In vivo Studies on Protein Synthesis in Developing Seeds of Canavalia gladiata D.C.

In successive stages of seed development of Canavalia gladiata,fruits were harvested and time-course changes of seed proteinaccumulation and SDS-gel electrophoretic patterns of the polypeptideswere examined. Albumin content increased gradually after thestage of 1.8 g fresh weight per seed, reached a peak, then decreasedat later stages. SDS-gel electrophoresis showed a clear changein albumin composition between the stages of 2.3 g and 3.6 gfresh weight per seed. Seed globulins were synthesized activelyafter the stage of 1.8 g fresh weight per seed, and the majorpolypeptides of seed globulins, including the main polypeptidesof canavalin, were first detected at this stage by gel electrophoresis.In contrast to the albumins, these polypeptides of globulinscontinued to accumulate until the maturation was completed.In vivo protein synthesis was also analyzed by SDS-gel electrophoresisand fluorography, after [³⁵S]methionine had been fed to cotyledonsections of maturing seeds at the stages of 3.4 g and 5.0 gfresh weight per seed. The results indicated that the synthesisof canavalin starts earlier than that of concanavalin A andalso ends earlier during seed development. (Received December 10, 1985; Accepted June 2, 1986)     

Moisture Loss as a Prerequisite for Seedling Growth in Soybean Seeds (Glycine max L. Merr.)

Rosenberg, L. A. and Rinne, R. W. 1986. Moisture loss as a prerequisitefor seedling growth in soybeanseeds (Glycine max L. Merr.).—J.exp. Bot. 37: 1663–1674. As soybean seeds [Glycine max (L.) Merr.] develop, they undergoa change in seed moisture. When excised prematurely from thepod and planted, seeds do not exhibit seedling growth until63 d after flowering (DAF) when the seed moisture has fallenbelow 60%. In contrast, seed germination (radicle protrusion)can occur when seeds as young as 35 DAF (68–79% moisture)are excised, but this germination docs not lead to comparableseedling growth frequencies unless seeds are first given a moistureloss treatment to artificially reduce their moisture below 60%.A moisture loss treatment applied at 35 DAF thus enables seedto undergo the transition from germination (cell expansion)to seedling growth (cell division and expansion) to the extentthat treated immature seed have a vigour index comparable toseeds matured on the plant (100%). The pattern of protein synthesisin vivo was examined in 35 DAF seed using [³⁵S]-methionine incorporation.When moisture loss treatment was applied for 24 h to 35 DAFseeds, seeds synthesized several new polypeptides when comparedwith untreated seeds at the same developmental stage. The sameseed samples showed 0% seedling growth in the absence of moistureloss treatment and 80% seedling growth when the treatment hadbeen applied. Moisture loss from soybean seeds appears to bea prerequisite for the synthesis of new proteins which may bepart of the metabolic process or processes that allow the soybeanseed to undergo the transition from seed germination to seedlinggrowth. Key words: Moisture loss, germination/growth, soybean     

A Comparison of Leek (Allium porrum) and Onion (Allium cepa) Seed Development

Leek and onion seed dry weight increased exponentially for thefirst 31 days after flowering (DAF) but thereafter the increasein dry weight was slower. Before maximum seed dry weight wasreached at 45 DAF in onion and 59 to 66 DAF in leek, seed moisturecontent, seed oxygen uptake and conductivity of the seed steepwater fell from initially high levels. Although some seeds germinated31 DAF in both species, full germination in both was not reacheduntil 66–80 DAF. Tolerance of the seed to artificial dryingimmediately after harvesting occurred 45 DAF in onion and 74DAF in leek. Free nuclear division continued in the endospermuntil 17–22 DAF in onion and until 31–35 DAF inleek but it was not until 45 DAF in onion and 66 DAF in leekthat the embryo and endosperm filled the cavity formed by thepericarp. After formation of cell walls in the endosperm thepattern of change in cell number in both species was similar.The shrunken appearance of the seed coat in leek, which occurredearly in seed development, was associated with the period offree nuclear division in the endosperm and, in addition, thepericarp was thinner than in onion. There was no evidence thatthe shrunken seed coat early in development was associated withself as opposed to open-pollination. Allium porrum, Allium cepa, seed development, endosperm, embryo, cell number, germination, respiration, seed leachates     

