Is Responsible for Apoplastic Semipermeability of the Endosperm Envelope of Muskmelon Seeds

Semipermeable cell walls or apoplastic “membranes” have been hypothesized to be present in various plant tissues. Although often associated with suberized or lignified walls, the wall component that confers osmotic semipermeability is not known. In muskmelon (Cucumis melo L.) seeds, a thin, membranous endosperm completely encloses the embryo, creating a semipermeable apoplastic envelope. When dead muskmelon seeds are allowed to imbibe, solutes leaking from the embryo are retained within the envelope, resulting in osmotic water uptake and swelling called osmotic distention (OD). The endosperm envelope of muskmelon seeds stained with aniline blue, which is specific for callose (β-1,3-glucan). Outside of the aniline-blue-stained layer was a Sudan III- and IV-staining (lipid-containing) layer. In young developing seeds 25 d after anthesis (DAA) that did not exhibit OD, the lipid layer was already present but callose had not been deposited. At 35 DAA, callose was detected as distinct vesicles or globules in the endosperm envelope. A thick callose layer was evident at 40 DAA, coinciding with development of the capacity for OD. Removal of the outer lipid layer by brief chloroform treatment resulted in more rapid water uptake by both viable and nonviable (boiled) seeds, but did not affect semipermeability of the endosperm envelope. The aniline-blue-staining layer was digested by β-1,3-glucanase, and these envelopes lost OD. Thus, apoplastic semipermeability of the muskmelon endosperm envelope is dependent on the deposition of a thick callose-containing layer outside of the endosperm cell walls.     

SEED DEVELOPMENT IN CITRUS WITH SPECIAL REFERENCE TO 2X X 4X CROSSES

Comparative cytological and histological studies during embryogenesis of seeds from 2x X 2x and 2x x 4x crosses indicate that the ratio of ploidy level between embryo and endosperm is the most important factor affecting the course of seed development. The crosses produced seeds with the expected ploidy relationships between embryo, endosperm, and maternal tissue of 2:3:2 and 3:4:2 as well as the anomalous relationships 3:5:2, 4:6:2, and 6:10:2. All but 3:4:2 resulted in normal, germinable seeds. The ploidy level of the maternal tissue in relation to that of the embryo or endosperm did not appear to have any effect on seed development. About 92–99 % of seeds from 2x x 4x crosses containing triploid embryos with tetraploid endosperm aborted at different stages of embryogenesis. The abortion in all cases was preceded by abnormalities in the tetraploid endosperm. It is postulated that the unbalance of chromosome number between embryo and endosperm disturbs physiological relationships between these two tissues, leading first to the abortion of the endosperm and then of the embryo.     

Brassinosteroids and gibberellins promote tobacco seed germination by distinct pathways

Seed germination of Nicotiana tabacum L. cv. Havana 425 is determined by the balance of forces between the growth potential of the embryo and the mechanical restraint of the micropylar endosperm. In contrast to the gibberellin GA4, the brassinosteroid (BR) brassinolide (BL) did not release photodormancy of dark-imbibed photodormant seeds. Brassinolide promoted seedling elongation and germination of non-photodormant seeds, but did not appreciably affect the induction of class I beta-1,3-glucanase (betaGLU I) in the micropylar endosperm. Brassinolide, but not GA4, accelerated endosperm rupture of tobacco seeds imbibed in the light. Brassinolide and GA4 promoted endosperm rupture of dark-imbibed non-photodormant seeds, but only GA4 enhanced betaGLU I induction. Promotion of endosperm rupture by BL was dose-dependent and 0.01 microM BL was most effective. Brassinolide and GA4 promoted abscisic acid (ABA)-inhibited dark-germination of non-photodormant seeds, but only GA4 replaced light in inducing betaGLU I. These results indicate that BRs and GAs promote tobacco seed germination by distinct signal transduction pathways and distinct mechanisms. Gibberellins and light seem to act in a common pathway to release photodormancy, whereas BRs do not release photodormancy. Induction of betaGLU I in the micropylar endosperm and promotion of release of 'coat-enhanced' dormancy seem to be associated with the GA-dependent pathway, but not with BR signalling. It is proposed that BRs promote seed germination by directly enhancing the growth potential of the emerging embryo in a GA- and betaGLU I-independent manner.     

