Brefeldin A induces reversible dissociation of the Golgi apparatus in the green algaMicrasterias

Summary The fungal metabolite brefeldin A (BFA) causes inhibition of cell growth inMicrasterias denticulata after 2 h incubation, combined with slight malformation of the cell shape. The BFA effects on cell development are accompanied by a gradual decrease in the number of Golgi cisternae and severe structural and morphological changes of the dictyosomes which are already visible after only 10 min exposure. When the treatment is prolonged the number of dictyosomes is markedly reduced, leading to almost complete loss of Golgi bodies, particularly in the young semicell. Groups of primary wall material-containing vesicles accumulated in areas of former dictyosomes, and previously unknown vesicular bodies are found. Restitution of almost normal dictyosomes occurs within 5 h when the cells are allowed to recover from BFA treatment.Micrasterias cells incubated in BFA at concentrations below 15 M maintain their ability to divide over several generations. Our results indicate that, of the various inhibitors of the secretory pathway tested against growingMicrasterias cells, BFA is the only drug which induces complete and reversible dissociation of dictyosomes in the growing semicell. This allows deductions about the function of the processes targeted by BFA during cell development inMicrasterias.Abbreviations BFA brefeldin A - CPA cyclopiazonic acid - ER endoplasmic reticulum - TM tunicamycin     

Morphology and function of the Golgi apparatus

The polymorphism of the dictyosomes in the root meristeme ofFagopyrum is connected with their various functions in secretory processes and cell differentiation. The dictyosomes containing vesicular dilatations of the cisternae, which in this object occur more frequently than in others, probably participate in a similar way as the Golgi apparatus of the animal cell in the formation of lysozomes, in the formation of elements belonging to the group of dense bodies analogical lysozomes. These bodies are present in large numbers in the cytoplasm of cells, containing dictyosomes with vesicular dilatations. The other forms of the dictyosomes reveal indications of their participation in the production of the carbohydrate material of the cell walls, like most dictyosomes of other plant objects. However, no fusion of the Golgi vesicles with the plasmalemma was observed. According to their morphological appearance the typical forms of dictyosomes were classified on the basis of their relationship to secretory processes. Simultaneously the morphology and function of the Golgi apparatus was compared in the animal and plant cell. Several morphological varieties of the dictyosomes of plant cells, observed after the action of pathogenic factors and the effect of the fixation procedures, were also noticed in small quantities in the cells of the investigated objects.     

Megasporogenesis in Arabidopsis thaliana L.: an ultrastructural study

 In this study, megasporogenesis of the plant model Arabidopsis thaliana was investigated by electron microscopy for the first time. The data described here could constitute a reference for future investigations of Arabidopsis mutants. During the beginning of meiosis the megaspore mother cell shows a polarity created by unequal distribution of organelles in the cytoplasm. Plastids accumulate in the chalazal region and long parallel saccules of endoplasmic reticulum, small vacuoles and some dictyosomes are found in the micropylar region. Plasmodesmata are abundant in the chalazal cell wall. The nucleus is almost centrally localized and contains a prominent excentric nucleolus and numerous typical synaptonemal complexes. After the second division of meiosis the four megaspores are separated by thin cell walls crossed by numerous plasmodesmata and do not show significant cellular organization. The young functional megaspore is characterized by a large nucleus and a large granular nucleolus. The cytoplasm is very electron dense due to the abundance of free ribosomes and contains the following randomly distributed organelles: mitochondria, a few short saccules of endoplasmic reticulum, dictyosomes and undifferentiated plastids. However, there is no apparent polarity, except for the distribution of some small vacuoles which are more abundant in the micropylar region of the cell. The degenerating megaspores are extremely electron dense and do not show any substructure. Received: 30 July 1998 / Revision accepted: 3 February 1999     

A Fine Structural Analysis of Auxin-induced Elongation of Cucumber Hypocotyls, and the Effects of Calcium Antagonists and Ionophores