Development of Desiccation Tolerance and Longevity in Seeds from Detached Capsules of Foxglove (Digitalis purpurea L)

Developing capsules of foxglove (Digitalis purpurea L.) weredetached at 4-d intervals between 12 and 28 d after flowering(DAF) and attached to canes within a natural foxglove standsuch that they were experiencing field conditions identicalto those experienced by normally developing, on-plant capsules.Seeds were subsequently harvested at 4-d intervals until 40total d after flowering (tDAF). Capsule detachment resultedin the cessation of dry matter accumulation; the mean dry weightof seeds from 12, 16, 20, 24, and 28 DAF-detached capsules was21, 32, 51, 60, and 79% respectively, of the mean dry weightof seeds during the post-abscission phase of normal, on-plantdevelopment. Nonetheless, seeds from detached capsules acquiredthe ability to germinate at harvest and tolerance to dryingunder seed conservation conditions (15% relative humidity and15 °C). The capability to withstand storage also arose followingcapsule detachment. Seed longevity increased the longer theperiod of detachment but, in the earlier-detached capsules (12,16, and 20 DAF) longevity subsequently declined. Only seedsfrom later detached capsules (24 and 28 DAF) acquired longevitieswhich were comparable with seeds from on-plant capsules, however,no seeds from detached capsules were as long lived as seedsfrom on-plant capsules harvested at 40 DAF. Digitalis purpurea L.; foxglove; capsule detachment; seed development; desiccation tolerance; longevity     

Development of the Recalcitrant (Homoiohydrous) Seeds of Avicennia marina: Anatomical, Ultrastructural and Biochemical Events Associated with Development from Histodifferentiation to Maturation

Early histodifferentiation of the embryo of Avicennia marina(Forssk) Vierh was characterized by the formation of endospermhaustoria Once the growth phase was initiated, subsequent embryodevelopment was extra-ovular The mature seed, therefore, wasenclosed by a pericarp originating entirely from the ovary wallGrowth and reserve deposition was not initiated until 45–50d after fruit set (DAFS), when respiratory activity had peakedWater content remained constant from the earliest stages ofembryogenesis to seed abscission and respiratory activity, althoughdeclining somewhat after the completion of histodifferentiation,remained relatively high throughout seed development The ultrastructureof the meristematic root primordia was indicative of metabolicactivity, remaining essentially similar in all respects fromthe end of histodifferentiation until the mature seeds wereabscised During this phase cotyledon cells became highly vacuolatedand the soluble sugars, which constituted the major nutrientreserves of mature seeds, increased considerably Seeds of A,marina initiate germination, without the requirement for additionalwater, as soon as they are shed It is proposed that the accumulationof soluble sugars, rather than insoluble complex reserves, isa major factor in the developmental strategy of these highlyrecalcitrant seeds Anatomy, Avicennia marina, biochemistry, homoiohydrous, recalcitrant, seed development/maturation, ultrastructure     

Freezing injury to ovules, pollen and seeds in winter rape

Winter rapeseed (Brassica napus, cv. Samouraï) flowersearly in spring and, under field conditions, short freezingperiods can occur. Unacclimatized plants were freeze-stressed(–3°C for 4 h) at different developmental stages ofbuds, open flowers and seeds. The dissection of pistils from stressed plants showed that freezingresults in shrivelled ovules. We assessed freezing injury onthe basis of per cent of shrivelled ovules: ovule sensitivitybegins early (8 d before anthesis) but increases up to anthesis.Crosspollination of stressed pistils with non-stressed pollenshowed that recording of freeze-injured ovules is a good methodfor early estimation of the effect of stress on seed yields. On the other hand, stress does not reduce the viability of pollen,except when it was applied at the binucleate pollen stage. Useof frozen pollen x nonstressed pistils has little effect onseed yields. Freeze injury on seeds was assessed by seed filling:seeds are very susceptible just after fertilization until 20d after fertilization (DAF). Freezing seems to inhibit seedfilling. A germination test of stressed seeds during their developmentindicated that embryo viability is not affected if the stressoccurs after 35 DAF. As the embryos develop, resistance to stressincreases. Key words: Brassica napus, rapeseed, freeze injury, pollen and ovule, seed filling     

