Acidification of the Medium Associated with Normal and Fusicoccin-Induced Seed Germination

Fusicoccin, a toxin stimulating cell enlargement and inducing proton extrusion in various plant tissues, has been shown to replace kinetin, gibberellic acid and red light in breaking seed dormancy. It also removes the inhibitory effect of abscisic acid. The present data also show that the stimulating effect of fucicoccin on embryo growth of decoated radish (Raphanus sativus L.) and maize (Zea mays) seeds and on the development of maize embryos is accompanied by an early, significant acidification of the medium. Acidification of the medium is also observed when fusicoccin reverses the abscisic acid-induced inhibition of germination. These results support the hypothesis that the mode of action of fusicoccin in promoting germination involves, as in stimulation of cell enlargement, the activation at the cell membrane level of proton extrusion processes. The physiological significance of fusicoccin-induced release of protons at the onset of germination is discussed in comparison with the results concerning the mechanism of action of fusicoccin on cell enlargement in other plant materials.     

Differential Expression of the Two Subunits of Tomato Polygalacturonase Isoenzyme 1 in Wild-Type and rin Tomato Fruit

The [beta] subunit of tomato (Lycopersicon esculentum Mill.) fruit polygalacturonase 1 is a cell wall glycoprotein that binds to and apparently regulates the catalytic PG2 polypeptide in vivo. [beta] Subunit and polygalacturonase 2 (PG2) expression have been investigated in both wild-type and ripening inhibitor (rin) mutant fruit. During fruit development and ripening, [beta] subunit expression was unrelated to expression of the catalytic PG2 protein. In wild-type fruit, [beta] subunit mRNA and protein were first detected early in development and increased to maximal levels before PG2 mRNA and protein were detected. At the onset of ripening [beta] subunit mRNA decreased dramatically, but [beta] subunit protein levels remained stable. In rin fruit, which fail to ripen, [beta] subunit expression was similar to that in wild type, although PG2 mRNA and protein were not detected. These data suggest that [beta] subunit expression is ethylene independent and regulated primarily by developmental cues. This conclusion is supported by results from ethylene-treated immature (20 days after pollination) wild-type and rin fruit in which no significant differences were observed in [beta] subunit expression patterns in response to ethylene treatment. Surprisingly, RNA blot analysis indicated that catalytic PG2 mRNA was induced in immature rin fruit after 3 d of exogenous ethylene treatment. In addition, [beta] subunit mRNA and protein were also detected at lower levels in root, leaf, and flower tissues of both genotypes, suggesting a broader functional role for the protein.     

Two Tomato Expansin Genes Show Divergent Expression and Localization in Embryos during Seed Development and Germination

Expansins are plant proteins that can induce extension of isolated cell walls and are proposed to mediate cell expansion. Three expansin genes were expressed in germinating tomato (Lycopersicon esculentum Mill.) seeds, one of which (LeEXP4) was expressed specifically in the endosperm cap tissue enclosing the radicle tip. The other two genes (LeEXP8 and LeEXP10) were expressed in the embryo and are further characterized here. LeEXP8 mRNA was not detected in developing or mature seeds but accumulated specifically in the radicle cortex during and after germination. In contrast, LeEXP10 mRNA was abundant at an early stage of seed development corresponding to the period of rapid embryo expansion; it then decreased during seed maturation and increased again during germination. When gibberellin-deficient (gib-1) mutant seeds were imbibed in water, LeEXP8 mRNA was not detected, but a low level of LeEXP10 mRNA was present. Expression of both genes increased when gib-1 seeds were imbibed in gibberellin. Abscisic acid did not prevent the initial expression of LeEXP8 and LeEXP10, but mRNA abundance of both genes subsequently decreased during extended incubation. The initial increase in LeEXP8, but not LeEXP10, mRNA accumulation was blocked by low water potential, but LeEXP10 mRNA amounts fell after longer incubation. When seeds were transferred from abscisic acid or low water potential solutions to water, abundance of both LeEXP8 and LeEXP10 mRNAs increased in association with germination. The tissue localization and expression patterns of both LeEXP8 and LeEXP10 suggest developmentally specific roles during embryo and seedling growth.     

