Changes in the Composition and Synthesis of Proteins in Cellular Membranes of Echinochloa crus-galli (L.) Beauv. Seeds during the Transition from Dormancy to Germination

The effect of alcohols which stimulate or have no effect on germination on the composition and synthetic pattern of proteins in the cellular membranes of Echinochloa crus-galli (L.) Beauv. seeds was studied. Imbibition of dry seeds was accompanied by an increase in the synthesis of proteins and by synthesis of new proteins in their intracellular membranes. The transition of the seeds from a dormant to a nondormant state was associated with synthesis of specific proteins and a decrease in content of others in the plasma membrane. The synthesis of a 23 kilodalton protein was strongly increased upon release from dormancy. The changes in the pattern of protein synthesis were not directly associated with the beginning of germination. The results suggest that the plasma membrane constitutes the first site in the seed cells, at which the stimulus from external factors affecting seed dormancy is detected.     

Role of Endogenous Plant Growth Regulators in Seed Dormancy of Avena fatua: II. Gibberellins

Gibberellin A₁ (GA₁) was identified by combined gas chromatographymass spectrometry as the major biologically active gibberellin (GA) in seeds of wild oat (Avena fatua L.) regardless of the depth of dormany or stage of imbibition. Both unimbibed dormant and nondromant seeds contained similar amounts of GA₁ as estimated by the d5-maize bioassay. During imbibition, the level of GA₁ declined in both dormant and non-dormant seeds, although the decline was more rapid in dormant seeds. Only in imbibing nondormant seeds did the GA biosynthesis inhibitor, 2-chloroethyltrimethyl ammonium chloride (CCC), cause a reduction in the level of GA₁ from that observed in control seeds. These results are interpreted as an indication that while afterripening does not cause a direct change in the levels of GAs during dry storage, it does induce a greater capacity for GA biosynthesis during imbibition.

Nondormant seeds imbibed in the presence of 50 millimolar CCC germinated equally as well as untreated seeds. When wild oat plants were fed CCC throughout the entire life cycle, viable seeds were produced that lacked detectable GA-like substances. These seeds afterripened at a slightly slower rate than the controls. Moreover, completely afterripened (nondormant) seeds from plants fed CCC continuously contained no detectable GA-like substances, and when these seeds germinated, dwarf seedlings were produced, indicating GA biosynthesis was inhibited during and after germination. In total, these results suggest that the increased capacity for GA biosynthesis observed in imbibing nondormant seeds is not a necessary prerequisite for germination. It is therefore possible that GA biosynthesis in imbibing nondormant seeds is one of many coordinated biochemical events that occur during germination rather than an initiator of the processes leading to germination.

     

Protein and nucleic acid synthesis during imbibition of dormant and after-ripened Vaccaria pyramidata seeds.

The regulation of nucleic acid and protein synthesis in dormant, thermodormant, and after-ripened embryos of Vaccaria pyramidata (Caryophyllaceae) has been studied. Germination of after-ripened V. pyramidata seeds is prevented by inhibitors of protein, RNA, and DNA synthesis. The synthesis of both protein and RNA is activated at the beginning of imbibition, whereas [³H]thymidine incorporation does not start until the second period of the imbibition phase. [³H]Thymidine incorporation is greatly reduced in embryos treated with cycloheximide or 6-methylpurine. There is no correlation between the level of [³H]uracil and l-[¹⁴C]leucine incorporation into macromolecules and the physiological state of the seeds: tRNA, ribosomal RNA, and poly(A)-containing RNA (probably mRNA) as well as proteins are synthesized at the same rate in both dormant and thermodormant embryos as in after-ripened embryos. The protein patterns of dormant and after-ripened embryos are similar, as shown by electrophoresis and electrofocusing of double-labeled proteins. The level of DNA synthesis, measured as [³H]thymidine incorporation, may, on the other hand, indicate the physiological activity of the seeds: [³H]Thymidine is incorporated at a high rate in after-ripened embryos only and remains at a low level in dormant or thermodormant embryos. This correlation is, however, observed only in the axes. DNA synthesis in the cotyledons does not show any relation to the developmental stage of the seeds. These results are discussed in relation to the regulation of dormancy and after-ripening of seeds.     

