Hours Of Imbibition Research Articles

Adenosine triphosphate and adenylate energy charge in relation to dormancy of wild oat seeds

Adenosine phosphates, expressed as ATP and energy charge levels, were compared in genetically identical dormant and nondormant caryopses of wild oat (Avena fatua L.). Air-dry nondormant caryopses had a larger pool of ATP and a higher energy charge than dormant caryopses. During the early hours of imbibition of water by dormant caryopses, ATP levels and energy charge increased indicating they are not limiting factors for germination. Moreover, during the induction of germination in dormant caryopses by gibberellic acid (GA3), changes in energy charge were similar to those observed on water, indicating that GA3 does not overcome dormancy simply by elevating energy charge levels. In a second study, ATP and energy charge were measured in initially dormant caryopses incubated under conditions of temperature that either removed or retained dormancy. Both embryo and endosperm extracts demonstrated correlations between levels of ATP or energy charge with temperature of incubation but not with the state of dormancy. The conclusions drawn from both lines of investigation do not support the hypothesis that release of dormancy is related to the metabolism of adenosine phosphates in caryopses of wild oat.

Newly Synthesized mRNA Is Translated during the Initial Imbibition Phase of Germinating Maize Embryo

RNA synthesis is activated in the cells of the plant embryo very soon after the start of seed imbibition. We previously reported that mainly heterogeneous nuclear RNA is synthesized in the radicle of Zea mays embryo during the first hours of germination. The present study was undertaken in order to detect the time of appearance of the newly synthesized messenger RNA in the polysomes of germinating maize axes.Free polysomes were prepared from embryonic axes rehydrated for 2 hours in the presence of radioactively labeled uridine. These polysomes were shown to be labeled and to contain labeled particles sedimenting, after dissociation with EDTA, in the 10S to 40S region of a sucrose gradient. The labeled polysomal RNA migrates heterogeneously in a gel with a mean size corresponding to about 16S, and 60% of these molecules are polyadenylated.The data indicate that the newly synthesized RNA associated with the polysomes after 2 h of germination consists of messenger RNA molecules. Analysis of the polysomes prepared 0.5 and 1 h after the start of imbibition suggests that translation of the newly synthesized messenger RNA probably occurs within the 1st hour of imbibition of the isolated axis, thus well before the completion of the initial water uptake.

Appearance and Disappearance of Cyanide-Resistant Respiration in Vigna mungo Cotyledons during and following Germination of the Axis

Mitochondrial preparations isolated from black gram (Vigna mungo L.) cotyledons exhibited cyanide-resistant respiration which was of mitochondrial origin. The appearance and the disappearance of this alternative respiration took place during and following imbibition. During the first 6 hours of imbibition, the respiration was completely inhibited by cyanide, but after this time the alternative respiration markedly developed, reaching a maximal cyanide-resistance 12 to 16 hours after the start of imbibition. Subsequently, the alternative respiration gradually disappeared. The actions of cycloheximide and chloramphenicol indicated that the appearance was dependent on cytoplasmic protein synthesis and that the disappearance depended on both cytoplasmic and mitochondrial protein synthesis. The alternative pathway contributed to state 4 respiration, but not to state 3 respiration, in mitochondria from 1-day-old cotyledons. On day 3, it contributed to neither state 3 nor state 4.

Influence of Seed Quality on Ethylene Production of Germinating Snap Beans

Abstract Deterioration of snap bean (Phaseolus vulgaris L.) seed quality during accelerated aging at 42°C and 100% relative humidity was accompanied by a decline in germination, radicle emergence, hypocotyl length, and ethylene production. Field emergence of 5 seed lots had a highly significant correlation with ethylene production rates when measured after 22 to 23.5 hours of imbibition at 25°. Seed lots that produced low levels of ethylene emerged poorly in the field. Results indicate that determination of ethylene production of imbibed seeds might be a useful method for detecting changes in seed vigor.