The fine structure of epidermal cells, particularly in relationto dictyosomes, has been examined in different regions of dark-growncucumber hypocotyls and in response to auxin treatment, usingboth dot overlay and image analysis techniques. The most noticeablechange in cell structure along the hypocotyls is the increasein vacuolar volume. The volume fraction occupied by dictyosomesand secretory vesicles also increased, whereas that for mitochondriaremained relatively constant. During auxin treatment, the volumefraction for dictyosomes showed an increase after 30 min followedby a fall, whereas that occupied by secretory vesicles fellsteadily over 90 min. The number of cisternae per dictyosomeshowed some increase after 2 h of auxin treatment, althoughthe increase in dictyosomal material with cell expansion waslargely accounted for by an increase in the number of dictyosomes. Auxin-stimulated elongation growth of the hypocotyls was inhibitedby a range of calcium antagonists, chelators and ionophores.The most marked inhibitions were observed with calcium chloride,the chelator chlortetracycline and the ionophores verapamil,nigericin and monensin. Linear transducer experiments showedthat these compounds generally caused an immediate reductionin the rate of growth. Fine structural observations carriedout on epidermal cells showed the most obvious effects withmonensin and nigericin which caused dictyosomes and secretoryvesicles to swell. EGTA and LaCl₃ caused secretory vesiclesto accumulate around dictyosomes, while the ionophore A23187had little effect. The results suggest that the concentration of Ca²⁺ in the cytoplasmmay be critical for cell elongation. Compounds which chelateCa²⁺ appear to be more effective inhibitors of growth in theinitial acid-induced phase, whereas those which affect ionicgradients are more disruptive in the second phase.Copyright1993, 1999 Academic Press Calcium, Cucumis sativus hypocotyle, dictyosomes, elongation growth, indoleacetic acid, stereology     

Ultrastructural observations of maize root tips following exposure to monensin

Summary Epidermal and outer rootcap cells of maize root tips were treated with the sodium selective ionophore, monensin, and the ultrastructural changes were studied. In the presence of 10^–5 to 10^–3 M monensin, dictyosomes became distorted, cisternae separated from the stack, and secretory vesicles were released. Released secretory vesicles disappeard from the cytoplasm suggesting that their transport to, and fusion with, the plasma membrane was unaffected. Monensin did not inhibit cytoplasmic streaming of the outer rootcap cells. No new secretory vesicles were formed on the remaining dictyosomes or dictyosome fragments. In contrast to results with animal cells, swelling of plant dictyosome cisternae was observed only after fixation in glutaraldehyde-osmium tetroxide and not after fixation in potassium permanganate. Other cell components were not altered structurally by monensin. The effects of monensin on the Golgi apparatus were reversible, and dictyosomes were either repaired or new dictyosomes were formed after the root tips were removed from the monensin.Dictyosomes in epidermal cells reacted in the same manner as those in the rootcap except that numerous secretory vesicles remained in the cytoplasm, mostly in association with dictyosome fragments. Some secretory vesicles increased in size but no evidence of vesicle-vesicle fusion was noted. Cell plate formation was partially inhibited or blocked by monensin.Mention of a commercial or proprietary product in this paper does not constitute an endorsement of this product by the USDA.     

Development,structure, and occurrence of secretory trichomes ofPharbitis

Summary Secretory trichomes develop from epidermal cells on the leaf primordia and stem ofPharbitis nil. Following an initial growth phase, trichomes begin active secretion of a protein-carbohydrate mucilage. This mucilage covers the shoot apex and developing leaves ofPharbitis.The secretory cells possess cellular organelles in forms usually associated with actively secreting cells: many mitochondria, an elaborate network of rough endoplasmic reticulum (RER), many free ribosomes, and numerous dictyosomes. The role of the dictyosomes is twofold: 1. dictyosome vesicles bud coated vesicles which transport materials from the cell and, 2. dictyosome vesicles coalesce, forming large storage vesicles. The storage vesicles are surrounded by, and often in contact with, poculiform RER. The RER forms an interconnected network throughout the cytoplasm, extending from the nuclear envelope to the plasmalemma. Distended profiles of RER are frequently in direct contact with the plasmalemma. Thus, inPharbitis secretory trichomes, it is the coated vesicles and RER which are active in secretion export. These findings imply a secretory pathway which deviates from the usual pattern in glandular cells.Predoctoral fellow of National Science Foundation during part of the investigation.     

