Ultrastructural Studies of Somatic Embryos of Eucalyptus nitens and Comparisons with Zygotic Embryos Found in Mature Seeds

Although somatic embryogenesis has been observed in tissuesfrom a limited number of Eucalyptus species cultured in vitro,no comparisons have been made of the morphology and structureof eucalypt somatic embryos and zygotic embryos found in matureseeds. We used scanning and transmission electron microscopy,in conjunction with histological analysis, to compare maturezygotic embryos with somatic embryos of the commercially-importanttemperate eucalypt Eucalyptus nitens. Apart from differencesin the nature of the outer coating enclosing both embryo types,somatic embryos of E. nitens were observed to have strong similaritieswith zygotic embryos in seeds in terms of their overall size,morphology and internal cellular organization. Many cells inboth sexually-produced and somatic embryos contained numerouslipid-rich globular bodies. The wider significance of theseobservations is discussed with regard to their potential applicationsin eucalypt plantation biotechnology programmes. Copyright 2000Annals of Botany Company Eucalyptus nitens, shining gum, somatic embryo, tissue culture, ultrastructure, zygotic embryo     

Effect of Gibberellic Acid on the Ultrastructure and {alpha}-Amylase Activity of Aleurone Cells of Avena sativa L.

The -amylase activity and ultrastructure of aleurone cells inseeds of Avena sativa L. were studied using seed halves withembryo (embryo seeds) which had imbibed water and seed halveswithout embryo (embryo-less seeds) which had imbibed water withor without GA₃. -Amylase activity was detected in the aleurone layers of embryoseeds that had imbibed water and embryo-less seeds that hadimbibed GA₃-water. The ultrastructure of aleurone cells withdetectable -amylase activity showed marked changes in the roughsurfaced endoplasmic reticulum (rER), in the flattened sacculesforming stacks and in the aleurone grains. The progressive changesin the rER were as follows: first, the number of slender rERincreased; then, the inner space became wider and showed roundor oval profile; and finally, the rER became slender again witha reduced number of adhering ribosomes. The flattened sacculesforming stacks were appressed to the surface of aleurone grains.With time, they decreased in number and finally disappeared.In parallel with the decrease of flattened saccules, digestionof proteinaceous material inside the aleurone grains proceeded. (Received February 24, 1987; Accepted September 3, 1987)     

Callose Deposition During Megagametogenesis in Two Species of Oenothera

Callose deposits are present both in degenerating megasporesof the heteropolar tetrad in Oenothera hookeri and in degeneratingembryo sacs of the homopolar developing tetrad in O biennis.They are partially continuous with the cell wall and partiallyenclosed in the degenerating cytoplasm and show electron opaquebands within a less electron opaque material Vesicles calledcallose grains are present in the degenerating cytoplasm ofthe embryo sac in O biennis These show an electron opaque fibnllaror granular core surrounded by a halo of low electron opacity Similarities in fine structure between callose deposits of femalegametophytes which follow the degenerating pathway of development,and callose plugs present in pollen tubes during their growth,are discussed. Oenothera, evening primrose, megagametogenesis, megasporogenesis, callose, ultrastructure     

Analysis of Variation in the Rates of Germination and Early Seedling Growth in Tomato

Studies have been made on the time to germination and earlyseedling growth of the tomato Lycopersicon esculentum var. Potentateand Lycopersicon pimpinellifolium var. Red Currant, as wellas on derivative generations from their hybridization. Althoughtime to germination showed a genetic component, the relationshipsbetween different genotypes was much influenced by environmentalfactors. A marked maternal effect on time to germination dueto the varying seed sizes was noted while variation betweendifferent genotypes of similar seeds size was ascribed to anendospermic effect. The F¹ hybrid embryo with L. Pimpinellifoliumas maternal parent contained more cells than that of the parentbut the hybrid cmbryo with L. esculentum as maternal parentcontained a similar number of cells to that of the parent itself.It is suggested that the results for embryo size both supportAshby‘s assertion that embryo size may be important indetermining heterosis, and also Hatcher’s findings in1940, that heterosis for hypocotyl extension is found in hybridsfrom parcnts of different sized seeds provided the small seededvariety is the maternal parent.     