Characterization of Solute Efflux from Dehydration Injured Soybean (Glycine max L. Merr) Seeds

Soybean (Glycine max L. Merr) seeds lose their tolerance of dehydration between 6 and 36 hours of imbibition. Soybean axes and cotyledons were excised 6 hours (tolerant of dehydration) and 36 hours (susceptible) after commencing imbibition and subsequently dehydrated to 10% moisture. Kinetics of the efflux of potassium, phosphate, amino acid, sugar, protein, and total electrolytes were compared in the four treatments during rehydration. Only slight differences were observed in the kinetics of solute efflux between the two cotyledon treatments dehydrated at 6 and 36 hours suggesting that the cotyledons may retain their tolerance of dehydration at this stage of germination. Several symptoms of injury were observed in the axes dehydrated at 36 hours. An increase in the initial leakage of solutes during rehydration, as quantified by the y-intercept of the linear regression line for solute efflux between 2 and 8 hours suggests an increased incidence of cell rupture. An increase in the rate of solute efflux (slope of regression line between 2 and 8 hours) from fully rehydrated axes was observed in comparison to axes dehydrated at 6 hours. The Arrhenius activation energy for potassium, phosphate, and amino acid efflux decreased and for protein remained unchanged. Both observations indicate an increase in membrane permeability in dehydration-injured tissue. Increasing the H(+) concentration of the external solution increased K(+) efflux from both control and dehydrated/rehydrated samples, increased sugar efflux from axes at 6 hours imbibition but decreased sugar efflux from axes at 36 hours imbibition, indicating changes in membrane properties during germination. The dehydration treatment did not alter the pattern of the pH response of axes dehydrated at 6 or 36 hours but did increase the quantity of potassium and sugar efflux from dehydration injured axes. These results are interpreted as indicating that dehydration of soybean axes at 36 hours of imbibition increased both the incidence of cell rupture during rehydration and altered membrane permeability of the rehydrated tissue.

Dehydration Injury in Germinating Soybean (Glycine max L. Merr.) Seeds

The sensitivity of soybean (Glycine max L. Merr. cv Maple Arrow) seeds to dehydration changed during germination. Seeds were tolerant of dehydration to 10% moisture if dried at 6 hours of imbibition, but were susceptible to dehydration injury if dried at 36 hours of imbibition. Dehydration injury appeared as loss of germination, slower growth rates of isolated axes, hypocotyl and root curling, and altered membrane permeability. Increased electrolyte leakage due to dehydration treatment was observed only from isolated axes but not from cotyledons, suggesting that cotyledons are more tolerant of dehydration. The transition from a dehydration-tolerant to a dehydration-susceptible state coincided with radicle elongation. However, the prevention of cell elongation by osmotic treatment in polyethylene glycol (-6 bars) or imbibition in 20 micrograms per milliliter cycloheximide did not prevent the loss of dehydration tolerance suggesting that neither cell elongation nor cytoplasmic protein synthesis was responsible for the change in sensitivity of soybean seeds to dehydration. Furthermore, the rate of dehydration or rate of rehydration did not alter the response to the dehydration stress.

Intracellular levels of polyadenylated RNA and loss of vigour in germinating wheat embryos