Release of heat pretreatment-induced dormancy in lettuce seeds by ethylene or cytokinin in relation to the production of ethylene and the synthesis of 1-aminocyclopropane-1-carboxylic acid during germination

The germination of lettuce (Lactuca sativa L.) seeds was greatly reduced when the seeds were heated at 97°C for 30 h prior to imbibition. This dormancy was effectively released when ethylene (1–100 ppm) or benzyladenine (BA) (0.005–0.05 mM) was applied during the imbibition period. Ethylene was not required during the early part of imbibition, but was essential during the period immediately prior to radicle protrusion. Treatment with 1-aminocyclopropane-1-carboxylic acid (ACC) (0.1–10 mM) stimulated germination, but was not as effective as ethylene or cytokinin treatment. During the germination of nondormant lettuce seeds, ethylene production increased rapidly and reached a peak at 24 h, which coincided with the emergence of the radicle, and then declined; the level of ACC increased as ethylene production rate increased, but remained at a high level after radicle protrusion. In heat-pretreated dormant lettuce seeds, the increases in percent germination, ethylene production, and ACC levels were all delayed and lower than those of nondormant seeds, and these increases were accelerated by treatment with ethylene or cytokinin.     

Untersuchungen über die Regulation der DNS-Synthese während des Aktivitätswechsels vonAgrostemma githago-Samen

The relationships between DNA synthesis and germination capacity ofAgrostemma seeds have been studied. Protein synthesis and RNA synthesis are activated at the very beginning of imbibition, whereas DNA synthesis starts in the second part of the imbibition phase. Agrostemma seeds inhibited by higher temperature (30° C), or aged seeds with a low germination capacity are characterized by a remarkably reduced protein synthesis. DNA synthesis is also reduced. The inhibition of protein-synthesis ofAgrostemma embryos fed with cycloheximid or actinomycin D causes a depression of DNA synthesis. These results indicate that the initiation of DNA synthesis of imbibingAgrostemma seeds depends on the synthesis of special proteins. Abscisic acid inhibits growth as well as DNA synthesis of isolatedAgrostemma embryos. Mitomycin inhibits germination and DNA synthesis to the same extent. Dormant seeds with an undiminished intensity of protein synthesis also show a reduced incorporation of³H-thymidine in DNA. We suggest that DNA synthesis of imbibed seeds, which is a necessary prerequisite for the radicle protrusion, is involved in the mechanism of afterripening of theAgrostemma seeds.     

Does dormancy protect seeds against attack by the pathogenic fungus <Emphasis Type="Italic">Fusarium tricinctum</Emphasis> in a semiarid grassland of Northwest China?

Aims

Soil fungal pathogens can result in the failure of seedling establishment, but the effects of fungicide applications on seed/seedling survival have differed among studies. We assumed that the variation may relate to seed dormancy/germination characteristics and hypothesized that nondormant germinating seeds are more likely to be killed by fungal pathogens than dormant seeds.

Methods

Dormant and nondormant seeds of Stipa bungeana and Lespedeza davurica were inoculated with a pathogenic fungus Fusarium tricinctum under laboratory and field conditions. The outcomes of seed/seedling fate and other parameters were evaluated.

Results

In the laboratory, nondormant seeds inoculated with F. tricinctum developed white tufts of mycelium on the radicles of germinating seeds causing them to quickly die, but dormant seeds remained intact. In contrast, in the field inoculation with F. tricinctum did not cause higher mortality of nondormant than dormant seeds but resulted in higher percentages of seedling death before they emerged from soil than the controls.

Conclusions

Our results suggest that dormancy protects seeds from being attacked by some pathogens by preventing germination, but the protection is lost once germination has commenced. Further study involving various plant species with more seeds is needed to assess the generality of this pathogen-seed interaction hypothesis.

     

Proteinbiosynthesen in dormanten und nachgereiften embryonen und samen von Agrostemma githago

Seed germination of Agrostemma githago is prevented by inhibitors of protein and RNA synthesis. Thus protein as well as RNA synthesis are essential prerequisites for germination. Early protein synthesis of Agrostemnia embryos can be completely inhibited by cycloheximide and cordycepin. During the aging of seeds there is a considerable decrease in germination capacity and protein synthesis. In dormant and afterripened embryos of Agrostemma githago¹⁴C-leucine and ¹⁴C-uracil are incorporated in protein and RNA respectively with nearly the same intensity, whereas RNA and protein synthesis of dormant seeds and embryos starts earlier than in those subjected to afterripening. ³H-uracil-labelled RNA from dormant and afterripened embryos are able to hybridize on oligo-dT-cellulose to the same extent. There is a similarity in the protein pattern of dormant and afterripened embryos revealed by electrophoresis in polyacrylamide gels of double-labelled proteins. According to these results dormancy of Agrostemma githago is not caused by a general but by a specific metabolic block.     