Nucleocytoplasmic Interaction During Maturation of the Egg of the Fern Histiopteris incisa (Thunb.) J. Smith

The maturing egg of Histiopteris incisa has many features incommon with that of Pteridium, with which the species may berelated. The nuclear evaginations are however conspicuous inrespect of the extensive development of internal membrane systems,involving dilatation of the perinuclear space. In some in stancesthe inner membrane of the nuclear envelope, the proliferationof which gives rise to the dilatations, was strikingly tenuous,possibly indicating an extensive interchange between the perinuclearspace and that part of the nucleoplasm sequestered in the evaginations.The evaginations frequently contain an aggregate of electron-opaquematerial resembling structurally and in staining propertiessimilar aggregates in Pteridium, but in contrast to Pteridiumthere was no evidence that these aggregates were organized inor near the nucleolus. Histiopteris inciso (Thunb.)J. Smith, Pteridophyta, egg maturation, nucleocytoplasmic interaction, ultrastructure     

An analysis of seed development in Pisum sativum XIX. Effect of mutant alleles at the r and rb loci on starch grain size and on the content and composition of starch in developing pea seeds

The effects of mutant alleles at the r and rb loci on starchgrain size and the levels of starch and amylose in developingpea (Pisum sativum L.) seeds have been examined. Four lines,near-isogenic except for genes at these loci, have been usedto show that both mutations reduce levels of starch throughoutembryo development and reduce levels still further when combinedin the ‘double mutant’. The reduction in starchcontent was due, at least in part, to a reduction in starchgranule size. Although the proportion of starch in mature embryoswas similar in the rrRbRb and RRrbrb lines, the starch contentdiffered between these two lines during development, as a percentageof embryo dry weight. This difference was due to a reductionin the absolute growth of the embryo caused by the rb mutation.Lines homozygous for the mutant r allele with either wild-type(RbRb) or mutant (rbrb) alleles at the rb locus contained increasedproportions of amylose in their starch throughout development,due to a reduced production of amylopectin. The presence ofthe rb mutation, however, also reduced the amount of amylosein relation to the reduction in total starch levels. Mutantalleles at both loci also reduced starch levels in the testaduring development, the reduction due to rb being more extreme.Reciprocal crosses showed a maternal effect of the rb mutationon final seed size and on the absolute amount of starch in theembryo. Key words: Pisum sativum L., seed, starch, development, mutant     

Ultrastructural Changes in Chloroplasts of Quercus rotundifolia Lam. in Response to Evernic Acid

The amount of total chlorophyll, chlorophylls a and b as wellas the ratio of a to b decreased in chloroplasts isolated fromQuercus rotundifolia leaves, kept for 17 d in a solution of35.5 µM evernic acid in 1 mM Na HCO₃, when compared withthe chloroplasts of control leaves (kept in NaHCO₃). The chloroplastsin the spongy parenchyma were smaller and the amount of starchand plastoglobuli lower. The number of grana per chloroplastsection, the number of thylakoids per grana and the height ofgrana stacks were also less in the chloroplasts of leaves treatedwith evernic acid. Quantitative ultrastructural differenceswere determined by means of electron microscopy and image analysistechniques. Quercus rotundifolia Lam., chloroplasts, ultrastructure, lichens, evernic acid     

Nucleic Acid Chemistry of the Ilyanassa Embryo

A morphological study of Ilyanassa embryogenesis was made fromsections of several stages and the nucleic acid content wasmeasured throughout the course of development. The kineticsof uptake and incorporation of radioactive precursors into RNAwas determined. The RNAs synthesized at different stages ofembryogenesis were fractionated by methylated albumin kieselguhrchromatography, and the polyadenylation of RNA during developmentwas measured. From these studies it was concluded that the Ilyanassa embryo,beginning with the 4-cell stage, synthesizes dRNA, rRNA, 4SRNA, and 5S RNA throughout all stages of development. The extensivesynthesis of dRNA and the higher proportion of RNAs polyadenylatedby the pregastrula embryo was considered to be of particularsignificance. The localization of nucleic acids and nucleic acid precursorsin the polar lobe and the effect of removing the polar lobeon nucleic synthesis was reviewed. Recent work on the ultrastructure of the Ilyanassa egg was reviewedin relation to the problem of ooplasmic segregation. The discoveryof a new cytoplasmic organelle in the Ilyanassa oocyte was reviewed.     

