Distortion of plant cell plate formation by the intracellular-calcium antagonist TMB-8

Summary Caulonema tip cells ofFunaria deposit new oblique cross walls of specific morphology and placement by a highly defined reorientation mechanism. In the presence of the purported intracellular Ca²⁺ antagonist 8-(N,N-diethylamino)octyl-3,4,5-trimethoxybenzoate (TMB-8), these cross walls form in the proper place but exhibit a distorted morphology. Video microscopy indicates that the deformation takes place during the reorientation of the cell plate from a perpendicular to an oblique configuration. Electron micrographs of TMB-8 treated cells indicate a stabilization of phragmoplast microtubules and a greater amount of vesicles and membrane in the developing cell plate. TMB-8 treated cells also show intense chlortetracycline fluorescence from mitochondria, vesicles and endoplasmic reticulum as compared to untreated cells indicating that TMB-8 is blocking release of Ca²⁺ from intracellular stores. It is concluded that this may cause distortation of cross walls as they form by delaying vesicle fusion, stabilizing microtubules, and increasing the amount of new wall material in the developing cell plate.Abbreviations CTC chlortetracycline - OsFeCN osmium ferricyanide method - TMB-8 8-(N,N-diethylamino)octyl-3,4,5-trimethoxybenzoate     

The structure of the endoplasmic reticulum revealed by osmium tetroxide-potassium ferricyanide staining

Postfixation of plant tissues with a mixture of osmium tetroxide and potassium ferricyanide (OsFeCN) yields a selective staining of the endoplasmic reticulum (ER) and nuclear envelope (NE). The other cytoplasmic organelles and inclusions are evident, but by comparison with the NE-ER they are weakly contrasted. Demarcation of the NE-ER results from the enhanced deposition of an electron-opaque reaction product on the inner leaflet of the membrane that extends into the cisternal space. The procedure thus renders the NE-ER readily apparent even when the elements are sectioned parallel to their surface and makes it possible to easily visualize their cellular pattern. Ultrastructural studies reveal with clarity tubular reticula and fenestrated lamellae that are extensively interconnected into one continuous membrane system. Problems with the OsFeCN procedure include the inability of the reagent to stain the NE-ER in all cells of a tissue, the occasional staining of non-ER such as dictyosomal cisternae and plastids, and the failure to selectively stain the NE-ER in protoplasts or single wall-less cells. Results obtained with OsFeCN are compared with other ER fixatives and stains including potassium permanganate and zinc iodide-osmium tetroxide. Despite its problems, under optimal circumstances OsFeCN is judged to be superior to other stain-fixatives for selectively contrasting the NE-ER compartment and is recommended generally for ultrastructural investigations.     

Studies on graft unions

Summary De novo formation of cytoplasmic cell connections are studied at the graft interface of 5 day old in vitro heterografts ofVicia faba onHelianthus annuus. Continuous and half plasmodesmata, both branched and unbranched, are described at various stages of development in non-division walls between unlike and like dedifferentiated callus cells. In apical portions of protruding callus cells and in the contact zone between opposing cells extremely thin wall parts with a striking ER/plasmalemma contact are observed. During subsequent thickening of the modified wall parts cytoplasmic strands enclosing constricted ER cisternae are entrapped within the newly deposited wall material. These cytoplasmic strands represent half plasmodesmata which—in case of fusion with corresponding structures of adjoining cells across the loosened wall matrix — form continuous cell connections. Golgi vesicles secreting wall material are involved in the process of forming half and continuous plasmodesmata, thus following the same mechanism of plasmodesmata development as described for isolated protoplasts in cell cultures. The findings suggest the existence of a unifying mechanism of secondary formation of plasmodesmata showing far-reaching similarities with the establishment of primary cell connections.     

