A structural study of germination in celery (Apium graveolens L.) seed with emphasis on endosperm breakdown

Germination of celery seed occurred after 6 d of imbibition in light. During this time the embryo enlarged at the expense of the adjacent endosperm cells and at the time of germination was 2–3 times as long as in the dry seed. Breakdown of the endosperm cells near the root cap preceeded radicle emergence. None of these changes occurred in darkness.Endosperm digestion began adjacent to the embryo and spread radially. In degrading cells, the aleurone grains often became larger and fewer in number. The cell walls were modified and appeared to undergo partial degradation. Ultimately the cells seemed to lose their contents. In cells adjacent to the root cap, similar changes occurred except there was a transient appearance of starch grains. Radial progression of endosperm breakdown also occurred in isolated endosperm treated with gibberellin A₄₊₇.The results indicate that (1) the stimulus for breakdown of celery endosperm emanates from the embryo in response to light; (2) the stimulus may be a gibberellin because changes in endosperm cells and the sequence of endosperm digestion during germination resemble the responses of isolated endosperm to gibberellin; and (3) the radial progression of endosperm breakdown during germination may be the result of a sequential response of cells to a uniformly applied stimulus rather than the result of gradual embryo expansion.     

Action of light and gibberellic acid on the growth of excised embryos from Phacelia tanacetifolia seeds

Summary Germination of Phacelia tanacetifolia seeds is inhibited by light. Embryos freed of endosperm grow irrespective of light. However, if they are held in 0.3 M mannitol plus 2% sucrose, light sensitivity is reinstated: growth (i. e., germination) occurs in darkness but not in light. Gibberellic acid (GA₃) releases the inhibition due to light. These results suggest that (a) the photoreceptor site of the seed is in the embryo; (b) GA₃ acts directly on the embryo; and (c) darkimbibed embryos appear to have a higher water-uptake potential.This work was supported in part by the Jane Coffin Childs Memorial Fund for Medical Research, and by the U.S. Atomic Energy Commission under Contract No. AT(11-1)-1338.     

Light or high (30°C) temperature effects on l-[U-C]leucine incorporation and protein patterns in embryos and endosperm during the early phase of germination of the negatively photoblastic and thermosensitive seeds of Phacelia tanacetifolia

The activities of l-[U-¹⁴C]leucine uptake and incorporation into proteins of embryos and endosperm of seeds of Phacelia tanacetifolia Benth. cv Bleu-Clair were analysed during the first 24 h of incubation under conditions optimal for germination (16°C in darkness) and in two inhibitory conditions: 16°C in the light and 30°C in darkness. Blocking germination induced by light or 30°C was accompanied by the inhibition of l-[U-¹⁴C]leucine uptake and incorporation in embryos. In the endosperm, the activation of l-[U-¹⁴C]leucine uptake was of the same magnitude for the non-inhibited and the light-inhibited seeds and much higher for the 30°C-inhibited seeds; the activation of l-[U-¹⁴C]leucine incorporation was quantitatively similar in all three conditions, with the patterns of newly synthesised proteins qualitatively different in the endosperm from light- or 30°C-inhibited seeds. The results showed that germination of P. tanacetifolia seeds is controlled by light or super-optimal temperature through the inhibition of the activation of transport and protein synthetic activities in embryo without effect on the endosperm. We suggest, on the basis of the translational activity, the possibility that in the inhibitory conditions the blockade of the embryo to operate as a sink affects the transition of the endosperm to operation as a source.     

Photoinhibition of radish (Raphanus sativus L) seed germination: control of growth potential by cell-wall yielding in the embryo

