Cytochemical Localization of Phenolic Compounds in Columella Cells of the Root Cap in Seeds of Brassica napus--Changes in the Localization of Phenolic Compounds during Germination

Changes in the localization of phenolic compounds were investigatedin columella cells of embryonic roots in 1-year-old Brassicanapus seeds during 48 h of imbibition and germination. In dry,dormant seeds, phenolic compounds were located in the apoplasticcompartment between the cell wall and plasmalemma in the outermostlayer of the columella. These apoplastic phenolic deposits disappearedduring the activation processes associated with imbibition andgermination, but new deposits appeared successively in the nucleus,ER cisternae, protein bodies and on the outer surface of theroot cap. A large number of phenolic deposits were observedin the outermost part of the columella, becoming less frequenttowards the initial centre. Their appearance coincided withrestoration of the ER and immediately preceded cytological activation.After the primary root had emerged from the seed coat, depositsof phenolic compounds disappeared from the cytoplasm, but simultaneouslyappeared in the vacuoles. Copyright 1999 Annals of Botany Company Brassica napus var. oleifera, root columella, germination, phenolic compounds localization.     

Fine Structural Details of Tannin Accumulations in Non-Dividing Cambial Cells

The fine structure of autumn cambial cells of Aesculus hippocastanumand Populus x euramericana reveals the presence of electron-densebodies, other than lipids, in both the cytoplasm and vacuoles.These deposits are identified as tannins following a cytochemicaltest with ferrous sulphate. Small tannin bodies associated withthe strands of ER and inside small vesicles are often evidentin the cytoplasm. The cytoplasmic tannins are deposited intovacuoles by fusion of the tannin-containing vesicles with thetonoplast. The positive reaction of tannin inclusions with periodicacid-thiocarbohydrazide-silver proteinate suggests their polysaccharidicnature. The observations support the role of ER in tannin synthesisand deposition into the central vacuole. Aesculus hippocastanum, Populus x euramericana, cambial cells, tannin, vacuoles     

Cytoplasmic and Apoplastic Location of Phenolic Compounds in the Covering Tissue of the Brassica napus Radicle Between Embryogenesis and Germination

The localization and intensity of cytoplasmic and apoplasticdeposits of phenolic compounds in Brassica napus L. change betweenembryogenesis and 36 h after seed germination. In the late stageof embryogenesis there were no phenolic compounds that wouldbe precipitated with caffeine, located either in the cytoplasmor outside the plasmalemma. Seeds collected at this stage rotduring germination. During seed maturation phenolic compoundswere localized in small vesicles which correspond to vesicular-shapedendoplasmic reticulum (ER) characteristic of this stage. Thiswas followed by slightly larger deposits in vacuoles, and inmature seed dark deposits accumulated outside the plasmalemma.In these dormant seeds the deposits were thus mostly betweenthe plasmalemma and the cell wall. After 3 h in water such darkdeposits appeared outside the cell wall on the embryo surface.After 6 h the cytoplasmic deposits were very few, and after24 h deposits reappeared in the round vesicles and long ER cisternae.After 36 h, when the emerging radicle and hypocotyl were 3 mmlong, there were large deposits of phenolic compounds in thevacuoles of various sizes. The occurrence of these depositsparalleled the previously demonstrated waves of embryo activityat the same stages of development, such as mitoses, synthesisof DNA, RNA, and protein, and mobilization of storage material. Embryogenesis, phenolic compounds, germination, seedling     

Ontogenesis of Tannin Coenocytes in Sambucus racemosa L. II. Mother Tannin Cells

Tannin coenocytes develop from mononucleate tannin mother cells.The process occurs within the whole of the first (youngest)internode and its development can be divided into three stages.In stage I the MTC is isodiametric and similar to the surroundingcells of the flank meristem, being present in the ninth celllayer from the apex surface. The nucleus becomes lanceolateand elongates, and large cytoplasmic vacuoles appear. A twofoldelongation of both the cell and nucleus continues in the secondstage, the cell-nucleus ratio indicating that it is due to theenlarged vacuole, which pushes a thin layer of cytoplasm closeto the cell wall. In this layer of cytoplasm dilated ER cisternaoccur together with small and large vacuoles, a fusion of thevacuoles increasing their volume. Such cells are diploid inspite of larger nuclear volume and rough structure of its chromatin. Sambucus racemosa L., tannin cells, development, ontogenesis     