Polyadenylated-RNA (Poly(A)(+)RNA) levels have been studied during the germination of wheat embryos of high viability but differing vigour. In high-vigour embryos imbibed at 20°C the level of poly(A)(+)RNA falls dramatically over the first hour of imbibition, then remains constant up to 3 h of imbibition before increasing rapidly to a level similar to that found in the quiescent state by 7 h of imbibition. Median-vigour embryos imbibed at 20°C show similar changes in poly(A)(+)RNA content but the initial decrease and subsequent increase in poly(A)(+)RNA levels are less marked. On imbibition at 10°C, the poly(A)(+)RNA content in high-vigour embryos decreases to a lesser extent during the first hour than at 20°C and the level increases more slowly over the next 6 h than during the same time period at 20°C. The level of poly(A)(+)RNA in medianvigour embryos remains constant over the first 4 h of germination and then falls to a level of about half that found in quiescent high-vigour embryos. Polyacrylamide gel electrophoresis of total-RNA samples shows that the polyadenylic acid (poly(A)) sequences occur in RNA species ranging in size from 35-7S. Polyacrylamide gel electrophoresis of isolated poly(A) sequences demonstrates the presence of two size classes of poly(A) in quiescent embryos, but at 20°C a more heterodisperse pattern appears by 2 h of imbibition. At 10°C, two size classes of poly(A) persist throughout the period studied in both high- and median-vigour embryos, although in median-vigour embryos the ratio of larger: smaller poly(A)-tail sizes decreases more rapidly than in high-vigour embryos.

Anesthetic Effects on Secondary Dormancy and Phytochrome Responses in Setaria faberi Seeds

Seeds of giant foxtail (Setaria faberi Herm.) entered secondary dormancy after pretreatment in H(2)O at 35 degrees C. Pretreatment in 0.1 m ethanol, or several other substances with anesthetic properties, prevented secondary dormancy induction. Pretreatment in 0.5 m ethanol inhibited germination in darkness, but germination could be stimulated by a red irradiation. Germination was initially insensitive to light. Two separate responses are indicated. The first, affected by a variety of substances and low (0.1 m or less) concentrations of ethanol, is related to anesthetic effects and prevention of secondary dormancy. The second, induction of response to red irradiation, is caused by 0.5 m ethanol and some closely related substances. The anesthetic effect is accomplished within the first 8 hours of imbibition while the phytochrome induction effect required treatment for more than 24 hours. Both responses were lost if the 35 degrees C imbibition began in H(2)O. Involvement of cell membranes is suggested in the prevention of secondary dormancy by anesthetics.

Distinctive characteristics of RNA and protein synthesis in pea cotyledons at early stages of germination

The RNA and protein synthesis in pea cotyledons at early imbibition were studied. The results obtained show that RNA and protein synthesis are reactivated at the beginning of imbibition. The initial transcription is independent of protein synthesis, after 8 h synthesis RNA depends on translation by 44 – 52%. α-Amanitin (5μg ml−1) inhibits3H-uridine incorporation by 75%. It indicates the predominance of mRNA synthesis in early transcription. The protein synthesis during first hours of imbibition depends significantly (by 50%) on RNA synthesis. It means newly-made mRNA is involved in the translation. Protein synthesis, which is independent of transcription, is provided by preformed mRNA. This fraction of translation decreases rapidly during imbibition. Thus, protein synthesis in pea cotyledons depends mainly on newly-synthesized mRNA. Thein vivo synthesized polypeptides were fractionated by SDS-gel electrophoresis and visualized by autoradiography. The pattern of protein synthesis after 4 h of imbibition is very complex. Newly-synthesized polypeptides vary in m.m. (from 10 000 to 100 000), subcellular localization and solubility.14C-labelled polypeptides do not belong to subunits of storage proteins. The patterns of proteins, synthesized at different germination time (from 2 to 24 h), have much in common, but definite quantitative and qualitative changes in messenger population can be observed. During maturation of seed, the pattern of protein synthesis in pea cotyledons is changed rather quantitatively than qualitatively. Drastic changes in population of mRNA are observed in late embryogenesis. Patterns of proteins synthesized in late embryogenesis and early germination are similar.

