Two C2H4-producing systems in cocklebur seeds

Summary C₂H₄ production of the embryonic axes and cotyledons excised from dormant and non-dormant cocklebur (Xanthium pennsylvanicum Wallr.) seeds was examined in relation to ambient O₂ tensions. There were two kinds of C₂H₄-producing systems, quasi-anaerobic and aerobic, in both organs. Regardless of the organ, the former activity was high in the dormant state and, particularly in axes, declined with after-ripening. On the other hand, the latter activity was almost insignificant in the dormant state, but increased with release from dormancy and the non-dormant axes exclusively produced C₂H₄ through this system. In the cotyledons, however, the former was still predominant even after they were fully after-ripened. Thus, the C₂H₄-producing systems were different in the seed organ and in the dormancy state.     

Protein Synthesis in Dormant and Non-Dormant Cocklebur Seed Segments

Using the axial and cotyledonary segments of lower cocklebur (Xanthium pensylvanicum Wallr.) seeds, protein synthesis as shown by incorporation of radioactive leucine was examined in relation to their dormant status. During the first 9 h of water imbibition, the protein synthesis was higher in the dormant axes than in the non-dormant, after- ripened ones. When imbibed for more than 12 h non-dormant axes had a higher activity than dormant ones. This was also the case with the cotyledonary segments. Cyctoheximide, an inhibitor of protein synthesis, blocked protein synthesis in the axial tissue regardless of its dormant status, and thereby inhibited germination of the non-dormant seeds. In the dormant seeds, however, cycloheximide at 3 mM slightly stimulated germination without stimulating the C₂H₄ production. Based on these results, it is suggested that in cocklebur seeds there may be some proteinaceous system which is involved in the maintenance of dormancy.     

Induction and release of secondary seed dormancy in genetically pure lines of Avena fatua

Induction and release of secondary dormancy in genetically pure dormant (AN-51, Mont 73) and non-dormant (CS-40, SH-430) lines of wild oat ( Avena fatua L.) were studied. These lines differed with regard to the optimal period of anaerobiosis necessary for induction of dormancy, and/or the degree (% of seeds acquiring dormancy) and duration of the dormancy induced. Secondary dormancy could be induced more effectively in the after-ripened seeds of dormant lines than in the non-dormant lines, where only a short-term dormancy could be induced (in 5–7 week-old-seeds). Higher anaerobiosis temperatures were more effective in inducing dormancy in all lines studied. Thus, as with primary dormancy, wild oat biotypes exhibit genetic variability in their secondary dormancy behaviour and factors like temperature can modify the expression of this trait.
The germination stimulants kinetin, isopentenyl adenine, sodium azide, potassium nitrate, ethanol and substituted phthalimides, which break primary dormancy in wild oats, stimulated germination of secondarily dormant seeds (line AN-51). Since these chemicals are structurally diverse, primary and secondary dormancies appear to be similar in part in their regulation.
Salicylhydroxamic acid, an inhibitor of cyanide-insensitive (alternative) respiration, did not inhibit: 1, spontaneous release of secondary dormancy in the line SH-430; and 2, stimulation of germination of secondarily dormant AN-51 seeds by various chemicals (except azide), suggesting that this respiratory pathway is not necessary for the release of induced dormancy.     

The physiological basis of seed dormancy in Avena fatua. IX. Characterization of two dormancy states

The dormancy-breaking effect of several known germination promoters was studied in 9 genetically pure lines of Avena fatua L. during a period of controlled after-ripening. Changes in the germination response show at least two dormancy states in the caryopses of these lines. The first state is overcome by a short period of after-ripening and is insensitive to nitrate and azide, while the second state is more persistent and is sensitive to nitrate and azide. Both states are sensitive to gibberellic acid (OA,) and ethanol. In the most dormant lines a third ethanol-insensitive dormancy state is present. The duration of both major dormancy states was related to several environmental factors influencing plant growth and seed storage. Duration was increased in caryopses produced from plants matured under low temperatures (15°C) and decreased in caryopses produced from plants matured under high temperatures (25°C). Duration was increased in caryopses after-ripened under low temperatures (4°C) and decreased in caryopses after-ripened under high temperatures (45°C). Dehulling the seeds prior to after-ripening reduced the duration of both major dormancy states. The multiple state dormancy system and its environmentally induced plasticity are discussed with reference to previous explanations of the dormancy mechanism in wild oats.     

Wheat seed proteins regulated by imbibition independent of dormancy status

Seed dormancy is an important trait in wheat (Trticum aestivum L.) and it can be released by germination-stimulating treatments such as after-ripening. Previously, we identified proteins specifically associated with after-ripening mediated developmental switches of wheat seeds from the state of dormancy to germination. Here, we report seed proteins that exhibited imbibition induced co-regulation in both dormant and after-ripened seeds of wheat, suggesting that the expression of these specific proteins/protein isoforms is not associated with the maintenance or release of seed dormancy in wheat.     