Development of Air Blisters in Pilea cadierei Gagnep. and Guillaumin

An ultrastructural study was conducted to determine the originand development of the air blisters in leaves of Pilea cadierei.At early developmental stages (characterized visually by ananthocyanin flush to the blistered region, in leaves 0.5–1.8cm long) two epidermal layers separated by a small intracellularspace from a sub-epidermal layer of phenolic cells are observed.Osmiophilic phenol aggregates occur in the central vacuole,minivacuoles (or provacuoles) and in the endoplasmic reticulaof the phenol cells. Plastids in the phenol-containing cellshave extremely osmiophilic thylakoids, perhaps indicative ofphenol—caffeine complexes in the plastid. Later developmentalstages (characterized by a silver coloration in the air blisters)show an increase in the intracellular space, due at least inpart to the apparent collapse of the second epidermal layerand breaking of cell connexions. Phenol-containing cells havetwo different types of electron-opaque deposit: a grey granulardeposit and a continuous caffeine—phenol agglutination.Plastids are shade type and are much less opaque than in earlierstages. Several possible reasons for the presence of air blistersin many genera are discussed. Pilea cadierei, air blisters, ultrastructure, phenolics, tannin     

Dilated Endoplasmic Reticulum Cisternae in Differentiating Xylem of Minor Veins of Mimosa pudica L. Leaf

The differentiating tracheary elements in the xylem of minorveins in Mimosa pudica L. contain, as is usual, complete nucleateprotoplasts. Within the latter, dictyosomes with associatedvesicles and endoplasmic reticulum (ER) are prominent components.The ER cisternae show an uncommon feature of developing voluminousdilations accumulating finely fibrous material. Similar dilatedER cisternae occur in parenchyma cells associated with the trachearyelements. Most of the dilated cisternae are elongated, taperingat both ends, and almost circular in transections. They varyin size. The largest measured was 10 µm long and 3 µmwide. In the tracheary elements the cisternae break down asthe protoplast disintegrates. For a time, mature cells containfibrous material, apparently the product of the dilated cisternae,at least in part. In the parenchyma cells the dilated cisternaeare released into the vacuoles after the associated trachearyelements reach maturity. They become structurally modified anddisintegrate. The timing in the appearance and disintegrationof the dilated ER cisternae suggests that these structures havesome function with reference to the differentiation of trachearyelements in the xylem.     

Compartmentation of Phenolic Compounds and Phenylalanine Ammonia-Lyase in Leaves of Phyllanthus tenellus Roxb. and their Induction by Copper Sulphate

The compartmentation of phenolic compounds in mature leavesof Phyllanthus tenellus and their induction by copper sulphatewere analysed at histological and subcellular levels. Lightand electron microscopy studies demonstrated that the vacuolesof spongy cells were the main sites of phenolic accumulation.Spraying plants with copper sulphate induced punctated lesionsformed by groups of necrotic cells which accumulated brownishsubstances. Histochemical tests and fluorescence microscopyanalysis of the sprayed leaves indicated that the phenolic compoundsincreased in spongy cells within the lesions. Ultrastructuralanalyses showed that 3 h after elicitation, the organelles ofthe cells within the lesion started to collapse and the contentof phenolic substances increased in the vacuole of spongy cells.Antibody against phenylalanine ammonia-lyase (PAL) from parsleycross-reacted with the crude extract of P. tenellus leaves.Two isoforms, one of 65 kD and the other of 66 kD, were identified.Immunocytochemical studies showed that PAL was synthesized inthe palisade and spongy cells, mainly in the cytoplasm and chloroplasts.The phytotoxicity of Cu²⁺ions induced the accumulation of PALin sub-cellular compartments of palisade cells. PAL accumulationstarted to increase 3 h after elicitation and reached a maximumafter 6 h, decreasing 12 h post-induction. The increase of PALwas more evident in cells within the necrotic punctated regionsthan in surrounding cells. Since the vacuole of palisade cellsdid not accumulate phenolic compounds, the in situ studies suggestedthat the end products of PAL synthesis play a role in palisadecell wall reinforcement or might accumulate in other tissues.The symptoms induced by copper sulphate suggest that this abioticelicitor may be a useful tool in the understanding of the regulationof biosynthetic phenolic pathways inP. tenellus . Copyright2000 Annals of Botany Company Cell death, copper sulphate, heavy metal, immunolabelling, phenolic compounds, phenylalanine ammonia-lyase,Phyllanthus , transmission electron microscopy, ultrastructure     

Developmental Features of the Primary Phloem in Phaseolus vulgaris L.