The biophysical mechanism underlying photoinhibition of radish (Raphanus sativus L.) seed germination was investigated using three cultivars differing in sensitivity to continuous irradiation with far-red light (high-irradiance reaction of phytochrome). Sensitivity of germination to the inhibitory action of light was assessed by probing germination under osmotic stress (incubation in media of low water potentials adjusted with polyethylene glycol 6000) and expressed in terms of ‘germination potential’ (positive value of the water potential at which germination is inhibited by 50%). Far-red light decreases the germination potential to various degrees in the different cultivars, reflecting the light-sensitivity of germination in water. Removal of the seed coat increases the germination potential by a constant amount in darkness and light. It is concluded that germination depends on the expansive force of the embryo which can be drastically diminished by far-red light. Seed-coat constraint and expansive force of the embryo interact additively on the level of the germination potential. Photoinhibition of germination was accompanied by an inhibition of water uptake into the seed. Analysis of seed water relations showed that osmotic pressure and turgor assumed higher levels in photoinhibited seeds, compared to seeds germinating in darkness, while the water potential was close to zero under both conditions. Far-red light produced a shift (to less negative values) in the curve relating water-uptake rate to external water potential, i.e. a reduction in the driving force for water uptake. It is concluded that photoinhibition of germination results from the maintenance of a high threshold of cell-wall extensibility in the embryo.     

Dissecting seed dormancy and germination in Aquilegia barbaricina,through thermal kinetics of embryo growth

• Threshold‐based thermal time models provide insight into the physiological switch from the dormant to the non‐dormant germinating seed.
• This approach was used to quantify the different growth responses of the embryo of seeds purported to have morphophysiological dormancy (MPD) through the complex phases of dormancy release and germination. Aquilegia barbaricina seeds were incubated at constant temperatures (10–25 °C) and 25/10 °C, without pre‐treatment, after warm+cold stratification (W+C) and GA₃ treatment. Embryo growth was assessed and the time of testa and endosperm rupture scored. Base temperatures (T_b) and thermal times for 50% (θ₅₀) of embryo growth and seed germination were calculated.
• W+C enabled slow embryo growth. W+C and GA₃ promoted rapid embryo growth and subsequent radicle emergence. The embryo internal growth base temperature (T_be) was ca. 5 °C for W+C and GA₃‐treated seeds. GA₃ treatment also resulted in similar T_b estimates for radicle emergence. The thermal times for embryo growth (θ_e50) and germination (θ_g50) were four‐ to six‐fold longer in the presence of GA₃ compared to W+C.
• A. barbaricina is characterised by a multi‐step seed germination. The slow embryo growth during W+C reflects continuation of the maternal programme of development, whilst the thermal kinetics of both embryo and radicle growth after the removal of physiological dormancy are distinctly different. The effects of W+C on the multiphasic germination response in MPD seeds are only partially mimicked by 250 mg·l^?1 GA₃. The thermal time approach could be a valid tool to model thermal kinetics of embryo growth and radicle protrusion.

     

ABA inhibits embryo cell expansion and early cell division events during coffee (Coffea arabica 'Rubi') seed germination

Background and Aims

Coffee seed germination represents an interplay between the embryo and the surrounding endosperm. A sequence of events in both parts of the seed determines whether germination will be successful or not. Following previous studies, the aim here was to further characterize the morphology of endosperm degradation and embryo growth with respect to morphology and cell cycle, and the influence of abscisic acid on these processes.

Methods

Growth of cells in a fixed region of the axis was quantified from light micrographs. Cell cycle events were measured by flow cytometry and by immunocytochemistry, using antibodies against β-tubulin. Aspects of the endosperm were visualized by light and scanning electron microscopy.

Key Results

The embryonic axis cells grew initially by isodiametric expansion. This event coincided with reorientation and increase in abundance of microtubules and with accumulation of β-tubulin. Radicle protrusion was characterized by a shift from isodiametric expansion to elongation of radicle cells and further accumulation of β-tubulin. Early cell division events started prior to radicle protrusion. Abscisic acid decreased the abundance of microtubules and inhibited the growth of the embryo cells, the reorganization of the microtubules, DNA replication in the embryonic axis, the formation of a protuberance and the completion of germination. The endosperm cap cells had smaller and thinner cell walls than the rest of the endosperm. Cells in the endosperm cap displayed compression followed by loss of cell integrity and the appearance of a protuberance prior to radicle protrusion.