Strip-shaped Projections at the Cytoplasmic Face of the Outer Membrane of the Generative Cell in Amaryllis belladonna

Strip-shaped projections are present at the cytoplasmic faceof the outer membrane of the generative cell in Amaryllis belladonna.This outer membrane is actually the inner plasma membrane ofthe vegetative cell which surrounds the generative cell. Theprojections are situated in groups and arranged parallel toeach other. Their predominant orientation is perpendicular tothe long axis of the generative cell. The projections are approximately35 nm high, and on average equally spaced 40 nm apart. Theirmaximum observed length, estimated from grazing sections ofgenerative cells, is 250 nm. Generative cell, outer membrane, Amaryllis belladonna, ultrastructure     

Ageing in Triticum durum Seeds: Behaviour of Embryos and Endosperms from Aged Seeds as Revealed by the Embryo-transplantation Technique

The embryo-transplantation technique has been used to studythe behaviour of Triticum embryos and endosperm during the courseof ageing of 1-, 2-, 3-, 4-, and 5-year-old seeds. The resultsshow that the ageing process in Triticum seeds is a progressivephenomenon. It affects both embryo and endosperm as shown by(a) the behaviour of old embryos transplanted on young endosperms,and (b) the deleterious effects of aged endosperms on youngembryos.     

The Golgi apparatus in the endomembrane-rich gastric parietal cells exist as functional stable mini-stacks dispersed throughout the cytoplasm

Background information. Acid‐secreting gastric parietal cells are polarized epithelial cells that harbour highly abundant and specialized, H⁺, K⁺ ATPase‐containing, tubulovesicular membranes in the apical cytoplasm. The Golgi apparatus has been implicated in the biogenesis of the tubulovesicular membranes; however, an unanswered question is how a typical Golgi organization could regulate normal membrane transport within the membrane‐dense cytoplasm of parietal cells. Results. Here, we demonstrate that the Golgi apparatus of parietal cells is not the typical juxta‐nuclear ribbon of stacks, but rather individual Golgi units are scattered throughout the cytoplasm. The Golgi membrane structures labelled with markers of both cis‐ and trans‐Golgi membrane, indicating the presence of intact Golgi stacks. The parietal cell Golgi stacks were closely aligned with the microtubule network and were shown to participate in both anterograde and retrograde transport pathways. Dispersed Golgi stacks were also observed in parietal cells from H⁺, K⁺ ATPase‐deficient mice that lack tubulovesicular membranes. Conclusions. These results indicate that the unusual organization of individual Golgi stacks dispersed throughout the cytoplasm of these terminally differentiated cells is likely to be a developmentally regulated event.     

Pathway of Photosynthate Transfer in the Developing Seed of Vicia faba L: A Structural Assessment of the Role of Transfer Cells in Unloading from the Seed Coat

Photosynthate movement within the coat of the developing seedof Vicia faba occurs radially inward from the restricted vascularsystem and laterally through the non-vascularized region ofthe seed coat prior to exchange to the seed apoplast. Thin-walledparenchyma/transfer cells line the entire inner surface of theseed coat and thus are located at the terminus of the photosynthatetransfer pathway. The principal cellular route of transfer withinthe seed coat and the role of the thin-walled parenchyma/transfercells in membrane exchange to the seed apoplast has been investigated.Sucrose fluxes, computed from estimates of the plasma membranesurface areas of the cell types of the pathway, the plasmodesmatalcross-sectional areas interconnecting contiguous cells and theobserved rate of sucrose delivery to the embryo indicate thatsieve element unloading and subsequent transfer to the thin-walledparenchyma/transfer cells is through the symplast. For the cellsof the ground tissue, plasmodesmatal density is consistentlyhigher on their anticlinal walls. This observation supportsthe reported pattern of lateral transfer through these tissuesin the non-vascular regions of the seed coat. Wall ingrowthsare initiated sequentially in the thin-walled parenchyma cellsto maintain 1–3 rows of thin-walled parenchyma/transfercells. The development of these wall ingrowths results in a58% increase in the plasma membrane surface area of these cellsand provides them with the capacity to act as the principalcellular site for membrane exchange of sucrose to the seed apoplast.This cellular route of symplastic transfer from the sieve elementsto the ground tissues where membrane exchange to the seed apoplastoccurs is consistent with that reported for Phaseolus vulgaris Key words: Cellular pathway, photosynthate transfer, transfer cell, Vicia seed coat     

Beneficial Effects of Moist Chilling on the Seeds of Black Spruce (Picea mariana [Mill.] B.S.P.)