Fine structure of the wet,detached cell stigma of the orchid Dendrobium speciosum Sm.

Summary The stigmatic surface of the orchid Dendrobium speciosum is a cup containing detached cells suspended in a mainly carbohydrate mucilage. The fine structure of the detached cells and their organelles is indicative of secretory cells. The cells contain numerous mitochondria with well-developed cristae, dictyosomes containing extensive cisternae, an extensive network of rough and smooth endoplasmic reticulum and free polysomes throughout. There are many amyloplasts in the vicinity of the nucleus. Vesicles are seen arising from the dictyosomes and endoplasmic reticulum. The plasmalemma is undulating, and vesicles can be seen in its vicinity, giving the typical appearance of a granulocrine secretory system. Cetylpyridinium chloride (CPC) fixation to immobilise acidic carbohydrates detected a highly electron-opaque layer surrounding each cell and globules dispersed through the cell wall. The walls of the detached cells show irregular surface projections which are the remains of pitfields. Biochemical analysis showed that carbohydrates and arabinogalactan proteins are major components of the mucilage.     

Dictyosome polarity and membrane differentiation in outer cap cells of the maize root tip

Outer rootcap cells of maize produce large numbers of secretory vesicles that ultimately fuse with the plasma membrane to discharge their product from the cell. As a result of the fusion, these vesicles contribute large quantities of membrane to the cell surface. In the present study, this phenomenon has been investigated using sections stained with phosphotungstic acid at low pH (PACP), a procedure in plant cells that specifically stains the plasma membrane. In the maize root tip, the PACP also stains the membranes of the secretory vesicles derived from Golgi apparatus to about the same density that it stains the plasma membrane. Additionally, the membranes of the secretory vesicles acquire the staining characteristic while still attached to the Golgi apparatus. The staining progresses across the dictyosome from the forming to the maturing pole, thus confirming the marked polarity of these dictyosomes. Interestingly, the PACP staining of Golgi apparatus is confined to the membranes of the secretory vesicles. It is largely absent from the central plates or peripheral tubules and provides an unambiguous example of lateral differentiation of membranes orthogonal to the major polarity axis. In the cytoplasm we could find no vesicles other than secretory vesicles bearing polysaccharide that were PACP positive. Even the occasional coated vesicle seen in the vicinity of the Golgi apparatus did not stain. Thus, if exocytotic vesicles are present in the maize root cap cell, they are formed in a manner where the PACP-staining constituent is not retained by the internalized membrane. The findings confirm dictyosome polarity in the maize root cap, provide evidence for membrane differentiation both across and at right angles to the major polarity axis, and suggest that endocytotic vesicles, if present, exclude the PACP-staining component.     

The synthesis of phaseolin: a model for the study of the plant secretory pathway

Abstract

Phascolin, the major seed storage protein of common bean (Phaseolus vulgaris), has been for many years one of the main working horses for studying protein synthesis, trafficking and structural maturation in the secretory pathway of higher plants. Recently, phaseolin has been used as a tool to determine molecular interactions between chaperones and newly-synthesised wild-type or structurally-defective secretory proteins in plant cells. Despite the vast amount of information available on the structure and the cell biology of phaseolin, the determinants for its sorting to the vacuole are still unknown.     

Ultrastructural evidence for the existence of two types of neurosecretory cells in the abdominal ganglia of the chelicerate arthropod,Limulus polyphemus

Evidence suggesting the existence of two types of neurosecretory cells in each abdominal ganglion of Limulus polyphemus has been obtained by light and electron microscopy. After Helly fixation the two cell types are readily distinguished from other neurons by the Azan method, but they react weakly when stained by paraldehyde fuchsin. Type I cells are larger, more regular in shape, and found more anteriorly in each ganglion. They contain apparently cylindrical secretory granules, many dictyosomes, and numerous cytoplasmic vesicles. Type II cells produce spherical granules, contain fewer dictyosomes, have less conspicuous cytoplasmic vesiculation and possess more prominent parallel arrays of rough endoplasmic reticulum. Granules similar to those found in both cell types are present in the neuropile and certain nerves, but the specific pathways of the axons of these cells have not yet been determined.     