Variation between Emex australis populations in seed dormancy/non-dormancy cycles

Abstract Seeds of four Western Australian accessions of Emex australis were stored outside and their germinability tested at 4–6 week intervals for 22 months. Accessions showed cyclical behaviour in germinability, with peaks in autumn/early winter and troughs in spring/summer. There was considerable variation between accessions in dormancy/non-dormancy cycles, but when plants from accessions with contrasting cycles were grown in a common environment, the cycles shown by their seeds were virtually identical. Thus, the parameters of seed dormancy/non-dormancy cycles in E. australis appear to be under environmental control. Only part of an E. australis seed population demonstrated cyclical changes in dormancy status, in contrast to many other annual species whose entire seed populations undergo such changes. Populations also contained individuals which were either continuously dormant or continuously non-dormant following a period of after-ripening. The latter seeds give E. australis the flexibility to recruit opportunistically after summer rainfall events in mediterranean-climate environments.     

Role of reactive oxygen species in the regulation of Arabidopsis seed dormancy

Freshly harvested seeds of Arabidopsis thaliana, Columbia (Col) accession were dormant when imbibed at 25°C in the dark. Their dormancy was alleviated by continuous light during imbibition or by 5 weeks of storage at 20°C (after-ripening). We investigated the possible role of reactive oxygen species (ROS) in the regulation of Col seed dormancy. After 24 h of imbibition at 25°C, non-dormant seeds produced more ROS than dormant seeds, and their catalase activity was lower. In situ ROS localization revealed that germination was associated with an accumulation of superoxide and hydrogen peroxide in the radicle. ROS production was temporally and spatially regulated: ROS were first localized within the cytoplasm upon imbibition of non-dormant seeds, then in the nucleus and finally in the cell wall, which suggests that ROS play different roles during germination. Imbibition of dormant and non-dormant seeds in the presence of ROS scavengers or donors, which inhibited or stimulated germination, respectively, confirmed the role of ROS in germination. Freshly harvested seeds of the mutants defective in catalase (cat2-1) and vitamin E (vte1-1) did not display dormancy; however, seeds of the NADPH oxidase mutants (rbohD) were deeply dormant. Expression of a set of genes related to dormancy upon imbibition in the cat2-1 and vet1-1 seeds revealed that their non-dormant phenotype was probably not related to ABA or gibberellin metabolism, but suggested that ROS could trigger germination through gibberellin signaling activation.     

Proteomic analysis of seed dormancy in Arabidopsis

The mechanisms controlling seed dormancy in Arabidopsis (Arabidopsis thaliana) have been characterized by proteomics using the dormant (D) accession Cvi originating from the Cape Verde Islands. Comparative studies carried out with freshly harvested dormant and after-ripened non-dormant (ND) seeds revealed a specific differential accumulation of 32 proteins. The data suggested that proteins associated with metabolic functions potentially involved in germination can accumulate during after-ripening in the dry state leading to dormancy release. Exogenous application of abscisic acid (ABA) to ND seeds strongly impeded their germination, which physiologically mimicked the behavior of D imbibed seeds. This application resulted in an alteration of the accumulation pattern of 71 proteins. There was a strong down-accumulation of a major part (90%) of these proteins, which were involved mainly in energetic and protein metabolisms. This feature suggested that exogenous ABA triggers proteolytic mechanisms in imbibed seeds. An analysis of de novo protein synthesis by two-dimensional gel electrophoresis in the presence of [(35)S]-methionine disclosed that exogenous ABA does not impede protein biosynthesis during imbibition. Furthermore, imbibed D seeds proved competent for de novo protein synthesis, demonstrating that impediment of protein translation was not the cause of the observed block of seed germination. However, the two-dimensional protein profiles were markedly different from those obtained with the ND seeds imbibed in ABA. Altogether, the data showed that the mechanisms blocking germination of the ND seeds by ABA application are different from those preventing germination of the D seeds imbibed in basal medium.     

Developmental Differences Between Germinating After-ripened and Dormant Excised Avena fatua L. Embryos

Based on physiological and molecular differences associatedwith the germination of after-ripened and dormant caryopsesand excised embryos, it has been hypothesized that various methodsof after-ripening are the only treatments that facilitate thetransition of dormant wild oat embryos to a non-dormant state.To further investigate this hypothesis, analytical methods wereused to evaluate physical and temporal changes associated withgermination and subsequent growth of after-ripened and dormantexcised embryos (AR-embryos and D-embryos, respectively) inducedto germinate with fructose (Fru) and/or gibberellic acid (GA).While chemical treatments of Fru, GA, and Fru+GA have littleeffect on the germination and short-term growth of AR-embryos,they do induce germination of D-embryos. Growth following germinationof D-embryos varied according to treatment with the combinationof Fru+GA inducing the greatest growth over the duration ofthe experiment. Even considering differences in the time tocomplete germination, growth of D-embryos was not comparablewith that of AR-embryos. This provides physical evidence thatchemical treatments induce germination without fulfilling therequirements for normal after-ripening-enhanced germination/growth,and indicates that fructose and/or gibberellic acid do not removethe dormancy-block or rate limiting step in the same manneras after-ripening. Avena fatua ; after-ripening; dormancy; fructose; germination; gibberellic acid; wild oats     