Studies in Wild Oat Seed Dormancy: I. THE ROLE OF ETHYLENE IN DORMANCY BREAKAGE AND GERMINATION OF WILD OAT SEEDS (AVENA FATUA L.)

Seed of Avena fatua were shown to exhibit a characteristic loss of dormancy during dry storage at 25 C, whereas similar seed stored at 5 C maintained dormancy. 2-Chloroethylphosphonic acid was shown to increase germination of partly dormant seed imbibed under certain temperature regimes; a similar effect could not be established for fully dormant or fully nondormant seed. Using gas-liquid chromatography, natural ethylene levels were followed during imbibition of fully dormant and nondormant seed. A large peak in production was observed in the period prior to radicle emergence in the case of the nondormant seed. Measurements of ethylene production taken at 15 C, following periods of after-ripening in moist soil at either 5 or 25 C, indicated that endogenous production was unlikely to be a main cause of dormancy breakage in this species. The possibility that endogenous ethylene could play a role in natural dormancy breakage in aged seeds is discussed. The practical possibilities of 2-chloroethylphosphonic acid as a dormancy breaking agent in a field situation are outlined.     

Protein and oil concentration of soybean seed cultured in vitro using nutrient solutions of differing glutamine concentration

Oil and protein are the most valuable components of soybean seed. Evidence indicates that growth and composition of soybean seed are controlled by supplies of carbon and nitrogen provided by the maternal plant to the seed, but it is difficult experimentally to control and quantify the precise amount of carbon and nitrogen provided to the seed by the whole plant. To examine whether oil and protein concentrations are affected by the supply of nitrogen to the seed, immature soybean seeds (Glycine max cv. Williams 82) were grown in vitro in nutrient solutions containing 20, 40, 60 or 80 mM of glutamine. The seeds were incubated in Erlenmeyer flasks for 8 days at 25°C. The rate of dry matter accumulation changed from 7.2 to 8.3 mg seed⁻¹ day⁻¹ as the glutamine concentration increased from 20 to 80 mM but the differences were not significant (P 0.05). Seed protein concentration increased as glutamine concentration increased from 294 mg g⁻¹ at 20 mM glutamine to as high as 445 mg g⁻¹ at 80 mM glutamine. Typical in vivo protein concentration of mature soybean seeds is about 400 mg g⁻¹. Oil and protein concentrations were negatively correlated (r²= 0.44), which indicates that oil and protein synthesis are interrelated. Protein synthesis was favoured over oil synthesis when nitrogen became more abundant. The seeds used in this study clearly demonstrated a capacity to respond to nitrogen availability with changes in seed protein concentration.     

The induction of protein X in DNA repair and cell division mutants of Escherichia coli

Certain temperature-sensitive Escherichia coli cell division mutants and DNA repair mutants were treated in several ways to alter DNA synthesis or cell division. The bacteria were pulsed with [³⁵S]methionine; then membrane proteins were prepared and examined using sodium dodecyl sulfate/polyacrylamide slab gels. Autoradiography was performed on the slab gels so that the rate of synthesis of protein X could be determined by microdensitometry.Several changes in the rate of synthesis of the 40,000 molecular weight protein X were found in the different mutants. The wild-type (rec⁺ and lex⁺) strains synthesized protein X in response to DNA synthesis inhibition. However, neither recA^? strains nor lex^? strains synthesized protein X.Both the filament forming, temperature-sensitive mutants tif^? and tsl^? (which was derived from lex^?) synthesized protein X when DNA synthesis was inhibited, but at rates different from the wild-type strains. Moreover, these strains also produced protein X at their non-permissive temperature, even though DNA synthesis was not inhibited. In the tif^? mutant, the rate of synthesis of protein X was influenced by the addition of nucleic acid precursors.A double mutant tsl^?recA^? produced protein X when DNA synthesis was inhibited, or at the non-permissive temperature (although DNA synthesis was normal). This was the only strain carrying a recA^? mutation capable of synthesizing protein X.From these results it is suggested that the genes lex, recA and tif comprise a system that controls DNA repair and limits DNA degradation by the recBC nuclease. The inducer of this control system might be a DNA degradation product.     