Endosperm protein of wheat seed as a determinant of seedling growth

Seed of a Mexican semidwarf wheat (Triticum aestivum L. cv. Inia 66), was obtained from a nitrogen fertilizer field trial grown in Mexico. A high positive correlation was obtained between seed protein content and seedling dry weight after 3 weeks growth (r = +0.92^**). The seedling dry weight was positively related to the protein content of the aleurone layer and endosperm, but not to the embryo. Small, 35 milligrams, high protein seeds (4.7 milligrams protein per seed) produced larger seedlings than large, 45 milligrams, low protein seeds (4.3 milligram protein per seed). There was no difference in the weight or protein content of embryos from low and high protein seeds and their growth was similar. Composite seeds of the two protein levels were produced by transferring embryos from one endosperm type to the other. After 4 weeks, there was no difference between the different embryo types grown on the same endosperm type. High protein endosperm produced more vigorous seedlings regardless of the embryo type grown on it, indicating that the factor(s) responsible for the greater growth of high protein seed is in the endosperm.     

Tissue-specific transcriptome analysis reveals cell wall metabolism, flavonol biosynthesis and defense responses are activated in the endosperm of germinating Arabidopsis thaliana seeds

Seed germination is a result of the competition of embryonic growth potential and mechanical constraint by surrounding tissues such as the endosperm. To understand the processes occurring in the endosperm during germination, we analyzed tiling array expression data on dissected endosperm and embryo from 6 and 24 h-imbibed Arabidopsis seeds. The genes preferentially expressed in the endosperm of both 6 and 24 h-imbibed seeds were enriched for those related to cell wall biosynthesis/modifications, flavonol biosynthesis, defense responses and cellular transport. Loss of function of AtXTH31/XTR8, an endosperm-specific gene for a putative xyloglucan endotransglycosylase/hydrolase, led to faster germination. This suggests that AtXTH31/XTR8 is involved in the reinforcement of the cell wall of the endosperm during germination. In vivo flavonol staining by diphenyl boric acid aminoethyl ester (DPBA) showed flavonols accumulated in the endosperm of both dormant and non-dormant seeds, suggesting that this event is independent of germination. Notably, DPBA fluorescence was also intense in the embryo, but the fluorescent region was diminished around the radicle and lower half of the hypocotyl during germination. DPBA fluorescence was localized in the vacuoles during germination. Vacuolation was not seen in imbibed dormant seeds, suggesting that vacuolation is associated with germination. A gene for δVPE (vacuolar processing enzyme), a caspase-1-like cysteine proteinase involved in cell death, is expressed specifically in endosperms of 24 h-imbibed seeds. The δvpe mutant showed retardation of vacuolation, but this mutation did not affect the kinetics of germination. This suggests that vacuolation is a consequence, and not a trigger, of germination.     

Significant contribution of boron stored in seeds to initial growth of rice seedlings

Rice plants are relatively insensitive to low boron environment. We examined the role of boron stored in seeds in growth of rice seedlings in regard to its insensitivity to low boron condition. In experiments using seeds without the removal of the endosperm, the effect of low boron treatments (0.18 and 0.03 μM B) was not evident on growth of seedlings until 5 weeks from germination. Analysis of boron distribution among different organs in rice grain demonstrated that the endosperm contained 92% of total boron in brown rice and husks contained twice as much boron as the brown rice. To examine the role of boron stored in the endosperm, endosperm were removed 5 days after de-husking and imbibition and the plants were subjected to boron treatments. The growth of the plants without endosperm was much reduced after 10 days under the 0.03 μM boron compared with the plants grown with the normal level (18 μM B). With the normal level of boron supply, the growth did not differ between the plants with and without endosperm, suggesting that growth difference between seedlings with and without endosperm under low B condition is not due to lack of starch or proteins, but due to lack of boron stored in seeds. Tracer experiments confirmed that a substantial amount of B was supplied from the seed to the seedling and that B limitation increased the B supply from seeds to seedlings, especially to roots. Our results established that boron stored in seeds, principally in the endosperm, contributes significantly to the growth of rice seedlings when external boron supply is limited.     