Certain developmental features of the primary phloem were examinedin Phaseolus vulgaris L., chiefly by the use of the pulvinusat the base of the petiole. The cells included in the studywere the sieve element, the companion cell, and the tannin cell.In the sieve element, the sieve plate shows the usual sequenceof conversion of plasmodesmatal canals into pores. The endoplasmicreticulum, which appears as flat cisternae associated with ribosomesin younger cells, later becomes in part stacked and in partaligned parallel with the walls as a network. The stacked ERprecedes the anastornosing parietal ER in time of development,but the parietal ER persists longer. Of the two forms of P-proteincharacteristic of a number of Fabaceae, the crystalline bodyappears considerably in advance of the body composed of tubules.Neither form of P-protein disperses completely in the maturecell, although the crystalline protein may spread out into aggregatesof fine fibrils. The companion cells show the typical denseprotoplasts and branched plasrnodesmatal connections with thesieve elements. The vacuome of these cells is dispersed intonumerous small vacuoles, many of which appear to be concernedwith autophagic digestion of protoplasmic material. The tannincells have large vacuoles in which the tannin material is located.The cells form vertical series in which the end walls becomeperforated.     

ONTOGENY,HISTOCHEMISTRY AND FINE STRUCTURE OF CELLULAR INCLUSIONS IN VEGETATIVE CELLS OF ANTITHAMNION DEFECTUM (CERAMIACEAE,RHODOPHYTA)1

The ultrastructure and histochemistry of developing and mature cell inclusions in vegetative cells of Antithamnion defectum Kylin were examined. Those studied were chloroplast inclusions, cytoplasmic crystals and spherical bodies within the vacuole. Chloroplasts of mature vegetative cells contain an interthylakoidal, apparently noncrystalline deposit of undetermined chemical identity. The bodies are parallel to the long axis of the plastid, are square (0.13 μm) in cross-section, and up to 3 μm long. Spherical vacuolar bodies (0.5–1.5 μum diam) are formed during early stages of vacuole formation by accumulation of protein deposits in swelling endoplasmic reticulum (ER) cisternae. Swelling of smooth ER contiguous to the ER containing the deposits results in the vacuole enclosing the spherical bodies. In mature cells, vesicles appear to be secreted into the preformed vacuole. Cytoplasmic proteinaceous crystalloids develop without a bounding membrane and may serve as protein reserves.     

The function of the aleurone layer during galactomannan mobilisation in germinating seeds of fenugreek (Trigonella foenum-graecum L.), crimson clover (Trifolium incarnatum L.) and lucerne (Medicago sativa L.): A correlative biochemical and ultrastructural study

Summary The reserve endosperm galactomannans of fenugreek (Trigonella foenum-graecum L.), crimson clover (Trifolium incarnatum L.) and lucerne (Medicago sativa L.) are broken down to free galactose and mannose in dry-isolated endosperms (devoid of embryo) incubated under germination conditions. Breakdown is prevented by inhibition of protein synthesis or of oxidative phosphorylation in the aleurone layer. Resting aleurone cells contain inter alia a large number of ribosomes more or less regularly distributed in the ground plasma. At the onset of germination, before galactomannan breakdown begins, polysomes are formed and seem, at least partly, to become associated with vesicles and flat cisternae both probably newly formed and derived from ER. Concurrently with galactomannan breakdown in the reserve cells, wall corrosion occurs in the aleurone layer, the contents of the aleurone grains disappear and the rough vesicles and cisternae proliferate. Later a large central vacuole is formed which incorporates smaller vacuoles emerging from the cytoplasm, and at the same time the rough ER vesicles and cisternae become highly distended.It is concluded that the cells of the aleurone layer are responsible for the synthesis and secretion into the storage cells of the enzymes necessary for galactomannan degradation. The physiology of galactomannan breakdown is compared and contrasted with that of starch mobilisation in the endosperm of germinating cereal grains.This is part three in a series of papers dealing with galactomannan metabolism. Part two: Planta (Berl.) 100, 131–142 (1971).     