Phospholipid metabolism of wheat grains: Enzymes of the CDP-diglyceride phospholipid biosynthetic pathway

Enzymes of the CDP-diglyceride pathway of phospholipid synthesis, CDP-diacylglycerol synthetase, CDP-diacylglycerol: glycerol 3-phosphate phosphatidyl-transferase and enzymes of phosphatidylserine formation were initially of relatively high specific activities in aleurone cells of wheat and declined upon imbibition. Enzyme activity of phosphatidylinositol synthesis was not detected in dry grains but was present upon imbibition. CDP-diacylglycerol: glycerol 3-phosphate phosphatidyltransferase shifted during imbibition from 85% of the activity in the supernatant of aleurone layers from dry seeds to 98% associated with large particle fractions after 36 hours of imbibition. Phosphatidylserine formation shifted from a dominant location in the 1,500 x g fraction in the dry seed to a predominantly “mitochondrial” location after 36 hours of imbibition. The subcellular distribution of CDP-diacylglycerol synthetase did not change appreciably upon imbibition from that of the dry seed, 75 to 80% of the activity was found in the supernatant. Only CDP-diacylglycerol: glycerol 3-phosphate phosphatidyltransferase showed increased specific activity late in the imbibition period. GA3 accelerated the decrease of already declining activities of the CDP-diglyceride enzymes and the changes in their patterns of distribution, augmented the activities of the phosphatidylinositol synthesizing enzyme, and both accelerated and augmented the increase in the activity of the enzyme of phosphatidylglycerol synthesis which occurred late in imbibition.

Oxidative Phosphorylation in Germinating Lettuce Seeds (Lactuca sativa) during the First Hours of Imbibition

Experiments with lettuce seeds during the first hours of imbibition showed that oxygen is necessary to sustain high adenine nucleotide ratios and consequently, energy charge values are higher than 0.8 as is usually the case in normally metabolizing tissues.The energy charge value (0.2) of dry seeds soaked in aerated water increased to normal values (0.8) within 30 minutes. The energy charge value of seeds imbibed under cyanide or nitrogen stayed at low values, about 0.3 for 30 minutes. Nitrogen and cyanide treatment of seeds imbibed in aerated water produced a decrease of energy charge to low values within 3 minutes. During the first minutes of imbibition, the oxygen uptake is cyanide-sensitive. The effect of the uncoupler carbonyl cyanide p-trifluoromethoxyphenyl hydrazone was not as clear-cut. However, results were obtained which agree with the occurrence of oxidative phosphorylation during the first hours of imbibition. These results indicate that a normal cytochromic pathway synthesizes ATP during the first minutes and hours following the imbibition of lettuce seeds.

Protein Synthesis and Phospholipids in Soybean Axes in Response to Imbibitional Chilling

The responses of two cultivars of soybean (Merr.) to a chilling treatment (4 C for first hour of imbibition) were compared. The germination of cv. Biloxi was unaffected by the treatment, while the germination of cv. Fiskeby was reduced. The phospholipid fatty acids of dry axes of the two cultivars were very similar, and, thus, could not be correlated with their responses to chilling. The fatty acid composition of chilling-tolerant Biloxi did not change over a subsequent 23-hour warm incubation, but there was a marked reduction in the unsaturated fatty acids of chilling-sensitive Fiskeby after 12 hours, which may be a symptom of deterioration. Protein synthesis in both cultivars was reduced by the chilling treatment. Redrying of Biloxi axes up to 18 hours after the onset of imbibition had no effect on their germination upon rehydration. Germination of Fiskeby axes was reduced by redrying after 8 hours of imbibition. After 7 months of dry storage of intact seeds, the sensitivity of the axes to chilling was retested. Biloxi axes had become chilling-sensitive, while the germination of Fiskeby axes was reduced to zero by the chilling treatment. A hypothesis is presented that imbibitional chilling sensitivity is an indication of reduced vigor, axes with a high vigor can tolerate the stress, while those without cannot.

Effects of temperature, red light and hormones on the accumulation of free amino acids in osmotically growth-inhibited embryonic axes of New York lettuce seeds.