Is The Barley Endosperm a Water Reservoir for the Embryo When Germinating Seeds Are Dried?

The water content of germinating seeds fluctuates in response to water potential changes in the surrounding environment. We tested the hypothesis that the endosperm functions as a water reservoir when imbibed seeds experience drying, and we characterized water uptake and movement within barley (Hordeum vulgare cv. Triumph) caryopses (hereafter referred to as seeds). Water movement into and through germinating barley seeds during imbibition and drying was determined gravimetrically and with the fluorescent dye trisodium 8-hydroxy-1,3,6-pyrenetrisulfonate (PTS). During imbibition, embryo tissues hydrated more rapidly and reached a higher water content (g H20/g dry weight) than did the endosperm, although the endosperm eventually contained nine times as much total water. When barley seeds that had imbibed for 12 h were exposed to moderate (-4 MPa) drying, PTS solution moved from the endosperm into the shoot meristem, radicle, and scutellum, but not vice versa. Radicle emergence and elongation proceeded for up to 8 h. With harsh (-150 MPa) drying, PTS concentrated almost exclusively in the radicle. These data illustrate that the endosperm is at least a temporary water storage compartment external to the embryo itself. We speculate that water supplied by the endosperm may be important in reducing the harmful effects of drying during the critical transition period when a germinating seed changes from a desiccation-tolerant to a desiccation-intolerant organism.     

Relationships between seed weight and endosperm characteristics in carrot

Carrot seed weight, over the range 0·37 to 3·77 mg, and endosperm volume were closely and linearly related to endosperm cell number, the relationships accounting for 77–85% and 84–94% of the variance in seed weight and endosperm volume, respectively. The relationships were similar for seeds of cvs Chantenay and Amsterdam, and for seeds from primary and secondary umbels but were affected by season of seed production through differences in cell volume.     

东北刺人参种子萌发影响因子的研究

东北刺人参(Oplopanax elatus Nakai)种子透水性良好,休眠后萌发不受其影响.种皮和胚乳的水提取物中存在萌发抑制物质,胚乳中提取物对白菜种子萌发的抑制效果比种皮更明显.种子自然脱落时胚尚未分化完全,处于心形胚阶段.种子需要先温暖层积以完成胚的分化与生长,然后转入低温层积完成生理后熟.同批种子胚的发育不完全同步,变温层积处理7个月有极少数种子萌发,连续变温层积处理17个月大部分种子萌发.不同年份受气候条件影响,种子产量和发芽率差异较大.种子耐贮性较强,贮藏2年的种子生活力变化不大,仍具有较高的萌发潜力.     

AAP1 regulates import of amino acids into developing Arabidopsis embryos

The embryo of Arabidopsis seeds is symplasmically isolated from the surrounding seed coat and endosperm, and uptake of nutrients from the seed apoplast is required for embryo growth and storage reserve accumulation. With the aim of understanding the importance of nitrogen (N) uptake into developing embryos, we analysed two mutants of AAP1 (At1g58360), an amino acid transporter that was localized to Arabidopsis embryos. In mature and desiccated aap1 seeds the total N and carbon content was reduced while the total free amino acid levels were strongly increased. Separately analysed embryos and seed coats/endosperm of mature seeds showed that the elevated amounts in amino acids were caused by an accumulation in the seed coat/endosperm, demonstrating that a decrease in uptake of amino acids by the aap1 embryo affects the N pool in the seed coat/endosperm. Also, the number of protein bodies was increased in the aap1 endosperm, suggesting that the accumulation of free amino acids triggered protein synthesis. Analysis of seed storage compounds revealed that the total fatty acid content was unchanged in aap1 seeds, but storage protein levels were decreased. Expression analysis of genes of seed N transport, metabolism and storage was in agreement with the biochemical data. In addition, seed weight, as well as total silique and seed number, was reduced in the mutants. Together, these results demonstrate that seed protein synthesis and seed weight is dependent on N availability and that AAP1-mediated uptake of amino acids by the embryo is important for storage protein synthesis and seed yield.     