Tomato Seed Germination. Osmotic Pretreatment and Far Red Inhibition

At 25 °C germination of tomato (Lycopersicon lycopersicum)seeds is inhibited by continuous and intermittent far red illumination.It is also inhibited by a single 30 min far red irradiationgiven 8 h from the start of imbibition. The incubation of seedsin a mannitol solution inhibitory for germination has no effecton the final germination percentage after seeds are subsequentlytransferred to water. A 30 min far red irradiation at the timeof transfer results in partial inhibition of germination. Thisinhibition can be released by the continuation of osmotic incubationfor several days before the transfer to water. At the end ofa 7 d dark period of osmotic incubation, inhibition of subsequentgermination in water can be realized only by continuous farred illumination. Seeds osmotically pretreated for 7 d and afterwardsdried-back show a mean time to 50% germination significantlylower than that of untreated seeds. Moreover, besides singleand intermittent, even continuous far red light has no inhibitoryeffect on the germination of these seeds. It is concluded that,in addition to the already known germination advantages, osmoticpresowing treatment also induces the ability of seeds to germinateunder unfavourable light conditi.     

Ammonium and Salt Inhibition of Some Physiological Processes Associated with Seed Germination

Seed germination was totally inhibited by high (0.1 N) concentrations of potassium or ammonium salts. Partial inhibition of germination occurred with low (< 0.01 N) concentrations of ammonium salts, but low concentrations of potassium salts did not affect germination. Except with extreme salt or ammonium injury, the inhibition of germination was at least partially reversible. Respiration of germinating seeds was inhibited by both species of salts. Seedling development was severely impaired by ammonium salts, but not by potassium salts.     

Cell-Free Synthesis of a Putative Precursor of Polygalacturonase in Tomato Fruits

By using a monospecific anti-polygalacturonase-2 antibody, a54K-dalton polypeptide was detected in in vitro translationproducts by a wheat germ cell-free translation system programmedwith polyadenylated RNA from ripe tomato pericarp tissue. Thisputative precursor of polygalacturonase was about 9K daltonslarger in molecular weight than polygalacturonase-2. (Received December 12, 1983; Accepted May 17, 1984)     

Polygalacturonase Gene Expression in Rutgers, rin, nor, and Nr Tomato Fruits

Polygalacturonase (PG) gene expression was studied in normally ripening tomato fruit (Lycopersicon esculentum Mill, cv Rutgers) and in three ripening-impaired mutants, rin, nor, and Nr. Normal and mutant fruit of identical chronological age were analyzed at 41, 49, and 62 days after pollination. These stages corresponded to mature-green, ripe, and overripe, respectively, for Rutgers. The amount of PG mRNA in Rutgers was highest at 49 days and accounted for 2.3% of the total mRNA mass but at 62 days had decreased to 0.004% of the total mRNA mass. In Nr, the amount of PG mRNA steadily increased between 41 and 62 days after pollination, reaching a maximum level of 0.5% of the total mRNA mass. The mutant nor exhibited barely detectable levels of PG mRNA at all stages tested. Surprisingly, PG mRNA, comprising approximately 0.06% of the mRNA mass, was detected in 49 day rin fruit. This mRNA accumulation occurred in the absence of elevated ethylene production by the fruit and resulted in the synthesis of enzymically active PG I. The different patterns of PG mRNA accumulation in the three mutants suggests that distinct molecular mechanisms contribute to reduced PG expression in each ripening-impaired mutant.     

Epigallocatechin-3-Gallate Alleviates Salinity-Retarded Seed Germination and Oxidative Stress in Tomato

Journal of Plant Growth Regulation - Salinity is a global environmental problem, restricting crop production in a vast area of agricultural land. Although epigallocatechin-3-gallate (EGCG), the...     