Free amino acids accumulated in non-thermodormant embryonic axes of New York lettuce seeds from the 16th to the 24th hour of imbibition, even when germination was prevented with 0.3 m mannitol. The accumulation of free amino acids in the embryonic axes was affected by temperature, red light and hormones (gibberellic acid, kinetin and abscisic acid), and was correlated with the growth potential of the axes. These data are evidence that the accumulation of free amino acids in the embryonic axes is not the result of axes-growth, but one of its causes.

Modifications ultrastructurales de graines d'Allium cepa, hydratées ou non, soumises à de basses températures (−196 °C)

Osmic fixation was used in an ultrastructural study of Allium cepa seeds, both in the quiescent state and at the beginning of imbibition, in order to complete earlier observations made after glutaraldehyde fixation. In embryonic radicles, various groups of weakly stained lamellar structures looked like endoplasmic reticulum profiles. In these quiescent tissues, osmic fixation revealed many vesicles which appeared to be atypical elements of endoplasmic reticulum.In the first few hours of imbibition, reactive structures were more numerous and more diverse. Cisternae and vesicles of endoplasmic reticulum became typical looking, such as classically described in active tissues of other species.When Allium cepa seeds were treated with liquid nitrogen (−196 °C), induced structural alterations depended on the initial state of hydration. Lamellar systems underwent morphological changes. Osmic fixation showed the presence of numerous opaque globules throughout the cytoplasm. These globules seemed to derive from cisternae that appeared to have become fragmented during freezing. [Journal translation]

Respiratory pathways in aged soybean seeds

AbstractRespiratory activities in soybean seeds (Glvcine max (L.) Merr. cv. Chippewa 64) have been examined in the first minutes after water imbibition and after three hours of imbibition, using either particles or intact axes. Cyanide and azide were utilized as inhibitors of the cytochrome oxidase pathway of respiration, and SHAM inhibitions were interpreted as effects on the alternative pathway, since in unaged axes inhibitions by SHAM were obtained only when respiratory activity had either been restricted with inhibitors of the cytochrome oxidase pathway or expanded by an uncoupler (CCCP). From the experiments with these inhibitors, it is suggested that unaged seeds utilize both respiration pathways in the cotyledon but only the cytochrome pathway in the axis. Accelerated aging causes a marked deterioration of respiration, especially that through the cytochrome pathway, and there is an associated engagement of the alternative pathway in the seed axis. It is suggested that the lowering of respiratory activity and the shift in respiratory pathways may play a major role in the decline of germinability and vigor.

Alternative Respiratory Pathway

Oxygen uptake during the first hours of imbibition in intact soybean and mung bean seeds showed a marked sensitivity to potassium cyanide but was unaffected by addition of either salicylhydroxamic acid or propyl gallate. However O(2) uptake by finely ground seed particles was very sensitive to the addition of either compound. The results indicated that O(2) uptake in intact, imbibing seeds was associated with a cyanide-sensitive process, most probably mitochondrial mediated respiration, and not the result of the cyanide-insensitive lipoxygenase activity which was readily detectable in ground seed particles.The antioxidant propyl gallate was found to inhibit specifically alternative pathway electron transfer in isolated mung bean mitochondria. Half-maximal inhibition occurred with 2 to 5 micromolar propyl gallate. Kinetic analysis indicated that propyl gallate inhibition of the alternative pathway occurred at, or very near, the site of inhibition of the alternative pathway by salicylhydroxamic acid.A high level of lipoxygenase activity was found to be associated with washed mitochondria isolated from a variety of etiolated plant tissues. Most of this lipoxygenase activity could be eliminated from mung bean mitochondria if the mitochondria were purified on a discontinuous sucrose gradient. This indicated that the mitochondrial-associated activity was probably the result of nonspecific adsorption of lipoxygenase onto the mitochondrial membranes during isolation.