Conclusions

Coffee seed germination is the result of isodiametric growth of the embryo followed by elongation, at the expense of integrity of endosperm cap cells. The cell cycle, including cell division, is initiated prior to radicle protrusion. ABA inhibits expansion of the embryo, and hence subsequent events, including germination.Key words: Abscisic acid, β-tubulin, Coffea arabica, coffee seed, cell morphology, germination, microtubules     

Light-controlled Cycocel Reversal of Coumarin Inhibition of Lettuce Seed Germination and Root Growth

Lettuce seed germination or lettuce root elongation after germination in water was inhibited by coumarin and these inhibitions were reversed by Cycocel. 2.53 × 10³ M Cycocel reversed the inhibition of seed germination by 6.8 × 10⁴ M coumarin. and 6.32 × lO^?4 M Cycocel reversed the inhibition of root elongation by 3.4 × 10³ M coumarin. No other analogs of Cycocel reversed these coumarin induced inhibitions of growth. Cycocel reversal of coumarin inhibition of lettuce seed germination occurred in red light but not in far-red light or darkness. The red-far-red system was photoreversible. Cycocel and kinetin appear to act similarly in reversing coumarin inhibition of germination. Gibberellin A₃ and IAA were unable to reverse these coumarin induced inhibitions. A common mechanism is suggested for Cycocel reversal of coumarin and lAA inhibition of growth.     

Effects of gibberellins, darkness and osmotica on endosperm rupture and class I β-1,3-glucanase induction in tobacco seed germination

The “Havana 425” cultivar of Nicotiana tabacum L. is photodormant. Gibberellins (e.g. 10^?5 M GA₄ or GA₇) can substitute for light in releasing dormancy. Measurements of β-1,3-glucanase activity, mRNA accumulation and the activity of the class I β-1,3-glucanase B promoter indicated that class I β-1,3-glucanases are induced by GA₄ in the dark in association with germination. As in the light, this induction occurred prior to endosperm rupture and was localized exclusively in the micropylar region of the endosperm where the radicle will penetrate. Abscisic acid (ABA, 10^?5 M) did not appreciably affect GA-induced release of photodormancy or seed-coat rupture, but it delayed endosperm rupture and inhibited the rate of class I β-1,3-glucanase accumulation. Seeds imbibed in the light in the presence of osmotica, e.g. 0.04 M polyethylene glycol 6000, showed delayed seed-coat and endosperm rupture, delayed onset of β-1,3-glucanase induction, and decreased rates of β-1,3-glucanase accumulation. These delays were shortened by GA₄ treatment. Our results suggest that GAs and ABA act at two distinct sites during germination and that expansive growth of the embryo acts in two ways by triggering β-1,3-glucanase induction and by providing force for endosperm penetration. This provides further support for our working hypothesis that class I β-1,3-glucanases promote endosperm weakening and facilitate radicle penetration.     

Embryo growth, testa permeability, and endosperm weakening are major targets for the environmentally regulated inhibition of Lepidium sativum seed germination by myrigalone A

Myrigalone A (MyA) is a rare flavonoid in fruit leachates of Myrica gale, a deciduous shrub adapted to flood-prone habitats. As a putative allelochemical it inhibits seed germination and seedling growth. Using Lepidium sativum as a model target species, experiments were conducted to investigate how environmental cues modulate MyA's interference with key processes of seed germination. Time course analyses of L. sativum testa and endosperm rupture under different light conditions and water potentials were combined with quantifying testa permeability, endosperm weakening, tissue-specific gibberellin (GA) and abscisic acid (ABA) contents, as well as embryo growth and apoplastic superoxide production important for cell expansion growth. Lepidium sativum testa permeability and early water uptake by imbibition is enhanced by MyA. During late germination, MyA inhibits endosperm weakening and embryo growth, both processes required for endosperm rupture. Inhibition of embryo cell expansion by MyA depends on environmental cues, which is evident from the light-modulated severity of the MyA-mediated inhibition of apoplastic superoxide accumulation. Several important key weakening and growth processes during early and late germination are targets for MyA. These effects are modulated by light conditions and ambient water potential. It is speculated that MyA is a soil seed bank-destroying allelochemical that secures the persistence of M. gale in its flood-prone environment.     