山茱萸果实发育过程中单宁物质的分布与积累特征

选取不同发育时期的山茱萸果实作为研究对象,采用果实形态观察法、显微及超微技术、组织化学定位法以及紫外分光光度计法对山茱萸果实发育过程中单宁物质分布及积累特征进行观察分析,并以单因素ANOVA检验不同发育时期单宁含量的差异,以揭示单宁物质在山茱萸果实发育中的变化规律,为山茱萸果实涩味调控机制研究提供理论依据。结果表明:(1)山茱萸果实发育过程中果皮颜色和果实体积变化明显,可将其发育过程划分为幼果期、中果期、成熟期3个时期;单宁物质主要分布在山茱萸果实中果皮的单宁细胞中。(2)在山茱萸果实发育过程中单宁细胞数目呈先增后减的变化趋势,幼果期单宁细胞从无到有,随着果实发育单宁细胞数目不断增多,至中果期单宁细胞数目开始减少。(3)单宁含量的变化规律与单宁细胞数目的变化一致,单宁含量在花后120 d时达到最多,随后逐渐减少。(4)单宁物质首先在细胞质的小液泡中积累,中央大液泡形成后则为单宁物质积累的主要场所,其积累形态主要有颗粒状、不规则状和板块状3种;单宁细胞中线粒体数目较多,中果期后期及成熟期在中央大液泡液泡膜附近有电子致密物质积累。研究认为,山茱萸果实中中果皮薄壁细胞为单宁物质积累的专属细胞,即单宁细胞,单宁物质的合成运输与液泡、囊泡以及线粒体的作用密切相关;成熟期山茱萸果实总单宁含量降低,涩味降低,表明单宁物质积累的动态变化与植物对环境的适应性和果实涩味息息相关,可结合代谢组和转录组的方法对山茱萸果实中单宁物质的合成机制进行进一步研究。     

Ultrastructure, Origin, and Composition of the Protein Bodies in the Ligule of Isoetes lacustris L.

The basal cells in the ligule of Isoetes lacustris contain numerousprotein bodies, the contents of which can be digested enzymicallyby pronase and are stained red by treatment with ninhydrin Schiff'sreagent. Two types of protein bodies can be distinguished ultrastructurally:spherically-shaped bodies with granular contents and spindle-likebodies with fibrillar contents. Both are ensheathed by singlemembranes and do not show any solid inclusions within theirmatrix. The protein bodies probably arise from dilatation of the endoplasmicreticulum (ER) cisternae. This conclusion is based upon threeobservations: (a) The protein bodies occasionally show membranecontinuity with the ER; (b) ribosomes and polysomes are frequentlyattached to the protein-body membranes; (c) the contents ofthe protein bodies and of the dilated ER cisternae show similarultrastructural features. The dilatation of the ER cisternae is assumed to be a resultof protein accumulation in the intracisternal space. Based upon the results of polyacrylamide gel electrophoresis,it is likely that the spherically-shaped protein bodies storepredominately two proteins with molecular weights of 51300 and55800 D, while the spindle-like bodies store two proteins withmolecular weights of 92000 and 98000 D. The results presented do not permit a definite conclusion regardingthe function of the ligule of Isoetes lacustris but it is suggestedthat it may have a nutritive role. Isoetes lacustris L., ligule, protein bodies, endoplasmic reticulum, ultrastructure     

Migration of Calcium and Its Role in the Regulation of Seismonasty in the Motor Cell of Mimosa pudica L

Volume and conformational changes of the contractile tannin vacuoles of the abaxial motor cells of the primary pulvinus of Mimosa pudica L. parallel the seismonastic leaf movement. Since such changes in cells and organelles of animal systems are often regulated by calcium, we studied Ca²⁺ movement in the motor cells and tissue. By fixation with Lillie's neutral buffered formalin, followed by staining with alizarin red sulfate (ARS), calcium was localized in the tannin vacuoles of the motor cells of the primary pulvinus. After treatment with ethylenediaminetetraacetate, 8-hydroxyquinoline, and several other calcium-complexing or extracting agents, the color reaction due to alizarin red sulfonate was no longer present. By using an analytical method, it was shown that the effluent from stimulated pulvini has significantly more Ca²⁺ than that from unstimulated controls. Ten millimolar LaCl₃ inhibits recovery of the tannin vacuole in vivo in 10 mm CaCl₂ or in distilled water. Quantitative data obtained by microspectrophotometry demonstrated calcium migration during the bending movement of the primary pulvinus. In the adaxial motor cells a small amount of calcium migrates from the tannin vacuole, and calcium on the cell wall moves to the central vacuole. In the abaxial half, a large amount of calcium from the tannin vacuole moves to the central vacuole of the motor cell. It is probable that the calcium binds to the microfibrillar contents of the central vacuole. These observations support the contention that Ca²⁺ migrates between the surface of the tannin vacuole and the inside of the central vacuole. The recovery and maintenance of the tannin vacuole in the spherical form may play a role in maintaining turgor in the motor cells of the abaxial half of the primary pulvinus of Mimosa.     