Reversible inhibition of secretion in root cap cells of cress after treatment with cytochalasin B: Support for the membrane flow concept

After treatment of cress roots with cytochalasin B (cytB) (25 μg/ml. 5.2 × 10^?5 M) for 4 h, marked structural changes are observed in the peripheral secretory calyptra cells. Deposits of slime outside the plasma membrane are smaller than in cells of untreated roots, whereas secretory vesicles accumulate within the treated cells. Dictyosomes are no longer present and the number of cisternae of rough endoplasmic reticulum surrounding the nucleus is increased at least three-fold. After an 8 h leaching of the drug, the structure of the secretory cells changes again. Accumulation of secretory vesicles no longer takes place, slime is deposited outside the plasma membrane and the number of ER cisternae surrounding the nucleus decreases. On the other hand, dictyosomes are now present. However, they are different from those in the hypertrophied stage of cells exhibiting high secretory activity, but are similar to those of an early developmental stage found at the beginning of the secretion process. This indicates that the dictyosomes are rebuilt during the leaching procedure. The results show that ER membranes accumulate near the nuclear envelope. They also indicate that bulk membrane material is transferred from the RER to the plasma membrane via dictyosome membranes and secretory vesicles, i.e. that membrane flow occurs in secretory cells of higher plants.     

Membranfluß und Nucleosiddiphosphatase-Reaktion in Wurzelhaaren von Lepidium sativum

Summary Root hairs of Lepidium sativum were incubated with a Wachstein-Meisel medium in experiments designed to localize the activity of nucleoside diphosphatase(s). Electron dense precipitates were found in the ER and in Golgi cisternae of the secretory face of the dictyosomes and their adjacent Golgi vesicles. Such precipitates were absent in the Golgi cisternae of the regeneration face of the dictyosomes and in the detached Golgi vesicles which extrude pectic cell wall substances. These results may be the consequence of the normal cycle of membrane compounds associated with the secretion in which the nucleoside diphosphatase(s) participate (by activation and inactivation) as one of the cycling components. Alternatively the nucleoside diphosphatase(s) may undergo a special cycle in which they are transferred from one cisterna or its vesicles to the next as part of the process of cisternal maturation.     

A COMPARISON OF ROOT CAP CELLS OF EPIPHYTIC,TERRESTRIAL AND AQUATIC PLANTS

Outer cells from the root cap of Cattleya orchids are characterized by their secretory activity. They are arranged in layers intercalated with layers of secretory product and form a protective mantle over the root tip. The ultrastructure of these cells is similar to those of terrestrial roots (for example Zea mays) in that they are characterized by copious quantities of endoplasmic reticulum and numerous dense-staining prevacuolar bodies. In contrast, most root cap cells of water hyacinth and duckweed are highly vacuolate with no dense-staining prevacuolar bodies. The endoplasmic reticulum is sparse and dictyosomes are small and without secretory activity.     

Ultrastructure and a possible function of the intercisternal elements in dictyosomes

The slime-producing dictyosomes in the placentary papillae of Aptenia cordifolia (L.f.) Schwant. show some structural peculiarities: (1) the number of their cisternae is conspicuously large in comparison with those of other cormophyta; (2) the spaces between the extremely flat vesicle-producing cisternae of the maturing face are considerably higher than those between the other cisternae; (3) the intercisternal elements show a pearl-string form rather than a fibrillar form-especially on tangential sections. Based on personal and on cited findings, the following hypothesis is developed: The intercisternal elements effect the compression of the central region of the secretory cisternae. This causes the production of vesicles to remain restricted to the marginal region of the cisternae, even if these cisternae contain hypertonic or soaking substances.     