Expression of a-Amylase in Phaseolus vulgaris and Vigna mungo Plants

Levels of starch and soluble sugar in pods of Phaseolus vulgarisand Vigna mungo plants were analyzed during the course of maturationof fruits. The results suggest that the immature pods of P.vulgaris function to some extent as temporary reservoirs ofcarbohydrates for growth of seeds. A less clear pattern of accumulationof starch was observed in pods of maturing fruits of Vigna mungo.Measurements of a-amylase activites in pods of maturing fruitsand immunoblotting with an antiserum against     

Nucleotide Sequence of cDNA for Concanavalin A from Canavalia gladiata Seeds

We have determined the nucleotide sequence of Con A cDNA forconcanavalin A (Con A) from Canavalia gladiata using a plasmid(pCONAl) that was isolated previously [Eur. J. Bio-chem. (1988)170: 515-520]. This sequence contains a 870-bp open readingframe, a 63-bp 5'-un-translated region and a 99-bp 3'-untranslatedregion. DNA blot analysis suggested that Con A is encoded bya small gene family. In contrast to the case of canavalin, thenucleotide sequence and the genomic organization of Con A geneare highly conserved between C. gladiata and C. ensifor-mis. (Received August 4, 1988; Accepted November 21, 1988)     

Developmental Changes in Relation to Desiccation Tolerance of Archontophoenix alexandrae (Palmae) Seeds

Germination of Archontophoenix alexandrae seeds and embryos were studied under gradient water content treatments throughout the seed development phases of maturation in 2005 to investigate seed desiccation tolerance and storage characteristics. During the maturation process, seed water content decreased gradually from55 DAF (days after flowering) to 70 DAF, and seeds reached the maximum dry-weight at 90 DAF. Seed germinability appeared after 60 DAF. Seeds germinated with a temperature range from15℃- 40℃ under alternating photoperiod (14 h light, 10 h dark, 12μmol m^{- 2}s ^{- 1} ), while the best germination percentage was obtained between 30℃- 35℃. A maximum germination capacity reached at 70 DAF. However, seed germination was greatly inhibited by light. Desiccation tolerance of seeds and embryos increasedgradually from 55 DAF to 90 DAF and reached the maximum at 90 DAF with a semilethal water content of 0.18 g/g ( seed) and 0.3 g/g ( embryo) respectively. Rapid dehydration maintained higher seed germination percentage than thatof slow dehydration when drying to the same water content. Seeds with without water content treatments failed to germinate after 1 month storage under - 18℃, whereas appropriate desiccation treatment prolonged seed longevity under 4℃, 10℃ and 15℃ storage temperatures. It revealed obviously the recalcitrant characteristics of Archontophoenix alexandrae seeds torage behaviour which are tolerant toward neither deep desiccation nor low temperatures.     

The Partial Characterization of the Major Seed Storage Proteins in Arabidopsis thaliana L.

This study was aimed at the characterization of the major storage proteins in Arabidopsis thaliana. Two major protein fractions, i.e., the fraction Ⅰ and Ⅱ proteins, were isolated from the extract of mature seeds of this plant by molecular seive gel filtration chromatography. Various polyacrylarnide gel electrophoretic techniques were used to study the properties and polypeptide compositions of these two protein fractions. In was shown that during the SDS gel electrophoresis, fraction Ⅰ protein was separated into 6 major bands with the mol. was. of 34, 31, 29, 28 and 19-20 kD, respectively, whereas Fraction Ⅱ protein migrated as 3 low mol. wt. bands (10-12 kD) on the same gel. Non-denaturing native gel electrophoresis revealed that fraction Ⅰ was a neutral protein and Fraction Ⅱ was a positively charged basic protein with an isoelectric point (pI) higher than 8.8. Fraction I protein was further separated into at least 16 polypeptides in isoelectric focusing/SDS two-dimensional gel electrophoresis, i.e. each SDS band contained 3-4 polypeptides with the same mol. wt. but different pis. This suggested a more complex polypeptide composition of this protein. The properties of fraction Ⅰ and Ⅱ proteins were in good accordance with that of the 12s and 1.7s storage globulins in seeds of many other dicotyledonous plants, and therefore had been characterized as the two major seed storage proteins in this species. These two storage globulins were shown to be accumulated within a defined period during the late stage of seed development (12-14 DAF) and became predominant protein components in mature seeds. In the mean time, a few points in relation to the polypeptide composition and subunit molecular configuration of the 12s globulin were noted.     