The physiological basis of seed dormancy in Avena fatua III. Action of nitrogenous compounds

Sodium nitrate and nitrite (50–100 m M ) induced germination in three out of four genetically pure dormant lines of Avena fatua L. The sensitivity to these treatments was low immediately ater harvest and increased markedly after six months of dry after-ripening. The observation that a fourth dormant line failed to respond suggests at least two metabolic blocks may be involved in expression of dormancy. An inhibitor of gibberellin biosynthesis, 2-chloroethyl trimethylammonium chloride, completely inhibited the dormancy-breaking effect by nitrate and nitrite, indicating a requirement for gibberellin biosynthesis. Among reduced nitrogenous compounds, ammonium chloride and urea failed to break dormancy in all partly after-ripened lines, suggesting that nitrate and nitrite may induce germination through their ability to act as electron acceptors. The sensitivity to all nitrogenous compounds tested increased with the length of after-ripening indicating that the depth of the second dormancy block amy decrease with the time of after-ripening. Other reduced nitrogenous compounds, thiourea and hydroxylamine hydrochloride, promoted some germination in the least dormant, partially after-ripened lines. The function of these compounds as electron acceptors and their similarity in activity to the cytochrome oxidase inhibitor, sodium azide, is discussed with reference to dormancy and the possible involvement of the alternative pathway of respiration.     

Secondary dormancy in Avena fatua: Induction and characteristics in genetically pure dormant lines

The induction of secondary dormancy in caryopses of genetically pure dormant lines of Avena fatua L. is described. Seeds harvested from mature plants were after-ripened under controlled conditions (26°C, 25% relative humidity) until fully non-dormant. Secondary dormancy was then induced into these caryopses by incubation on moist filter papers in an aspirated nitrogen atmosphere at 20°C over periods from 3 h to 14 days. These caryopses failed to germinate when returned to an aerobic environment. The dose-response curves for gibberellic acid, sodium azide, sodium nitrite, sodium nitrate and ethanol show that all of these treatments can overcome the induced secondary dormancy. Drying increased the sensitivity of secondary dormant caryopses to these treatments. These treatments overcame secondary dormancy at all times, indicating the presence of only one of the two known blocks to germination that exist during primary dormancy. Similarities between primary and secondary dormancy in A. fatua are discussed.     

No contribution of the pentose phosphate pathway in dormancy-breaking of cocklebur seeds

The role of the oxidative pentose phosphate (PP) pathway in the dormancy-breaking of cocklebur (Xanthium pennsylvanicum Wallr.) seeds was investigated. D-[1-¹⁴C]-glucose or D-[6-¹⁴C]-glucose was fed to dormant and non-dormant lower seeds or to their axial or cotyledonary segments which were imbibed for different durations, and C₆/C₁ ratios of respired ¹⁴CO₂ as an index of the PP pathway activity were calculated. Contrary to expectation, there was no significant difference in the C₆/C₁ ratios between the dormant and non-dormant seeds or segments during a water imbition period of 24 h, although the PP pathway actually operated already in an early stage of water imbibition. Also concerning the activities of G6PDH and 6PGDH, the key enzymes of this pathway, no difference between the dormant and non-dormant seeds was found. It was thus concluded that, unlike other seeds, there is no contribution of the PP pathway to the regulation of dormancy of the cocklebur seed.     

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.     

Changes in protein synthesis during the development of Agrostemma githago seeds

The transition from the growth to the maturation phase in developing seeds of Agrostemma githago L. was found to coincide with major changes in the rate of protein synthesis, the kinds of proteins synthesized, and the composition of the non-proteinbound amino acid pool. Coincident changes were observed in viability and the ability to withstand desiccation. Desiccated mature Agrostemma seeds are dormant and need at least three months of after-ripening. In imbibed dormant and after-ripened seeds no synthesis of storage proteins was observed with the exception of one particular set of storage proteins. Dormant and after-ripened seeds synthesized the same kinds of proteins during early imbibition, indicating an almost identical metabolic state which differs considerably from that of developing seeds.     

Sodium nitroprusside, cyanide, nitrite, and nitrate break Arabidopsis seed dormancy in a nitric oxide-dependent manner

The seeds of many plant species are dormant at maturity and dormancy loss is a prerequisite for germination. Numerous environmental and chemical treatments are known to lessen or remove seed dormancy, but the biochemical changes that occur during this change of state are poorly understood. Several lines of research have implicated nitric oxide (NO) as a participant in this process. Here, we show that dormant seeds of Arabidopsis thaliana (L.) Heynh. will germinate following treatment with the NO donor sodium nitroprusside (SNP), cyanide (CN), nitrite or nitrate. In all cases, the NO scavenger c-PTIO effectively promotes the maintenance of seed dormancy. c-PTIO does not, however, inhibit germination of fully after-ripened seeds, and c-PTIO does not interact directly with nitrite, nitrate or CN. We also show that volatile CN effectively breaks dormancy of Arabidopsis seeds, and that CN is the volatile compound in SNP that promotes dormancy loss. Our data support the hypothesis that NO is a signaling molecule that plays an important role in the loss of seed dormancy.