东北刺人参种子萌发影响因子的研究

东北刺人参(Oplopanax elatus Nakai)种子透水性良好,休眠后萌发不受其影响。种皮和胚乳的水提取物中存在萌发抑制物质,胚乳中提取物对白菜种子萌发的抑制效果比种皮更明显。种子自然脱落时胚尚未分化完全,处于心形胚阶段。种子需要先温暖层积以完成胚的分化与生长,然后转入低温层积完成生理后熟。同批种子胚的发育不完全同步,变温层积处理7个月有极少数种子萌发,连续变温层积处理17个月大部分种子萌发。不同年份受气候条件影响,种子产量和发芽率差异较大。种子耐贮性较强,贮藏2年的种子生活力变化不大,仍具有较高的萌发潜力。     

Studies of sugars and sorbitol in developing corn kernels

Sugars and sorbitol were determined on corn (Zea mays L.) kernels harvested at various developmental stages, using sugary (su), sugary-sugary enhancer (su se), and starchy (Su) cultivars. In all cultivars tested, the sorbitol content increased from trace amounts in unpollinated ovules to a maximum at about the time that rapid starch synthesis was proceeding. Thereafter, sorbitol and sugars decreased continuously to the mature dry stage. Sorbitol in the su se kernels was higher than that of other cultivars from 28 days postpollination onwards; sucrose and maltose were higher from 21 days onwards. [¹⁴C]Sorbitol was recovered from kernel base, pedicel, and endosperm of IL677a (su se) kernels after allowing a flag leaf to fix ¹⁴CO₂ photosynthetically. No [¹⁴C]sorbitol was detected in the shank of the ear, and none was detected by the gas chromatograph. [¹⁴C]Sucrose was the predominant labeled substance recovered from the kernel base, pedicel, and endosperm tissues during the 10-h chase period, as well as from the shank of the ear, and nonradioactive sucrose was the predominant ethanol-soluble compound detected by the gas chromatograph. Hence, sorbitol appears not to be translocated from corn leaves as it is in certain woody plants of the rose family. The altered sugar profile of su se kernels may be related to reduced starch synthesis, but the biochemical mechanism is not yet known.     

Effect of laser irradiation on microviscosity of aqueous medium in imbibing maize seeds as studied with a spin probe method

A spin probe method was used to study microviscosity of aqueous medium in embryos and endosperm of maize (Zea mays L.) seeds after their irradiation with a Lvov-1 Elektronika laser device. Based on parameters of electron spin resonance (ESR) spectra of nitroxyl radicals (probes) absorbed by imbibing seeds during water uptake, correlation times τ_C were determined for the rotational diffusion of probes in embryos and endosperm of seeds. The τ_C values were found smaller in embryos of irradiated seeds than in untreated seeds; the dependence of τ_C on the duration of seed imbibition was determined. It is concluded that laser irradiation of seeds decreases the microviscosity of aqueous medium in embryo cells and elevates the mobility of the probes. Effect of laser irradiation on τ_C in seed endosperm was less pronounced but also led to the increase in probe mobility.     

Galactomannan hydrolyzing activity develops during priming in the micropylar endosperm tip of tomato seeds

Development of galactomannan hydrolyzing activity was followed in seeds of tomato [ Lycopersicon esculentum (L.) Mill. cv. Toyonishiki] during priming and germination. The activity developed in seeds that were being primed in polyethylene glycol (-0.8 MPa). The activity was detected exclusively in the endosperm portion just adjacent to the radicle tip. Part of the activity remained active after desiccation of the primed seeds. After transfer to water, the activity in the primed seeds immediately began to increase, while in unprimed seeds the beginning of the increase in activity was delayed by about 1 day. In scanning electron microscopy, the inner surface of the cell walls of the micropylar endosperm portion appeared eroded in primed seeds that had been imbibed in water for 16 h (before germination), but not in unprimed seeds imbibed for the same period. These results support the hypothesis that galactomannan hydrolyzing enzyme, which is believed to be responsible for breakdown of tomato endosperm cell walls and hence for the weakening of mechanical restraint against radicle growth, may be involved in the improved germination of primed tomato seeds.