Studies on Changing Protein Levels in Developing and Germinating Seeds of Lathyrus sativus L.

SDS-polyacrylamide gel electrophoresis of seed protein extracts showed synthesis of major legumin and vicilin polypeptides from 30 OAF onwards though very faint polypeptide bands could be seen at early stage of 10 OAF. Globulin synthesis enhanced and dominated in the second half of seed development. In germinating seeds, legumin polypeptides (mol wt 75–66, 45–41 kD) and vlcllin polypeptides (mol wt 54 kD) were degraded Increasingly from 2nd to 6th day of germination. Globulins and glutellns decreased and albumins Increased from 2nd to 6th day of seed germination.     

Influence of systemic insecticides on lipid biosynthesis in seeds of Indian mustard (Brassica juncea L.)

The systemic insecticides namely phorate (Thimet 10 G) oxydemeton methyl (metasystox 25 EC) and dimethoate (Rogor 30 EC) decreased oil content in the developing seeds of Indian mustard (Brassica juncea L.) but showed an increase in the mature seeds. The inhibitory effect in the developing seed was accompanied by an increase in soluble sugars and a corresponding decrease in malate dehydrogenase and G6P dehydrogenase activity. The relative proportion of triacylglycerols and glycolipids decreased significantly while that of phospholipids and free fatty acids increased in the developing seeds. In the mature seeds, the proportion of triacylglycerols did not change appreciably from that in control. The erucic acid synthesis which was less at 10 and 20 days after fertilization (DAF) increased at 30 DAF with oxydemeton methyl and dimethoate; phorate was ineffective. In mature seeds, the proportion of erucic acid increased at the cost of linoleic and linolenic acids. All the insecticides appreciably decreased the rate of [1-¹⁴C]acetate incorporation into lipids both in vivo and in vitro experiments. In the in vivo experiment, the synthesis of polar lipids was enhanced at 10 and 20 DAF, and the higher doses of oxydemeton methyl and dimethoate at 30 DAF. On the other hand, the ¹⁴C-incorporation into triacylglycerols showed an opposite trend to that of polar lipids. In the in vitro experiment, oxydemeton methyl and dimethoate enhanced the synthesis of polar lipids at 10 and 20 DAF while these inhibited it at 30 DAF. The synthesis of triacylglycerols was inhibited by the use of these insecticides.     

Comparison of in vitro translation products of Sarcocystis gigantea and Sarcocystis tenella.

Poly(A)+ RNA was purified from cystozoites of Sarcocystis gigantea and Sarcocystis tenella and used to in vitro translate polypeptides in a wheat germ and a rabbit reticulocyte translation system. The in vitro translated polypeptides were compared by sodium dodecyl sulphate-polyacrylamide gel electrophoresis. The S. tenella mRNA translated at least two polypeptides (mol. wt about 80,000 and 21,500) in both translation systems that were not translated by the S. gigantea mRNA. To study co-translational and initial post-translational processing in Sarcocystis, the poly(A)+ RNA preparations were in vitro translated in the rabbit reticulocyte translation system in the presence or absence of canine microsomal membranes. Based on electrophoresis, there appeared to be modification of at least some Sarcocystis polypeptides in the mol. wt range 17,000-30,000. In addition, the translation products were immunoprecipitated with a homologous and a heterologous antiserum. The immunoprecipitated polypeptides were compared by electrophoresis and the S. tenella translation products contained at least one unique antigenic polypeptide with a mol. wt of about 34,700 that was not processed by the microsomal membranes. These results suggest that there is at least one polypeptide that is a candidate for use as an antigen for the differentiation of S. gigantea and S. tenella infections in sheep.     

Biochemical Characteristics Associated with the Development of the Desiccation-sensitive Seeds of Machilus thunbergii Sieb. & Zucc.