番茄果实中乙烯与多聚半乳糖醛酸酶的关系

乙烯与多聚半乳糖醛酸酶(PG)都是果实成熟过程中关键的调节因子.一方面,在有乙烯合成缺陷的转反义ACS番茄和乙烯感受缺陷的Nr突变体番茄果实中PG基因表达量都明显下降,PG酶活性明显降低;用外源乙烯(100 μL/L)处理绿熟期番茄果实使PG基因的表达明显增强,而1-甲基环丙烯(1-MCP,1 μL/L)处理转色期番茄果实明显抑制PG基因表达.另一方面,转反义PG基因番茄果实乙烯释放量在授粉后低于其野生型,番茄乙烯受体基因LeETR4和乙烯反应因子LeERF2基因表达量比野生种低.PG降解果胶的产物D-GA(100 mg/L)促进未熟期番茄果实中的乙烯生成和LeETR4、LeERF2基因的表达.     

Involvement of Endomannanase in the Control of Tomato Seed Germination under Low Temperature Conditions

The cause of differences in germination rates in a cold-toleranttomato line (PI341988), a control line (UC82B), and six progenylines stemming from crosses and backcrosses between the twoparent lines was investigated. Pursuant to earlier work showingthat differences in germination ability at 12°C are dueto the barrier imposed by the endosperm layer, we analysed theactivity of cell-wall-hydrolysing enzymes extracted from theselines. A significant increase in endomannanase activity wasfound in plant line PI341988 prior to germination at 12°C.Extracts of PI341988 seeds that had imbibed at either 12 or25°C exhibited higher endomannanase activity than theircounterparts from plant line UC82B. Moreover, a positive relationshipwas found between germination ability at low temperature andendomannanase activity in the six progeny lines. Analysis ofendomannanase activity in sub-regions of the seed indicatedthat the increase in activity prior to germination was higherin the micropylar endosperm cap than in the rest of the seed.Exogenous application of mannanase originating from soil-bornebacteria increased germination rates under both moderate andlow temperature conditions. Cellulase (endo-1,4-ß-glucanase)activity was also found to be higher in plant line PI341988.However, the activity of this enzyme probably increases aftergermination and it is therefore not considered as a key enzymecontrolling germination at low temperatures.Copyright 1995,1999 Academic Press Lycopersicon esculentum, tomato, seed germination, cell wall     

Polygalacturonase Isozymes and Pectin Depolymerization in Transgenic rin Tomato Fruit

We have previously described the construction and expression of a chimeric gene that allows developmentally regulated expression of tomato (Lycopersicon esculentum) polygalacturonase in ripening-impaired, mutant (rin) tomato fruit (JJ Giovannoni, D DellaPenna, AB Bennett, RL Fischer [1989] The Plant Cell 1: 53-63). We now show that expression of the chimeric polygalacturonase gene in rin tomato fruit resulted in the accumulation of all three polygalacturonase isozymes (PG1, PG2A, and PG2B). Polyuronide solubilization and polyuronide depolymerization both reached their maximal levels in transgenic rin fruit prior to the appearance of PG2 isozymes. These results demonstrate that PG1, PG2A, and PG2B all arise by differential processing of a single gene product and further suggest that the PG1 isozyme is sufficient to carry out both polyuronide solubilization and depolymerization in vivo.     

Mycena subpiligera sp. nov., a Symbiotic Species from China Associated with the Seed Germination of Gastrodia elata

Mycena subpiligera, a new taxon in sect. Fragilipedes that can strongly enhance the germination efficiency of Gastrodia elata seeds, was discovered in subtropical areas of China. As revealed by a morphological comparison with related Mycena species as well as maximum likelihood (ML) and Bayesian phylogenetic analyses based on sequences of the internal transcribed spacer (ITS) and the large subunit (LSU) regions of nuclear ribosomal RNA, the new taxon can be distinguished from phenotypically similar and phylogenetically related species. Optimal cultural conditions for M. subpiligera basidiomata are reported, and the germination rate of the new species is compared with that of M. citrinomarginata.     