Ultrastructure and Secretion of the Secretory Cells of Two Species ofFagoniaL. (Zygophyllaceae)

The ultrastructure and secretion of the secretory cells of theglandular trichomes ofFagonia mollisandF. glutinosawere studied.The most important finding of this study is that two speciesof the same genus produce the lipophilic component of the secretorymaterial in completely different ways and at different siteswithin the cell. In the early stages of development of secretorycells ofF. mollis,numerous mitochondria, containing myelin-likestructures, occur in the basal part of the cell. Above them,highly-elongate elements, which are suspected to develop frommitochondria with myelin-like structures, are present. Thesehave been termed ‘modified mitochondria’. It issuggested that the myelin-like structures are precursors ofthe lipophilic material ofF. mollis.InF. glutinosa,the lipophilicmaterial appears first in the plastids as plastoglobuli. Polysaccharidesappear to be produced by dictyosomes in both species. Secretionof the secretory substance to the outside of the protoplastappears to be granulocrine.Copyright 1998 Annals of Botany Company Fagonia mollis,Fagonia glutinosa,glandular trichomes, secretory cell, mitochondria, modified mitochondria, plastids, dictyosomes, lipophilic material, myelin-like structures, polysaccharides     

Numerical and structural changes in dictyosomes during zygospore germination ofClosterium ehrenbergii

Summary Numerical and structural changes in dictyosomes during the germination of zygospores inClosterium ehrenbergii were examined by electron microscopy. In the dormant mature zygospores, two parallel cisternac were seen which were derived from the disorganization of dictyosomes during the maturation of zygospores. After the induction of germination, the two parallel cisternae developed into dictyosomes with ten or eleven cisternae. The dictyosomes doubled in number by division every day for four days and reached, at the time of germination, a density of distribution similar to that found in the youngest zygospore. On the 4th day after the induction of germination, dictyosomes produced two kinds of vesicles which appear to be involved in the formation of new cell wall layers. The germination of the zygospore was effected by the escape of the cell covered with the new cell wall layers through the broken old cell wall layers.     

AN ULTRASTRUCTURAL BASIS FOR HYPHAL TIP GROWTH IN PYTHIUM ULTIMUM

Hyphae of the fungus Pythium ultimum extend by tip growth. The use of surface markers demonstrates that cell expansion is limited to the curved portion of the hyphal apex. Growing and non-growing regions are reflected in internal organization as detected by light and electron microscopy. The young hypha consists of three regions: an apical zone, a subapical zone and a zone of vacuolation. The apical zone is characterized by an accumulation of cytoplasmic vesicles, often to the exclusion of other organelles and ribosomes. Vesicle membranes are occasionally continuous with plasma membrane. The subapical zone is non-vacuolate and rich in a variety of protoplasmic components. Dictyosomes are positioned adjacent to endoplasmic reticulum or nuclear envelope, and vesicles occur at the peripheries of dictyosomes. A pattern of secretory vesicle formation by dictyosomes is described which accounts for the formation of hyphal tip vesicles. Farther from the hyphal apex the subapical zone merges into the zone of vacuolation. As hyphae age vacuolation increases, lipid accumulations appear, and the proportional volume of cytoplasm is reduced accordingly. The findings are integrated into a general hypothesis to explain the genesis and participation of cell components involved directly in hyphal tip growth: Membrane material from the endoplasmic reticulum is transferred to dictyosome cisternae by blebbing; cisternal membranes are transformed from ER-like to plasma membrane-like during cisternal maturation; secretory vesicles released from dictyosomes migrate to the hyphal apex, fuse with the plasma membrane, and liberate their contents into the wall region. This allows a plasma membrane increase at the hyphal apex equal to the membrane surface of the incorporated vesicles as well as a contribution of the vesicle contents to surface expansion.     