Ultrastructural studies of a vesicle system associated with endoplasmic reticulum in exo-erythrocytic forms of Plasmodium berghei

Fine structural studies of a specialized vesicle system associated with the endoplasmic reticulum (ER) of exo-erythrocytic Plasmodium berghei suggest that this system may be the equivalent of a Golgi apparatus. Patches of ER, randomly distributed in the cytoplasm of developing parasites, are formed of smooth and ribosome-studded cisternae intermingled with each other. The vesicle systems are located between as well as at the edges of ER aggregates and appear to be in different stages of budding from the cisternae. Prolonged osmication reveals distinct staining of the nuclear envelope and ER of the parasites as well as part of the Golgi apparatus of the hepatocytes. However, the small vesicles associated with the parasite's ER are unstained, as are the coated vesicles in the Golgi region of the liver cell. These sites in the parasite cytoplasm seem comparable to the concave surface of the Golgi apparatus in liver cells. The pinched-off vesicles fuse with others to form the prominent peripheral vacuolization characteristic of the nearly mature exoerythrocytic form. The formation of these peripheral vacuoles and their subsequent fusion with the parasite membrane may be an exocytosis mechanism supplying the rapidly expanding parasite with new plasma membrane material.     

Membranes in the miotic apparatus of barley cells

Membranes in the mitotic apparatus have been investigated ultrastructually in dividing cells of barley (Hordeum vulgare). After osmium tetroxide- potassium ferricyanide or ferrocyanide postfixation (OsFeCN) of material that had been fixed in glutaraldehyde in the presence of Ca(++), the nuclear envolope (NE)-endoplasmic reticulum (ER) complex is selectively stained, permitting observations on the cellular pattern and structural ramifications of this membrane system that have not been previously recognized. Specifically, it is observed that membrane system that have not been previously recognized. Specifically, it is observed that during mitosis the NE-ER forms a continuous membrane system that ensheathes and isolates the mitotic apparatus (MA). Elements of ER progressively accumulate in the region of the spindle pole, becoming most concentrated by early anaphase. Within the MA itself, there are striking spindle- membrane associations; in particular, tubular elements of predominantly smooth NE-ER invade the spindle interior selectively along kinetochore microtubules. The membrane elements at the pole and surrounding the MA consist of tubular reticulum and fenestrated lamellae. Membranes of the MA thus resemble in considerable detail the tubular network and fenestrated elements of the sarcoplasmic reticulum of muscle. It is suggested that the NE-ER of the dividing barley cell may function in one or both of the following ways: (a) to control the concentration of free Ca(++) in the MA and (b) to serve as an anchor to chromosome motion.     

Electron tomographic analysis of somatic cell plate formation in meristematic cells of Arabidopsis preserved by high-pressure freezing

We have investigated the process of somatic-type cytokinesis in Arabidopsis (Arabidopsis thaliana) meristem cells with a three-dimensional resolution of approximately 7 nm by electron tomography of high-pressure frozen/freeze-substituted samples. Our data demonstrate that this process can be divided into four phases: phragmoplast initials, solid phragmoplast, transitional phragmoplast, and ring-shaped phragmoplast. Phragmoplast initials arise from clusters of polar microtubules (MTs) during late anaphase. At their equatorial planes, cell plate assembly sites are formed, consisting of a filamentous ribosome-excluding cell plate assembly matrix (CPAM) and Golgi-derived vesicles. The CPAM, which is found only around growing cell plate regions, is suggested to be responsible for regulating cell plate growth. Virtually all phragmoplast MTs terminate inside the CPAM. This association directs vesicles to the CPAM and thereby to the growing cell plate. Cell plate formation within the CPAM appears to be initiated by the tethering of vesicles by exocyst-like complexes. After vesicle fusion, hourglass-shaped vesicle intermediates are stretched to dumbbells by a mechanism that appears to involve the expansion of dynamin-like springs. This stretching process reduces vesicle volume by approximately 50%. At the same time, the lateral expansion of the phragmoplast initials and their CPAMs gives rise to the solid phragmoplast. Later arriving vesicles begin to fuse to the bulbous ends of the dumbbells, giving rise to the tubulo-vesicular membrane network (TVN). During the transitional phragmoplast stage, the CPAM and MTs disassemble and then reform in a peripheral ring phragmoplast configuration. This creates the centrifugally expanding peripheral cell plate growth zone, which leads to cell plate fusion with the cell wall. Simultaneously, the central TVN begins to mature into a tubular network, and ultimately into a planar fenestrated sheet (PFS), through the removal of membrane via clathrin-coated vesicles and by callose synthesis. Small secondary CPAMs with attached MTs arise de novo over remaining large fenestrae to focus local growth to these regions. When all of the fenestrae are closed, the new cell wall is complete. Few endoplasmic reticulum (ER) membranes are seen associated with the phragmoplast initials and with the TVN cell plate that is formed within the solid phragmoplast. ER progressively accumulates thereafter, reaching a maximum during the late PFS stage, when most cell plate growth is completed.     