Inhibition of pregerminative processes in dormant and thermodormant Agrostemma githago seeds

Imbibed non-dormant seeds do not germinate immediately after completion of water uptake and reactivation of their metabolism. During the lag-period the seeds apparently undergo processes which are essential for germination. The extent to which these pregerminative processes occur in dormant seeds (freshly harvested seeds) and thermodormant seeds (afterripened seeds imbibed at a supra-optimal temperature) of Agrostemma githago was determined. The pregerminative processes were inhibited almost completely in dormant seeds, but only to 50% or less in thermodormant seeds. When seeds were progressing through the pregerminative processes, the axes showed a higher rate of protein synthesis than axes of blocked seeds. However, this increased rate of protein synthesis was a late event and neither necessary nor sufficient for germination.     

Abscisic Acid levels and seed dormancy

Dormant seeds from Fraxinus species require cold-temperature after-ripening prior to germination. Earlier, we found that abscisic acid (ABA) will inhibit germination of excised nondormant embryos and that this can be reversed with a combination of gibberellic acid and kinetin. Using Milborrow's quantitative “racemate dilution” method the ABA concentration in 3 types of Fraxinus seed and pericarp were determined. While ABA was present in all tissues, the highest concentration was found in the seed and pericarp of dormant F. americana. During the chilling treatment of F. americana the ABA levels decreased 37% in the pericarp and 68% in the seed. The ABA concentration of the seed of the nondormant species, F. ornus, is as low as that found in F. americana seeds after cold treatment. Experiments with exogenously added ABA solutions indicate that it is unlikely that the ABA in the pericarp functions in the regulation of seed dormancy. However, the ABA in the seed does seem to have a regulatory role in germination.     

Storage Protein Synthesis during Oat (Avena sativa L.) Seed Development

Oat (Avena sativa L.) seeds harvested at 2-day intervals from anthesis to maturity were tested for their ability to incorporate [³⁵S]sulfate into protein. Incorporation of [³⁵S]sulfate into TCA-insoluble material began 2 to 4 days postanthesis (DPA), reached a peak 14 to 16 DPA, and was barely detectable by 24 DPA. Incorporation of label into globulin was parallel to total protein accumulation, and averaged about 85% of the total protein synthesis. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of total protein extracted from developing seeds indicated that some polypeptides coinciding with the α and β globulin subunits were present 2 to 4 DPA, but the full complement of globulin polypeptides was not present until 10 DPA. Immunoprecipitation of in vivo labeled seed extracts showed that globulin polypeptides and the 59 kilodalton precursor were present at early stages of development (4 DPA). Quantitation of dot blot analysis, using an oat globulin cDNA clone as a probe, indicated that one species of oat globulin mRNA was most abundant 15 DPA, which is during the peak time of storage protein synthesis.     

Potassium-Dependent Increase in RNA and Protein Synthesis in the Early Phase of Incubation of the Thermodormant Phacelia tanacetifolia Seeds

The seeds of Phacelia tanacetifolia Benth. cv Bleu Clair incubated at 30°C in the dark did not germinate and did not activate K⁺ uptake capacity. The administration of 1 millimolar K⁺ in the early phase of incubation stimulated RNA and protein synthesis. The possible role of K⁺ in promoting the marcromolecular syntheses during the early phase of germination is discussed.     

Reprogramming of Protein Synthesis from a Developmental to a Germinative Mode Induced by Desiccation of the Axes of Phaseolus vulgaris

Immature seeds of Phaseolus vulgaris cv Taylor's Horticultural removed from the pod at 32 days of development do not germinate unless first subjected to desiccation. Our results show that premature drying not only redirects metabolism from a developmental to a germination program but it does so permanently, thus effecting an irreversible switch. This is shown by in vitro protein synthesis, and analysis of poly(A)⁺ mRNA with a cDNA probe specific for phaseolin message. For example, the pattern of proteins synthesized in vitro by the mRNA fraction from fresh and prematurely dried axes show strong similarities; on the other hand, the mRNA population from rehydrated axes code for a different set of proteins. Also, the message for phaseolin is preserved following the normal maturation process and premature desiccation of seeds. Following rehydration of immature seeds at the desiccation-tolerant stage, this message is no longer detectable in the axes.     