The role of calcium in exocytosis and endocytosis in plant cells

The role of calcium in the individual cellular events leading to exocytosis is considered. Both vesicle movement processes and vesicle fusion at the cell surface require calcium for completion of specific events in this pathway. Our knowledge of these events is incomplete. In particular the movement of secretory vesicles by the cytoskeleton in response to added calcium is a key event that is beyond our comprehension at present. At the whole cell level, it is shown that external calcium, at the appropriate concentration, is required to elicit secretion at optimal rates. In both plant and animal cells secretion appears to be dependent on, or is triggered by, a rise in the level of internal free calcium ions from about 10^-7 to 10^-6M or even higher. In these eukaryotes internal organelles take up calcium and maintain a low level of calcium in the cell, offsetting the inflow of calcium from the plasma membrane. In some systems the inflow is restricted to a certain part of the plasma membrane, which then acts as a focus for exocytosis and, thereby, establishes a cellular polarity. In plant tissues there appears to be a requirement for some circulation of calcium within the apoplast, to sustain secretion. Recent papers on endocytosis have confirmed its occurrence in plant cells and made significant advances in isolating and characterising the clathrin coats of the coated vesicles involved in the uptake. There is no evidence, at present, for a direct role for calcium in these events. Indirectly, calcium stimulates exocytosis, and hence the delivery of excess membrane to the cell surface, which may be retrieved by an increase in the rate of endocytosis. Quantitative comparisons of the membrane flow occurring in these pathways are not available. Several plant cellular systems have been employed to study secretion and some of these may prove to be superior model systems for the investigation of certain aspects of the control of exocytosis and endocytosis by calcium ions.     

The electron microscopic and classic Golgi apparatus

The relationship of the membrane structure, designated in electron microscopy as the Golgi apparatus, to the classic Golgi apparatus in the light microscope were studied withFagopyrum. Comparison of these structures in plant cells with the same or similar structures in animal cells led to the following conclusions: there exist two groups of formations, impregnable with osmium or silver, considered as the classic Golgi apparatus. The first group contains the active membrane structures. These are the dictyosomes and the anastomosing form of the electron microscopic Golgi apparatus. To this group belongs also the endoplasmatic reticulum, which in plant cells forms dense vacuoles, having the appearance of the classic Golgi apparatus, and in animal cells occasionally has a similar arrangement as the anastomosing form of the Golgi apparatus. The second group comprises formation containing reserve and secretion material, i.e. predominantly products of the activity of the electron microscopic Golgi apparatus and of the endoplasmic reticulum (matter of the dense vacuoles, lipochondria, secretory granula etc.). In the plant cells, especially ofFygopyrum, the dictyosomes contained in the structures of the first group are separated from the formations of a reserve character in the second group, formed in the lumen of the endoplasmic reticulum (dense vacuoles). The identity of the dictyosomes with the osmiophilic platelets, considered by some authors in the light microscope as the classic Golgi apparatus, has not been proved up to present, because of the one-sidedness of the methods used nowadays. WithFagopyrum no foundation has been observed for the assumed formation of net-form structures by grouping of the dictyosomes. Structures similar to the net-form of the classic Golgi apparatus in the animal cell form only dense vacuoles. On the basis of the differentiation of both types of formations in the plant cell, the foundations were laid for the characterization of the classic Golgi apparatus in the animal cell. The net-form of the classic Golgi apparatus in the animal cell is obviously not artificial, but reflects the ultrastructural arrangement of the electron microscopic Golgi apparatus or of the endoplasmic reticulum. The problem of the suitability and specification of the name Golgi apparatus in the animal and plant cell was also discussed. In contrast to the opinion of some authors, it does not appear useful to remove the name golgi apparatus, designating the dictyosomes and the anastomosing forms of the smooth membranes.     

Development and ultrastructure of glandular trichomes in<Emphasis Type="Italic">pelargonium x fragrans</Emphasis> ‘mabel grey’ (geraniaceae)

The glandular trichomes of leaves fromPelargonium xfragrans ‘Mabel Grey’ (Geraniaceae) were examined by light, scanning, and transmission electron microscopy. These trichomes had unicellular globular heads and stalks of different lengths and features. Two types were classified: Type I, with an elongated, large head and a short (100 μm), cylindrical stalk that was more apparent on the adaxial surface; and Type II, with a spherical, small head and a long (300μm), conical stalk that was more pronounced on the abaxial surface. The ultrastructure of secretory cells from both types was distinguished by a well-developed endoplasmic reticulum, mitochondria, plastids, dictyosomes, and numerous vacuoles that likely were involved in the storage and transport of lipophilic substances. Plasmodesmata were frequent on the walls of the secretory and stalked cells. Here, we discuss the implication of structural differentiation in these trichomes.