The herbicide dichlobenil disrupts cell plate formation: immunogold characterization

Summary We have utilized light and transmission electron microscopy and immunocytochemistry to examine onion roots treated with the herbicide dichlobenil (2,6-dichlorobenzonitrile; DCB), a purported disrupter of cellulose biosynthesis. The most salient effect of DCB is observed on cell plate formation, the process that gives rise to new cell walls. In the presence of DCB, cell plates develop normally up to the tubular network stage. They are the result of fusion of Golgi-derived vesicles and the accumulation of callose and the first strands of cellulose. The DCB-treated cell plates retain the reticulate and malleable nature of the tubular network/early fenestrated plate stage of cell plate formation, but fail to display signs of the stiffening and straightening associated with an accumulation of cellulose. Instead, the malleable cell plates in the DCB-treated cells retain a wavy architecture, accumulate pockets of electron opaque material, and produce plasmodesmata in abnormal orientations. Immunocytochemical investigations of the abnormal cell plates formed after DCB treatment show 20-fold increase in the level of callose labelling found in the control cell plates. Xyloglucans and rhamnogalacturonans can be detected in the partially-formed cell plates, with the labelling density of xyloglucan 4–5 times greater than in the control cell plates and that of the rhamnogalacturonans being similar to the controls. These data support the hypothesis that DCB inhibits cellulose biosynthesis as a primary mechanism of action, and that in the absence of cellulose synthesis the cell plates fail to mature and to give rise to new cross walls.Abbreviations DCB dichlorobenzonitrile - PGA/RGI polygalacturonic acid/rhamnogalacturonan I     

Ultrastructural Studies of a Vesicle System Associated with Endoplasmic Reticulum in Exo-Erythrocytic Forms of Plasmodium berghei1

ABSTRACT. Fine structural studies of a specialized vesicle system associated with the endoplasmic reticulum (ER) of exo-erythrocytic Plasmodium berghei suggest that this system may be the equivalent of a Golgi apparatus. Patches of ER, randomly distributed in the cytoplasm of developing parasites, are formed of smooth and ribosome-studded cisternae intermingled with each other. The vesicle systems are located between as well as at the edges of ER aggregates and appear to be in different stages of budding from the cisternae. Prolonged osmication reveals distinct staining of the nuclear envelope and ER of the parasites as well as part of the Golgi apparatus of the hepatocytes. However, the small vesicles associated with the parasite's ER are unstained, as are the coated vesicles in the Golgi region of the liver cell. These sites in the parasite cytoplasm seem comparable to the concave surface of the Golgi apparatus in liver cells. The pinched-off vesicles fuse with others to form the prominent peripheral vacuolization characteristic of the nearly mature exo-erythrocytic form. The formation of these peripheral vacuoles and their subsequent fusion with the parasite membrane may be an exocytosis mechanism supplying the rapidly expanding parasite with new plasma membrane material.     

Caffeine inhibition of cytokinesis: Ultrastructure of cell plate formation/degradation

Summary Caffeine is a potent inhibitor of cell plate formation in dividing plant cells. Previous studies living cells reveal that the drug always permits the cell plate to arise and grow normally until about 80% complete, but then causes it to break down. In the present investigation we examine this formation/degradation cycle at the ultrastructure level. Our results show that during the formation phase the caffeine treated plate is indistinguishable from untreated controls. Phragmoplast microtubules arise and align in the interzone, Golgi vesicles are produced and aggregate in a line that defines the young cell plate, and considerable fusion of these vesicles occurs to form islands of plate material. However, under the influence of caffeine these islands do not fuse to form the enlarged lamellar expanses characteristic of maturing cell plates. Instead, the partially fused material reverts to small vesicles which appear to become resorbed by the cellular membrane systems. The resorption process continues leaving no evidence of the previously developing plate, although occasionally we observe a stub of fused vesicles attached to the parent wall. Following cell plate disintegration the reformed nuclei move close together and occupy the central region of the cell. These observations focus attention on the consolidation phase of cell plate formation as the one being maximally affected by caffeine.Dedicated to the memory of Professor Oswald Kiermayer     