Effects of ethylene and some other environmental factors on different stages of germination in red root pigweed (Amaranthus retroflexus L.) seeds*

Abstract. Ethylene was found to promote two distinct processes during germination of redroot pigweed (Amarantus retroflexus L.) seeds: embryo expansion that splits the seed coat (incomplete germination), and radicle penetration through the more elastic endosperm (complete germination). The two events can be separated in time by subjecting seeds to low water potential or low CO₂ levels, which arrest germination of some seeds at the incomplete stage. Ethylene applications to incompletely germinated seeds promote complete germination, with a response threshold near 0.02 cm³ m^?3 and saturation near 0.5 cm³ m^?3. Higher ethylene concentrations (0.5 to 50 cm³ m^?3) given during the first day of seed imbibition also increase the percentage of seeds which initiate embryo expansion and split the seed coat. Light and elevated CO₂ also promote radicle penetration of the endosperm in seeds incubated under water stress. The results support the view that the germination pause at the incomplete stage is an adaptation to environmental stresses that can be overcome with exogenous ethylene or certain other stimuli.     

Gibberellin-induced separation of cells in isolated endosperm of celery seed

Summary The mature seed of celery (Apium graveolens, L.) contains a small axile linear embryo surrounded by endosperm which occpies the bulk of the seed. The endosperm is living and consists of mostly large angular thick-walled cells containing aleurone grains (often with globoids) and lipid droplets. — Using de-embryonated seeds, it has been shown that the endosperm was induced to break down by gibberellin. The aleurone grains became swollen and lost their contents and the bulk of each cell wall was hydrolyzed. However, a thin resistant layer of wall remained around each protoplast. The wall hydrolysis caused the endosperm to break down into individual cells which could be plasmolyzed and therefore appeared to be still living. All cells of the endosperm responded to gibberellin in a similar way although the cells near the radicle appeared to degrade more rapidly than those elsewhere. There was no change in the absence of the hormone. The response was apparently specific to gibberellin and did not occur in the presence of ethylene, kinetin, abscisic acid and indole acetic acid. The results were the same in light and in darkness. — It has been thought that endosperm breakdown during germination of seed such as celery involved release of hydrolases from the expanding embryo. The results of this study indicate that endosperm breakdown might be caused by hydrolases arising in the endosperm itself in response to gibberellin released from the embryo.Abbreviations GA gibberellic acid - PAS periodic acid-Schiff reagent Most of this work was performed in the Department of Vegetable Crops at the Volcani Center, Bet Dagan, while J.V. Jacobsen was a Research Fellow there.     

Seed germination ecology of the threatened endemic Iberian <Emphasis Type="Italic">Delphinium fissum</Emphasis> subsp. <Emphasis Type="Italic">sordidum</Emphasis> (Ranunculaceae)

Seeds of Delphinium fissum subsp. sordidum are physiologically dormant at maturity, with underdeveloped embryos; thus they have morphophysiological dormancy (MPD). The aims of this study were to determine the requirements for embryo growth, dormancy break and germination, to characterise the type of seed dormancy and to evaluate the effects of light, seed age, pollination mechanism, and inter-annual and inter-population variability on germinative ability. After 3 months of incubation at 5°C (cold stratification) in darkness conditions, the mean embryo length increased from 5.6 to 2.07 mm, with 76% of seeds germinating. Conversely, embryos of seeds incubated during 3 months at 20/7 or 28/14°C hardly grew and no germination was recorded. Since cold stratification was the only requirement for the loss of MPD, and both dry storage in laboratory conditions and warm stratification prior to cold stratification shortened the cold stratification period required for germination, it could be concluded that D. fissum subsp. sordidum seeds have intermediate complex MPD. Cold stratification and incubation in darkness conditions promoted higher germination percentages than those in light. In addition, germinative ability increased with seed age up to 8 months (reaching 96% at 5°C in darkness), showed a pronounced inter-annual and inter-population variability, as well as a significant decrease in seeds coming from pollination by geitonogamy. High temperatures (25/10 or 28/14°C) induced seeds to secondary dormancy, so seedling emergence in the greenhouse was restricted to February–March. The requirements for dormancy break and germination reflect an adaptation to trigger germination in late winter. This study is the first one to document a gradual increase in germination percentage with seed age for plant species with intermediate complex MPD.     