Moist chilling (cold stratification) is well-known as a simpleand effective means of overcoming physiological seed dormancyand enhancing germination of many temperate tree and shrub species.However, the apparent activation of intracellular repair mechanismsin non-dormant black spruce (Picea mariana) seeds followingmoist chilling, has not previously been reported. The presentcontribution records the beneficial effects of moist chillingafter accelerated ageing (40°C/98% relative humidity) for3–10 d of black spruce seeds. This treatment was maximallyeffective after 7 d of accelerated ageing, increasing germinationfrom 43 to 61%. Ultrastructural examination of embryo cellsindicated that intracellular activation occurred during moistchilling even when seeds had not been subjected to acceleratedageing, and that the increasing levels of damage accumulatingover the 3–7 d period of ageing were substantially reversedduring subsequent moist chilling. The results of cold stratificationof non-dormant seeds are discussed in terms of the value ofthis practice in nursery seedling production from seeds thatare of less than the highest quality.Copyright 2000 Annals ofBotany Company Accelerated ageing, black spruce, cold stratification, embryonic axes, germination, moist chilling, Picea mariana, repair mechanisms, ultrastructure     

Isolation and Characterization of Components of the Lytic Compartment of Several Plant Tissues Including Separations with a Zonal Rotor

The ultrastructural localization of acid phosphatase in cellsof pea and mung bean root tips, wheat coleoptiles, cauliflowerflorets and potato shoots, occurred in association with theinner surface of membranes of vacuoles, within multivesicularbodies and in the Golgi stacks and associated vesicles. Tissue homogenization techniques were varied and behaviour ofcomponents of the lytic compartment in density gradients ofvarious types was systematically examined. A basic fractionationmethod, employing linear Ficoll density gradients, was developedand modified for routine use with the various plant tissues,and the results compared with those obtained on sucrose gradients.Two predominant bands of components of the lytic compartmentwere isolated; one of these equilibrated at a density of 1·06g cm^–3 and contained vacuoles and tonoplast components,the other band, with an isopycnic equilibrium density of 1·11g cm^–3, was rich in mitochondria, Golgi stacks and associatedvesicles. Sucrose gradients gave broadly similar separations,but of lower resolution, and with increased density of the components.Prior fractionation of homogenates by differential centrifugationgave poor subsequent separations of the lytic compartment indensity gradient ultracentrifugation. The bulk isolation of components of the lytic compartment frompotato shoots and cauliflower curd, using a zonal density gradienttechnique, is described and its possible applications are discussed. An improved method of rapid fixation of fractions enabled theuse of a modified Gomori lead-salt method for ultrastructurallocalization of ß-glycerophosphatase activity in isolatedelements of the lytic compartment. Lytic component, Golgi apparatus, density gradient fractionation, ultrastructure, acid hydrolases, ß-glycerophosphatase, Pisium sativum L., Phaseolus aureus L., Triticum aestivum L., Solanum tuberosum L., Brassica oleracea L.     

Ultrastructure of the Developing and Fertilized Embryo Sac of Amaranthus hypochondriacus L.

Amaranthus hypochondriacus embryo sac development was investigatedbefore and after fertilization. During the early stages of development,the young embryo sac displays three antipodal cells at the chalazalpole that degenerate very early in the maturation process, beforethe synergids and egg cell are completely differentiated. Themature embryo sac is composed only of the female germ unit.The synergid cells organize a filiform apparatus accompaniedby the presence of mitochondria and dictyosomes with numerousvesicles. The involvement of the synergids in transport andsecretory functions related to pollen tube attraction and guidance,are discussed. The egg cell is located at the micropylar polenear the synergids and displays exposed plasma membranes atthe chalazal pole. The fertilized egg cell does not exhibitmarked changes after fertilization except for the closure ofthe cell wall. The central cell is the largest cell of thisvery long embryo sac. The fused nucleus is close to the eggapparatus before fertilization and displays a remarkable chalazalmigration after gamete delivery. The ultrastructure of the centralcell cytoplasm and the numerous wall ingrowths around this cellsuggest an important role in nutrient transportation. Aftergamete delivery, the embryo sac displays electron dense bodiesthat aggregate within the intercellular space between the synergids,egg cell and central cell. These bodies, that appear in theembryo sac of several plants, are probably involved in gametedelivery for double fertilization. The possibility of biparentalinheritance of mitochondria in this plant is also discussed.Copyright 1999 Annals of Botany Company Amaranthus hypochondriacus, grain amaranth, embryo sac, fertilization.     