Biochemical properties, i.e. endogenous abscisic acid, proline,sugars, respiration, adenosine phosphates and adenylate energycharge, and growth and moisture content were measured duringthe development of seeds of Machilus thunbergii. As dry matteraccumulated in the embryo during development, moisture content,ABA, proline, respiration and sugars all declined. At maturity,the dry mass of the seeds failed to attain a plateau beforethe period of natural seed shedding; the axis and cotyledonsreached moisture contents of 58 and 45%, respectively. Dryingof immature seeds at 73% relative humidity and 25 °C for30 d resulted in a complete loss of viability at all developmentalstages tested with the exception of mature seeds that were ableto tolerate a 5% decrease in moisture content before germinationdeclined. ABA was detected in all embryos tested, with a maximum value16.·16 µg g^-1 d. wt about midway through development.Although the presence of ABA induced no tolerance to desiccationof mature seeds, it did coincide with decreased content of waterin the developing seeds and decreased respiration. Desiccationdamage of M. thunbergii seeds occurred when moisture contentwas still high (45%) and this damage was not related to theabsence of oligosaccharides in the mature seeds. We concludethat developing embryos and mature seeds of M. thunbergii haveproperties common to many recalcitrant seeds, with seeds beingsensitive to desiccation at all stages, having a prominent ABApeak, little proline, lacking oligosaccharides, and specifically,little dormancy and a moderate rate of respiration of matureseeds (0·9 µmol O₂ min^-1 g^-1 f. wt). Adenosinetriphosphate content and energy charge decreased from stagefour to stage eight of seed development, then increased againto 103 nmol g^-1 d. wt and 0·73, respectively, in matureseeds. The moderate energy charge observed in mature seeds indicatesthat continuous metabolism is also a characteristic of recalcitrantseeds.Copyright 1995, 1999 Academic Press Machilus thunbergii, seed development, recalcitrant seed, abscisic acid, energy charge     

Photoperiodically Induced Changes in Seed Growth Rate of Soybean as Related to Endogenous Concentrations of ABA and Sucrose in Seed Tissues

Short-day photoperiods can increase the partitioning of assimilatesto filling seeds of soybean (Glycine max L. Merr.), resultingin higher seed growth rates. The plant growth substance ABAhas been implicated in the regulation of assimilate transferwithin filling soybean seeds. Thus, we hypothesized that anincreased concentration of endogenous ABA in seeds may enhancesucrose accumulation and seed growth rate of soybeans exposedto short-day photoperiods. Plants of cv. Hood 75 were grownin a greenhouse under an 8-h short-day photoperiod (SD) until11 d after anthesis (DAA) of the first flower, when half ofthe plants were transferred to a night-interruption (NI) treatment(3 h of low-intensity light inserted into the middle of thedark period). Plants remaining in SD throughout seed developmenthad seed growth rates 43% higher than that of plants shiftedto NI (7·6 mg seed^–1 d^–1 vs. 5·3 mgseed^–1 d^–1). On a tissue-water basis, the concentrationof ABA in SD seeds increased rapidly from 7.6 µmol l^–1at 11 DAA to 65·2 µmol l^–1 at 18 DAA, butthen declined to 6·6 µmol l^–1 by 39 DAA.In contrast, the concentration of ABA increased more slowlyin NI seeds, reaching only 47·4 µmol l^–1by 18 DAA, peaking at 57·0 µmol l^–1 on 25DAA, and declining to 10·2 µmol l^–1 by 39DAA. The concentration of sucrose in SD embryos peaked at 73·5mmol l^–1 on 25 DAA and remained relatively constant forthe remainder of the seed-filling period. In NI, the concentrationof sucrose reached only 38·3 mmol 1^–1 by 25 DAA,and peaked at 61·5 µmol l^–1 on 32 DAA. Thusin both SD and NI, sucrose accumulated in embryos only afterthe peak in ABA concentration, suggesting that ABA may havestimulated sucrose movement to the seeds. The earlier accumulationof ABA and sucrose in SD suggests that ABA may have increasedassimilate availability during the critical cell-division period,thus regulating cotyledon cell number and subsequent seed growthrate for the remainder of the seed-filling period. Glycine max L. Merr. cv. Hood 75, soybean, assimilate partitioning, abscisic acid, photoperiod, source-sink     

Effects of Nutrient Supply on Seed Growth of Field Bean (Vicia faba L.). 1. Effects of Different Sulphur and Methionine Regimes on Cotyledon Growth, Protein and Uncombined Amino Acids In vivo