Changes in low-molecular-weight thiol-disulphide redox couples are part of bread wheat seed germination and early seedling growth

The tripeptide antioxidant glutathione (γ-l-glutamyl-l-cysteinyl-glycine; GSH) essentially contributes to thiol-disulphide conversions, which are involved in the control of seed development, germination, and seedling establishment. However, the relative contribution of GSH metabolism in different seed structures is not fully understood. We studied the GSH/glutathione disulphide (GSSG) redox couple and associated low-molecular-weight (LMW) thiols and disulphides related to GSH metabolism in bread wheat (Triticum aestivum L.) seeds, focussing on redox changes in the embryo and endosperm during germination. In dry seeds, GSH was the predominant LMW thiol and, 15?h after the onset of imbibition, embryos of non-germinated seeds contained 12 times more LMW thiols than the endosperm. In germinated seeds, the embryo contained 17 and 11 times more LMW thiols than the endosperm after 15 and 48?h, respectively. This resulted in the embryo having significantly more reducing half-cell reduction potentials of GSH/GSSG and cysteine (Cys)/cystine (CySS) redox couples (E_GSSG/2GSH and E_CySS/2Cys, respectively). Upon seed germination and early seedling growth, Cys and CySS concentrations significantly increased in both embryo and endosperm, progressively contributing to the cellular LMW thiol-disulphide redox environment (E_{thiol-disulphide}). The changes in E_CySS/2Cys could be related to the mobilisation of storage proteins in the endosperm during early seedling growth. We suggest that E_GSSG/2GSH and E_CySS/2Cys can be used as markers of the physiological and developmental stage of embryo and endosperm. We also present a model of interaction between LMW thiols and disulphides with hydrogen peroxide (H₂O₂) in redox regulation of bread wheat germination and early seedling growth.     

Physical and physiological dormancy in black henbane (<Emphasis Type="Italic">Hyoscyamus niger</Emphasis> L.) seeds

Aim of this study was to investigate the nature of dormancy in black henbane (Hyoscyamus niger) seeds which have low germination rate under normal laboratory conditions. To do this, before placing the seeds in Petri dishes, they were soaked in 5,10 and 15 mg/L GA; 1,2 and 3% H₂SO₄, 15 mg/L GA + 1% H₂SO₄, 0.01 M KNO₃ solutions, tap water, 40, 50 and 60°C hot water for 30 min. The study was performed under both continuous illumination and darkness in growth chambers to evaluate the effect of light on germination rate. The results showed that H₂SO₄ and GA treatments were the most important factors affecting seed germination and their germination enhancing effects were more evident in darkness. The results also suggested that black henbane seeds exhibit double dormancy involving a hard seed coat and a partially dormant embryo and have a partial dark requirement to germinate.     

The control of seed germination in Trollius ledebouri: The breaking of dormancy

Seeds of Trollius ledebouri exhibit low germination when maintained on moistened filter paper. Dormancy can be overcome by the application of gibberellins A₄+A₇ or by testa removal. Germination is characterised by a change in the anatomy of the seed and by specific alterations in the protein complement of the endosperm tissue. These anatomical and biochemical changes are also exhibited by isolated endosperm tissue maintained in the absence of the embryo. The observations described are discussed in relation to the interaction between the endosperm and the embryo in the control of seed germination in T. ledebouri.Abbreviation GA gibberellin     

Physiological Studies with Heavy Water. II. Germination and Growth Inhibition of Barley by D2O

Seed germination and barley seedling growth in various D₂O concentrations have been studied. It was observed that the emergence of root and shoot was delayed, there being greater delay in shoot than in root emergence. A complete block was observed in germination in pure D₂O and the germination rate was slowed down significantly in lower concentrations. An initial germination delay by different D₂O concentrations seemed to cause a subsequent retardation in the growth measured as shoot and root length. A comparison of root and shoot length with their respective dry weights suggested that the growth by cell division/elongation might have been affected more than the transport of food materials from the endosperm to the embryo. Analysis of the total sugars of the endosperm and the embryo at 8 hour intervals showed that while the hydrolysis of starch to sugars was progressively decreased by increasing D₂O concentrations, the transport rate from endosperm to embryo showed a sharp inhibition in 50% D₂O and above. This indicated that the inhibition in the transport of materials, besides less hydrolysis of reserve food materials, may also be a causal factor of germination and growth inhibition in D₂O.     