Auffällige ER-Konfigurationen in Protein-Polysaccharidschleim-sezernierenden pflanzlichen Drüsen

In the ovary of Aptenia cordifolia and Platythyra haeckeliana placentary papillae produce a slime containing polysaccharides and proteins. These papillae show two types of conspicuous vacuoles enclosed by rough ER cisternae and complexes of concentrically arranged rough ER. The enclosed vacuoles probably play an important role in the accumulation of the polysaccharide-protein slime. In the case of storage vesicles (first vacuole type) derivates of the Golgi apparatus are enclosed by ER. In other instances (second vacuole type) ER cisternae which have lost their membrane-bound ribosomes seem to delimit protoplasmic regions free of organelles.

     

Cytoplasmic deletion processes during spermatogenesis in mosses

Comparative ultrastructural observations reveal that cytoplasmic deletion during spermatogenesis in Sphagnum and other mosses (Bryopsida) has two distinct phases. In young spermatids, Golgi-derived vesicles produce the mucopolysaccharide sheaths in which the gametes are liberated. Golgi bodies, however, play no part in removal of cytoplasm during gamete maturation. Rounding off of the cells during this process results in a 50% reduction in volume. Mid-spermatid stages in Sphagnum are characterised by the sequential loss of Golgi bodies and endoplasmic reticulum (ER) but no further diminution of the cytoplasm. The final stages of nuclear metamorphosis and chromatin condensation, in late spermatids, are marked by the sudden appearance, in the otherwise featureless central cytoplasm, of a membrane vesicle complex (MVC) comprising cisternae, tubules, and smooth and coated vesicles. Following repositioning of the MVC beneath the plasma membrane, rapid shrinkage of the cytoplasm is associated with the presence of vesicle fusion profiles at the cell surface. The MVC is considered to be intimately involved in cytoplasmic breakdown and loss. Acid phosphatase activity can be detected throughout spermatogenesis. Spermatogenous cells and young spermatids possess relatively low levels of the enzyme, restricted to the ER and perinuclear space, but particularly high levels occur in the MVC region of late spermatids of Sphagnum. The deletion process in Bryopsida is much more gradual than that of Sphagnum. Mid-spermatids contain sheets of ER, Golgi with small vesicles, and irregular cisternae associated with coated vesicles. Vacuoles derived either from dilation of the ER or the coated vesicle complexes gradually increase in size and number at the expense of the cytoplasm. During the early stages of chromatin condensation, a large central vacuole opens onto the anterior face of the gametes. Further discharge of vesicles continues throughout gamete maturation. A comparative survey of spermatogenesis in land plants indicates that cytoplasmic deletion is achieved in different ways in different groups. We speculate that the spermatozoids of the common ancestor of archegoniate plants probably possessed large amounts of cytoplasm. The deletion mechanisms may have originated from a contractile vacuole apparatus.     

Isolation of Vacuoles from Immature Apple Fruit Flesh and Compartmentation of Sugars, Organic Acids, Phenolic Compounds and Amino Acids

Vacuoles of immature apple fruit (Malus pumila Mill. var. domesticaSchneid.) were obtained by purification using Ficoll densitygradient centrifugation after lysis of the protoplasts by bothmild osmotic shock and the addition of EDTA and BSA. The recoverywas about 35% of the protoplasts. The isolated vacuoles hada mean diameter of about 100 µm. The distribution of sugars, organic acids, phenolic compoundsand amino acids in the vacuole, the cytoplasm and the free spacewas determined. Almost all of the fructose and glucose, themajor sugars of the tissue, were found in the vacuole. Sorbitolwas mainly located in the free space and the vacuole, and sucrosein the free space and the cytoplasm. More than 90% of the malicacid, the main organic acid, was located in the vacuole. Almostall of the phenolic compounds were also deposited in the vacuole. The volumes of the vacuole, the cytoplasm and the free spacein the whole tissue were calculated from the cell numbers ofthe whole tissue, the volume of the isolated protoplasts, andthe volume of the vacuoles present in the protoplast. The soluteconcentration in each compartment was estimated: vacuoles, 888mM; cytoplasm, 37 mM; free space, 57 mM. How these compartmentationsof solutes affected the translocation of sugars into the fruitand the cell expansion is discussed. ¹This paper is contribution A-159 of the Fruit Tree ResearchStation. (Received July 7, 1983; Accepted November 14, 1983)     