Studies on the Sporogenous Lineage in the Moss Timmiella barbuloides IX. Development of the Tapetum

The ultrastructure of tapetal cells in Timmiela barbuloideswas investigated in relation to events of sporogenesis. Aftertheir establishment both internally and externally to the sporogonialinitials, tapetal cells enlarge and assume a permanently polarizedorganization after completion of meiosis. A large vacuole isformed in the cell region distal to the spore sac, the nucleusbecomes centrally located, and amyloplasts lie in the cytoplasmadjacent to the spore sac. An extensive endomembrane systemdevelops in tapetal cells during the stage of exine depositionin spore tetrads. Sheets of rough endoplasmic reticulum developfirst around the nucleus then also in close proximity to theplasma membrane abutting the spore sac. Concomitantly, interveningdictyosomes produce a variety of vesicles. Unusual structureswith vesicle-like profiles also occur in the inner tapetum cellwalls abutting the spore sac. At the same time most of the starchis lost from the plastids in which grana-fretwork systems develop.A massive secretion of extremely electron-opaque material isassociated with perine deposition onto the free spore surfaces.Degeneration of the tapetal cells during the terminal stagesof spore maturation is marked by distortion of the organelles,increase in vacuolation and the appearance of electron-opaquematerial between the sheets of endoplasmic reticulum.Copyright1994, 1999 Academic Press Bryophytes, endomembrane dynamics, Timmiella, ultrastructure, development, tapetum     

An Analysis of Seed Development in Pisum sativum III. The Relationship Between the r Locus, the Water Content and the Osmotic Potential of Seed Tissues in vivo and in vitro

The water content and osmotic potential (₂) of the differentparts of the pea fruit have been measured during developmentof the seed in two lines near-isogenic except for the r locus.During the early development of both genotypes, the testa possesseda more negative ₂ than either embryo, endosperm or pod while,at stages when liquid endosperm was present, the embryo alwaysmaintained ₂, more negative than the endosperm. A clear effectof the r locus on water content and ₂ was only observed in embryotissue — wrinkled (rr) embryos possessing more water andmaintaining a more negative ₂ than round (RR) for most of thedevelopmental period studied. The more negative ₂ of wrinkledembryos correlated with their greater uptake of water in vivo. When cultured in vitro, the embryos of both genotypes showedmaximum growth (fresh or dry weight) if 10 per cent sucrosewas added to the medium (equivalent to about — 1.2 MPa).Round embryos, however, increased in weight more than wrinkledat all sucrose concentrations examined. Cultured embryos maintaineda similar or more negative ₂ than their surrounding medium;wrinkled more negative than round. Embryo culture, osmotic potential, Pisum sativum, pea, r locus, seed development, tissue culture, water content     

Cytochemical localization of ATPase plasma membrane and acid phosphatase by cerium-based method in a salt-adapted cell line of Pisum sativum

In order to study a possible application of cerium-based techniquesin plant cells, ATPase and acid phosphatase activities havebeen compared in two cell lines of Pisum sativum calli, onesensitive to NaCI and the other selected to be grown under salinity(85 mM NaCI). ATPase activity was unchanged and localized inthe plasma membrane of both cell lines. Acid phosphatase activitywas significantly increased in the salt-selected line and localizedin the cell walls, Golgi complex, multivesicular bodies andvacuoles. These results indicated a possible involvement ofboth activities in the maintenance of cell growth in the selectedline under saline conditions. Key words: Acid phosphatase, Pisum sativum, plasma membrane ATPase, salt stress, ultrastructure     

A Comparative Ultrastructural Study of Cyanidium caldarium and the Unicellular Red Alga Rhodosorus marinus

The presence of nucleus, chloroplasts, mitochondria, microbodiesand a vacuole are confirmed in Cyanidium caldarium strain CCAP1355/1. Chloroplasts usually contain parallel rows of unstackedthylakoids surrounded by a peripheral thylakoid, although theconcentric arrangement has also been observed. The chloroplastenvelope consists of two closely appressed membranes which canonly be resolved after gluteraldehyde—osmium fixation.The chloroplast of Rhodosorus marinus also contains parallel,unstacked thylakoids surrounded by a peripheral thylakoid but,in addition, a prominent, stalked pyrenoid projects into thecytoplasm and is bounded by both the peripheral thylakoid andthe chloroplast envelope. This structure is covered by a capof starch grains and contains membranous vesicles in the matrix.Phycobilisomes are absent from the chloroplasts of both algae.The recognition of C. caldarium as a rhodophyte is supportedby these observations. Cyanidium caldarium, Rhodosorus marinus, chloroplast ultrastructure