Purine Nucleotide Metabolism of Germinating Soybean Embryonic Axes

Isolated soybean (Glycine max L. cv. Kent) embyronic axes metabolized [(14)C]glycine to ATP within the 1 hour of imbibition. Radioactivity was not detected in GTP until the 3rd hour. Throughout most of the first 24 hours of germination about 10 to 26 times as much label from [(14)C]glycine appears in ATP as GTP. About five times as much [(14)C]hypoxanthine and [(14)C]inosine was converted into GTP as into ATP in embryonic axes. Two independent pools of IMP appear to be used in purine nucleotide synthesis of soybean axes.

Rapid Release of Protease Inhibitors from Soybeans

Specific antisera were prepared against the Bowman-Birk trypsin inhibitor and four other trypsin inhibitors of low molecular weight isolated from soybeans (Glycine max L. cv. Tracy). These antisera were used to detect the presence and amount of the inhibitors in: (a) seeds and protein extracts of soybean meal; (b) seedlings; and (c) the water surrounding the seeds and roots of seedlings. Lectin activities in seeds, seedlings, and water were also determined at the same time as the protease inhibitor activities. By competitive inhibition of immunoprecipitation, the combined five low molecular weight protease inhibitors were found to constitute the following percentages of proteins (w/w): 6.3% in defatted soybean meal; 8.1% of the protein extracted from the meal by a buffer of pH 8.6; 8.3, 14.7, 15.2, 16.1, 17.2, and 18.9% of the protein in a lyophilisate of water in which seeds were incubated for 4, 8, 12, 16, 20, and 24 hours, respectively; 8.2% in a lyophilisate of water in which roots of seedlings grew for 20 days; 1.5% in cotyledons; and less than 0.1% in epicotyls, hypocotyls, and roots of 12-day-old seedlings. Hemagglutination activities, expressed as the lowest amount of protein required to give a positive agglutination of 0.2 ml of 2% rabbit red blood cells, were as follows: purified soybean lectin, 0.08 mug; lyophilisate of water in which seeds were incubated for 4, 8, 12, 16, 20, and 24 hours, 10, 2.5, 5, 5, and 2.5 mug, respectively; lyophilisate of water in which roots grew for 20 days, 5 mug; 12-day-old cotyledons, roots, epicotyls, and hypocotyls, 12.5, 100, >1,000, and >500 mug, respectively. The results indicate that a large amount of protease inhibitors as well as lectins are released from seeds during the first 8 hours of imbibition. Neither lima bean trypsin inhibitor (mol wt, 10,000) nor Kunitz soybean trypsin inhibitor (mol wt, 21,500) showed competitive inhibition in tests with antisera against low molecular weight soybean protease inhibitors.

Mode of Action of Gibberellic Acid and Light on Lettuce Seed Germination

The seeds of lettuce (Lactuca sativa L. cv. Grand Rapids) germinate in darkness at 25 C when treated by gibberellic acid (GA(3)) for 1 hour following 2 hours of imbibition. The time of GA(3) application influences the rate and the final percentage of seeds that germinate. In contrast, red light illumination given at different times affects only the rate and not the final germination percentage. The early process(es) of germination initiated by GA(3) or light treatment can be arrested by subjecting the treated seeds to a nongerminative temperature of 35 C. The results suggest differences in the mode of action of light and GA(3) during germination. They indicate that different kinds of processes are involved in the biochemical control of germination.

Decline in ribonucleic acid and protein synthesis with loss of viability during the early hours of imbibition of rye (Secale cereale L.) embryos.

A decline in protein synthesis and slow germination accompanies loss of viability in embryos of rye (Secale cereale L.) grains. Associated with this, incorporation of precursors into all the major classes of RNA is decreased and the processing of precursor rRNA to 25S and 18S RNA is retarded. Embryos that just reach 0% viability still synthesize some low-molecular-weight non-nucleolar material, although they do not synthesize protein. It is suggested that early-synthesized RNA could play a major part in determining the extent of protein synthesis at early germination, and thereby regulate the rate at which germination can proceed.