Possible Involvement of the Alternative Respiration System in the Ethylene-stimulated Germination of Cocklebur Seeds

Respiration of nondormant upper cocklebur (Xanthium pensylvanicum Wallr.) seeds was enhanced by exogenous C₂H₄, proportionally to the concentration of C₂H₄ and the duration of presoaking of the seeds. Benzohydroxamic acid (BHM) and salicylhydroxamic acid (SHM), inhibitors of alternative respiration, inhibited both the germination of nondormant lower cocklebur seeds and the respiration of the upper seeds presoaked for periods of 12 to 30 hours. Both the growth and respiration of axial and cotyledonary tissues were also inhibited by BHM. Moreover, BHM inhibited both the C₂H₄-induced germination of the upper seeds and their C₂H₄-stimulated respiration; the inhibition occurred only with concomitant addition of C₂H₄ and BHM. The respiration of seeds with a secondary dormancy induced by presoaking for prolonged periods was markedly stimulated by C₂H₄ but not suppressed by BHM. It was suggested that the alternative respiration system may be involved in the normal germination process of cocklebur seeds, secondary dormancy may result from its inactivation, and C₂H₄ may exert its germination-promoting action by stimulating the alternative respiration. The effects of BHM and SHM can suggest but not prove the involvement of the alternative respiration in seed germination.     

The effect of dormancy on the heat shock response in gladiolus cormels

Cormels of Gladiolus X gandavensis Van Houtte respond to heat shock by an induced synthesis of heat shock proteins. Synthesis of some of the non-heat shock proteins is concomitantly reduced. The ability of dormant cormels to synthesize heat shock proteins (hsps) and to repress the synthesis of non-hsps is greater than that of nondormant ones. A hsp of apparent molecular weight 68 kilodaltons is synthesized only in dormant cormels or in cormels that lost their dormancy after long storage at 25°C. The synthesis of hsps at 40°C, but not at 25°C, is promoted by abscisic acid in nondormant cormels. Methionine incorporation into hsps declines after a 4-hour incubation period at 40°C. Induction of hsps is stronger if exposure to extreme temperature is done gradually.     

GERMINATION ECOLOGY OF SEDUM PULCHELLUM MICHX. (CRASSULACEAE)

Factors controlling the timing of seed germination were investigated in the small succulent winter annual Sedum pulchellum Michx. (Crassulaceae) in its natural habitat on unshaded limestone outcrops in northcentral Kentucky. At maturity in early July the dormant seeds are not dispersed but are retained in the fruits on the standing dead plants until September and October. Many, but not all, of the seeds afterripen in the fruits during summer, and at the time of dispersal some of them are dormant and some are nondormant. Germination and annual population establishment occur in September and October from seed reserves that have been in the soil for one or more years and from seeds produced in the current year. Germination of nondormant seeds may be prevented in autumn by lack of the appropriate combination of environmental factors including light, temperature and soil moisture in the seed's microsite. The effect of low winter temperatures on ungerminated seeds in the population is to induce nondormant seeds into secondary dormancy and to prevent afterripening of dormant seeds. Thus, in spring all the seeds in the population's seed reserve are dormant. During spring and summer some of these seeds afterripen, and they germinate in autumn when, and if, germination requirements are fulfilled.     

MECHANISM OF SEED DORMANCY IN AMBROSIA ARTEMISIIFOLIA

Dormancy in Ambrosia artemisiifolia seeds was broken by 8 weeks of stratification. Germination of nondormant seeds was greater in light than in continuous darkness. Embryos of freshly harvested seeds were nondormant. Leaching and scarification did not stimulate germination of the dormant seeds. Exogenous gibberellin (GA₃) slightly increased germination of intact dormant seeds, and the effect was greatly increased by scarification. Germination was greater in the light in both tests. Exogenous indoleacetic acid did not stimulate germination of dormant seeds. Endogenous gibberellin and auxin content increased during stratification, and there was also a significant increase in GA during post-stratification at a favorable germination temperature. Inhibitors in the dormant seeds decreased during stratification and post-stratification. The high concentration of chlorogenic acid present in dormant seeds increased slightly during stratification. An unknown phenol very similar to chlorogenic acid in fluorescence and U.V. absorption significantly increased after 2 weeks of stratification. A significant decrease in the concentration of a second unidentified phenol occurred after 2 weeks of stratification. It is proposed that dormancy in Ambrosia artemisiifolia may be controlled by an inhibitor-promoter complex. The dormant seed is characterized by high inhibitor and low promoter levels. In the nondormant seed the balance was shifted to favor the promoter. Evidence suggests that the inhibitor involved may be abscisic acid and the promoters may be gibberellin and auxin. The content of auxin may be partially controlled by the concentration of phenols.