The effects of exogenous methionine on growth of developingVicia faba seeds in vivo was studied. Methionine (51 or 102µmol in total) injected into the base of the pod overa period of 15 d (26–40 d after pollination) decreasedgrowth and protein accumulation in proximal seeds but increasedin distal seeds. Distal seeds of methionine injected pods accumulatedmore d. wt and protein than distal seeds of water-injected pods.However, on a pod basis methionine reduced overall seed growthand protein accumulation. Exogenous methionine caused a relative increase in legumin buta decrease in vicilin. Injected methionine also affected thecomposition of uncombined amino acids, especially those derivedfrom aspartic acid. In contrast, the amino acid compositionof the protein fraction did not change appreciably. The data suggest that seed growth is not limited by methioninebiosynthesis but seed protein composition is regulated by methionine. Vicia faba L., field bean, cotyledon, growth, in vitro culture, uncombined amino acids, protein composition, legumin, vicilin, methionine, sulphur     

Seed Development in Podocarpus henkelii: an Ultrastructural and Biochemical Study

Biochemical and fine-structural studies were undertaken on theembryo and female gametophyte of seeds of Podocarpus henkeliiduring the 24-week period of post-fertilization growth and development. Although the d. wt of the embryo and female gametophyte showeda steady increase during development, the levels of sugars,amino acids, proteins and lipids differed between seed partsand showed changes during the four sampling intervals. Onlystarch was seen to increase steadily throughout development.Lipid levels were high in the mature embryo and free amino acidsshowed a steady increase until the seeds were shed. At thisstage seeds were fully hydrated, possessed abundant reservesand all organelles appeared fully functional. This was interpretedas part of the development strategy of neotonous (recalcitrant)seeds, namely, the maintenance of full metabolic competencefor continued growth in the absence of development arrest whichfollows drying in orthodox seeds. Podocarpus henkelii, yellow wood, embryo and endosperm growth, neotonous seed, recalcitrance     

Changes in levels of Proteases and Their Inhibitors during Development of the Seeds of Job's Tears (Coix lacryma-jobi L. var. Ma-yuen Stapf) and Their Localization and Interactions

Changes in levels and localization of proteases and a trypsininhibitor (JBTI) in the developing seeds of Job's tears (Coixlacryma-jobi L. var. Ma-yuen Stapf) were followed, and theirinteractions were examined. The JBTI was induced from the middlestage of development (12 DAF, i.e., 12 days after flowering)and increased until the late stage of development (24 DAF),and was localized in the germ. Two groups of proteases (A, thosewith molecular weights of 55–70 kDa; and B, those withmolecular weights greater than 94 kDa) were detected by activestaining of substrate-containing SDS-polyacrylamide gels. Thegroup of larger proteases seems to consist of three members(B₁, B₂₎ B₃, with increasing molecular weights). Band A wasobserved only in the early stage of development (until DAF 9),band B₂ persisted during all stages, while bands B, and B₃ werepresent only during early and late stages, respectively. Thegroup A proteases and one of the group B proteases (probablyB₁) were inhibited by diisopropyl-fluorophosphate (DFP), bytrypsin inhibitors from soybean and rice, and by JBTI. The proteasesthat were present in the seeds of Job's tears at a late stageseemed to be localized in the germ. (Received September 9, 1988; Accepted April 19, 1989)     

Canatoxin-, concanavalin A- and canavalin-cross-reactive materials during maturation of Canavalia brasiliensis (Mart.) seeds

The distribution of three cross-reactive materials (CRMs), a toxic protein analogous to canatoxin, CNTX-CRM, a lectin analogous to concanavalin A, Con A-CRM, and a major storage protein, canavalin-CRM, was investigated during successive stages of maturation of Canavalia brasiliensis Mart. seeds. The data obtained by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and immunological analyses indicated that these proteins share extensive homology with the analogous proteins found in Canavalia ensiformis seeds. The changes in CNTX-CRM and Con A-CRM levels throughout the maturation process were assayed by rocket immunoelectrophoresis. Synthesis of Con A-CRM was detectable at 30 days post-anthesis (DPA) while its hemagglutinating activity appeared only at 35 DPA. The CNTX-CRM was detected as a biologically active protein from 30 DPA onwards. The behavior of CNTX-CRM during maturation of C. brasiliensis seeds was quite distinct from that of Con A-CRM, pointing to different biological roles of these proteins in the seed.