Polygalacturonase Activity in Ripening Tomato Fruit Determined using Pericarp Discs

Discs prepared from the outer pericarp of tomato (Lycopersiconesculentum Mill. cv. Sunny) and placed in buffer exhibit anenzymic release of pectin fragments. Over a 2.5 h period at34 °C, discs from mature-green, 4 d and 10 d postbreakerfruit released approximately 90, 440 and 675 µg galacturonicacid equivalents (g^–1 disc fr. wt.), respectively. Bio-GelP-2 chromatography of the products revealed the presence ofpolymeric, oligomeric and monomeric uronic acids. The similarityof these products to those released from isolated, enzymatically-activecell walls and from enzymatically-inactive walls treated withpurified PG 2 provides evidence for the participation of polygalacturonase(PG, E. C. 3.2.1.15 [EC] ) in the release of pectin from disc tissue. The magnitude of pectin release from external pericarp discswas found to parallel ripening and increase progressively indiscs from the blossom, equatorial and shoulder regions, respectively.The use of discs and other systems to estimate in vivo PG activitywill be discussed. Key words: Cell wall, polyuronides     

番茄离体花柄脱落区多聚半乳糖醛酸酶提取纯化体系的优化

多聚半乳糖醛酸酶是植物器官脱落过程中的重要水解酶,实验以20μL.L-1乙烯处理的离体番茄花柄为试材,建立了与脱落相关的多聚半乳糖醛酸酶提取与纯化体系:以50 mmol.L-1乙酸缓冲液(pH=5.5)为提取液,加入0.1 mol.L-1NaCl、1 mmol.L-1DTT提取效果较好;将酶的粗提液低温浓缩后,经Sephadex G-75凝胶层析分离纯化,最佳流速为0.2 mL.min-1,适宜上样量为3.5 mL;再将凝胶层析分离的活性部分低温浓缩后,经CM Sepharose CL-6B离子交换层析再次纯化,流速为0.3 mL.min-1、洗脱液pH值5.5纯化效果最好。经上述提取纯化过程,得到了分子质量为30.2 kD的多聚半乳糖醛酸酶蛋白。该提取纯化体系为探讨与脱落相关酶的性质及其活性调控提供了参考。     

Analysis of Transgenic Tomatoes with Expression of Antisense Polygalacturonase Gene

Authors have been transfered antisense polygalacturonase (PG, E, C, 3, 2, 1, 15) gene into two tomato cultivars (Lycopersicon esculentum cv. “Lichun” and “Qingfeng”) by Agrobacterium-mediated transformation and obtained homozygotes with expression of antisense PG gene. Analysis of Northern blot and PG activity showed that antisense PG gene had stably expressed in T0～T2 generations of transgenic tomatoes, and this expression could be inherited and enhanced in the progeny. For example, PG activity of To, T1 and T2 generation fruits in line T2-19 of"Lichun” was dropped to 27%, 20% and 4% respectively. But the remarkable decrement of PG activity did not have obvious inhibitory effect on normal fruit softening. The transgenic plants of T0～T2 generations grew and developed normally. Average yield per plant and average weight per fruit in these transgenic plants were a little more than those in nontransgenic control. The content determination of vitamin C, soluble sugar and soluble solid substances indicated that transgenic fruits kept the original texture and taste. The contents of vitamin C and soluble solid substances of “Lichun” T2-19 and soluble sugar of “Qingfeng” T2-46 were higher than those of the control. Stored in 28～32℃ for 30 days, all of nontransgenic fruits and about 50% of the transgenic ones were infected by pathologcal bacteria, a few of remainders could be perserved for 50 days. These evidences suggested that the resistance of the transgenic fruits to pathological bacteria during preservation has been strengthened. The approach which the authors used to study regulation of tomato fruit ripening will be desirable to improve the storability of certain famous and precious fruits.     

Polygalacturonase mRNA of Tomato: Size and Content in Ripe Fruits

In ripe tomato fruits, polygalacturonase (PG) mRNA comprisedabout 1% of the translatable RNAs in the poly(A)(+)RNA fraction.Sucrose density gradient centrifugation showed that this PGmRNA is similar in size to 18S rRNA, which suggests the presenceof a non-coding region. (Received June 19, 1984; Accepted October 23, 1984)