Secretory cells in Piper umbellatum (Piperaceae) leaves: A new example for the development of idioblasts

This work aims to investigate the origins and development of secretory cells in Piper umbellatum (L.) Miq. (Piperaceae) leaves as well as the course and the nature of their secretion. The results were compared with studies in oil-secreting cells of several species. Fully expanded fresh leaves were sectioned and subjected to different histochemical tests. Leaves in different developmental stages were fixed and processed for study under light and scanning and transmission electron microscopy techniques. The secretory cells show mixed secretion made up of hydrophobic (oleoresin) and hydrophilic (phenolic compounds and alkaloids) compounds. Secretory cells originate either from the protodermis or the ground meristem. The growth of these cells occurs primarily by increasing the volume of the central vacuole, which corresponds to an extraplasmatic space connected to a protuberance of the wall. Electron-opaque compounds are observed initially in leucoplasts, while electron-dense compounds occur in small vesicles in the cytoplasm. Both are accumulated in the central vacuole which is already developed. Besides the mixed chemical nature of the secretion identified in secretory cells of P. umbellatum leaves, these secretory cells differ from those that have already been described mainly because of the development of the central vacuole prior to the accumulation of the secretion.     

Localization of Phenolic Compounds in the Covering Tissues of the Embryo of Brassica napus (L.) during Different Stages of Embryogenesis and Seed Maturation

During embryogenesis and maturation of an embryo the tissuescovering it produce phenolic compounds the localization of whichchanges during maturation of the embryo. In the ovary containinga globular embryo, phenolics are located in the epidermis ofthe integumentum externum and the innermost layer of the integumentuminternum. In the ovule at the stage at which heart- and torpedo-shapedembryos are present, phenolic compounds are visible in the stellarcells, the innermost cells of the integumentum internum andthe endosperm. In hard, green seeds, after the integumentuminternum and layers over the stellar cells gradually disappear,the remaining tissue contains cell walls impregnated with phenolics.Mature, black seeds contain only one distinct layer of cells—stellarcells, which, like the other compressed cell walls, are impregnatedwith phenolics. In this way they constitute a barrier betweenthe embryo and its environment.Copyright 1994, 1999 AcademicPress Brassica napus, seed coat, integumentum, phenolic compounds     

Lysosomal nature of plant vacuoles II. Acid hydrolases in the central vacuole of internodal cells of Charophyta

Contents of the central vacuole (cell sap) were separated frominternodal cells of Nitella and Chara. Most of the acid phosphataseand carboxypeptidase, which are marker enzymes for lysosomes,were detected in the separated cell sap. In contrast, most ofthe catalase, cytochrome oxidase, NADPH₂-cytochrome c reductase,and glucose-6-phosphate dehydrogenase were detected in partsof the cell other than the cell sap: which shows there is relativelylittle contamination of the cytoplasm in the separated cellsap. Enzymatic properties of the carboxypeptidase in Nitellaaxilliformis were similar to those of carboxypeptidases in higherplants and those of cathepsin A in animal lysosomes. These resultsindicate that the central vacuole of Charophyta has some propertiesof the lysosome. ¹ Preliminary results of this paper were given in reference(5). (Received February 3, 1975; )     

Fine Structure of Nucellar Cells During Development of the Embryo Sac in Oenothera biennis L.

Developing nucellar cells in Oenothera biennis L. present distinctpatterns of differentiation at the chalaza and around the embryosac. The cytoplasm of nucellar cells surrounding the tetradof megaspores displays cytolysomes, lipid bodies and membranesof smooth ER enveloping different cytoplasmic components. Concomitantto the differentiation of the embryo sac the nucellar cellsconstituting these ‘parietal layers’ undergo cytoplasmicdegeneration with shrinkage and flattening. In addition to theregular nucellar cells the chalaza at this stage presents threeother types: One with pycnotic nuclei, paramural bodies andcytoplasm filled with polymorphic vacuoles containing membranes,granular or flocculent material and multivesicular bodies. Asecond cell type shows swollen perinuclear cisternae aroundtheir pycnotic nuclei and large cytoplasmic vacuoles accumulatingtannins. The third type of cells is characterized by large numbersof starch grains and advanced disorganization of cytoplasmicorganelles; these cells probably become reservoirs after death. Oenothera biennis L., evening primrose, embryo sac, nucellus, cytolysomes, ultrastructure