MG2+, K+-DEPENDENT PHOSPHATASE ACTIVITY DURING SEED DEVELOPMENT IN NICOTIANA TABACUM

Cation-dependent phosphatase activity was followed in developing Nicotiana tabacum (tobacco) seeds from pre-pollination to seed maturity. Enzyme activity increased rapidly up to 6 days post-pollination, then dropped off until 13 days after pollination. At 15 days post-pollination, enzyme activity had increased again, and at 17 days post-pollination, it reached its highest level. At 19 days post-pollination (seed maturity) enzyme activity was still 28% higher than that of the pre-pollination stage. Since the two peaks in enzyme activity correlate very highly with endosperm growth (first peak) and embryo growth (second peak), it is suggested that the Mg²⁺, K⁺-dependent phosphatase activity detected in this study is associated with the active transport of nutrients to the endosperm and embryo during seed development. The relatively high amount of enzyme activity remaining in the mature seed may represent a store of transport phosphatase that will be available for immediate use at the time of germination and early seedling growth.     

The Mechanism of Low Temperature Dormancy in Mature Seeds of Syringa Species

The mechanism of seed dormancy at low temperatures (15-9°C) was investigated in the seeds of Syringa josikaea, S. reflexa and S. vulgaris. Low temperature dormancy in Syringa species was mainly imposed by endosperm embedding the radicle. Different degrees of embryo dormancy may occur in S. reflexa seeds. In most cases the low temperature dormancy was broken completely by removing the endosperm around the radicle. The endosperm did not seem to contain significant quantities of germination inhibitors, and the results indicate that it prevents germination mainly due to its mechanical resistance. The mechanical resistance of endosperm did not change during chilling or during induction of dormancy by high temperature incubation. The strength of the endosperm decreased rapidly in non-dormant seeds before visible germination. Similar changes were not observed in dormant seeds. Generally, the strength of the endosperm was lower in the non- (or less) dormant species S. josikaea and S. vulgaris than in the more dormant S. reflexa seeds. The growth potential of the embryos, measured as their ability to germinate in osmotic solutions (mannitol or polyethylen glycol 4000), was increased by chilling and by GA₃-treatment. The growth potential of untreated S. josikaea and S. vulgaris embryos was generally higher than that of S. reflexa embryos. Acid ethyl acetate fractions of methanol extracts from embryos of all three species contained substances with GA³-like activity in the lettuce hypocotyl test. The activity was found at Rf 0.9–1.0 on paper chromatograms run in distilled water. No significant changes in the activity were detected during chilling or prior to visible germination.     

Two Sites of Hormonal Action during Germination of Chenopodium album Seeds

During the overall germination process in the light-requiring seeds of Chenopodium album L. two sites of hormonal action can be distinguished. The start of visible growth is induced in darkness by the gibberellins 4 and 7 (GA₄₊₇) or by ethylene. Added cytokinins are inactive although 8-¹⁴C-kinetin shows rapid penetration into the seed and rapid turnover. The induction of growth by GA₄₊₇ and the uptake of 8-methylene-¹⁴C-GA₃ are enhanced at lower pH values. Prolonged incubation in darkness results in a decreased responsiveness of the seeds towards GA₄₊₇ and ethylene. A second site of hormonal action is located during the progress of growth inside the covering structures. ABA inhibits radicle growth, while GA₄₊₇, GA₃, kinetin, zeatin and ethylene reduce the inhibition.     

Hoary cress (Cardaria draba (L.) Desv.) seed germination ecology,longevity and seedling emergence

The effects of gibberellic acid (GA₃), potassium nitrate (KNO₃), prechilling, temperature, salt stress and osmotic potential on seed germination and sowing depth on seedling emergence and burial depth on seed viability of hoary cress (Cardaria draba (L.) Desv.), were studied in a series of laboratory, glasshouse and outdoor experiments. The optimal temperature for hoary cress seed germination was 20°C, both in light/dark and darkness regimes. Seed germination of hoary cress at 400 ppm concentration of GA₃ in a light/dark regime was maximal. Potassium nitrate concentrations increased the percentage of germination in comparison with the control treatment. Increasing the duration of dry prechilling to 30 and 45 days promoted the seed germination of hoary cress. Germination of hoary cress markedly decreased as salt and drought stress increased. Seed germination of hoary cress occurred at a range of pH from 3 to 11. Seedling emergence significantly decreased as planting depth increased. Total seed viability decreased with increasing burial depth. The maximum increase in mortality occurred in seeds that were buried at 5‐cm depth.