Caffeine inhibits cell plate formation by disrupting membrane reorganization just after the vesicle fusion step

Summary We have re-examined the effects of caffeine on cell plate formation in synchronized tobacco BY-2 cells by means of cryofixation, immunocytochemistry, and calcium staining techniques. Because cryofixation preserves structural intermediates of cell plates that are not seen in chemically fixed cells, this methodology has enabled us to define not only when caffeine acts but also which assembly steps are inhibited. Caffeine acts at an early stage of cytokinesis, just after the Golgi-derived vesicles have arrived at the cell equator and begun to fuse with each other via thin (20 nm) membrane tubules. This initial round of fusions produces a delicate membrane network which in control cells is rapidly converted in a more substantial tubulo-vesicular network covered by a thick, fuzzy coat on its cytoplasmic surface. Caffeine disrupts the conversion of the fragile, thin, fusion tube-generated membrane network into the more stable tubulo-vesicular network, the assembly of its fuzzy coat, and the budding of clathrin-coated vesicles from its surface. Normally, the tubulo-vesicular network also provides the structural framework for calcium-dependent callose synthases that deposit a callose layer over the lumenal surface of the cell plate membranes. In the presence of caffeine, no stabilizing callose layer is formed, and the thin tubule membrane network fragments into vesicles of variable sizes. Cell plates in caffeine-treated cells stained with chlortetracycline, a fluorescent stain of membrane-associated calcium, also display a significant reduction in fluorescence at the cell plate, suggesting a major decrease in cell plate membrane-associated calcium. However, this latter finding needs to be confirmed by more sophisticated calcium measuring techniques. Current theories of the mechanism of action of caffeine, including its ability to disrupt local calcium gradients, are discussed within the new ultrastructural context that this study provides. Our findings, finally, suggest a new method for isolating just fused but not further matured cell plate forming vesicles for biochemical studies.Dedicated to Professor Eldon H. Newcomb in recognition of his contributions to cell biology     

Cytokinesis in tobacco BY-2 and root tip cells: a new model of cell plate formation in higher plants

Cell plate formation in tobacco root tips and synchronized dividing suspension cultured tobacco BY-2 cells was examined using cryofixation and immunocytochemical methods. Due to the much improved preservation of the cells, many new structural intermediates have been resolved, which has led to a new model of cell plate formation in higher plants. Our electron micrographs demonstrate that cell plate formation consists of the following stages: (1) the arrival of Golgi-derived vesicles in the equatorial plane, (2) the formation of thin (20 +/- 6 nm) tubes that grow out of individual vesicles and fuse with others giving rise to a continuous, interwoven, tubulo-vesicular network, (3) the consolidation of the tubulo-vesicular network into an interwoven smooth tubular network rich in callose and then into a fenestrated plate-like structure, (4) the formation of hundreds of finger-like projections at the margins of the cell plate that fuse with the parent cell membrane, and (5) cell plate maturation that includes closing of the plate fenestrae and cellulose synthesis. Although this is a temporal chain of events, a developing cell plate may be simultaneously involved in all of these stages because cell plate formation starts in the cell center and then progresses centrifugally towards the cell periphery. The "leading edge" of the expanding cell plate is associated with the phragmoplast microtubule domain that becomes concentrically displaced during this process. Thus, cell plate formation can be summarized into two phases: first the formation of a membrane network in association with the phragmoplast microtubule domain; second, cell wall assembly within this network after displacement of the microtubules. The phragmoplast microtubules end in a filamentous matrix that encompasses the delicate tubulo-vesicular networks but not the tubular networks and fenestrated plates. Clathrin-coated buds/vesicles and multivesicular bodies are also typical features of the network stages of cell plate formation, suggesting that excess membrane material may be recycled in a selective manner. Immunolabeling data indicate that callose is the predominant lumenal component of forming cell plates and that it forms a coat-like structure on the membrane surface. We postulate that callose both helps to mechanically stabilize the early delicate membrane networks of forming cell plates, and to create a spreading force that widens the tubules and converts them into plate-like structures. Cellulose is first detected in the late smooth tubular network stage and its appearance seems to coincide with the flattening and stiffening of the cell plate.     

Three-dimensional analysis of syncytial-type cell plates during endosperm cellularization visualized by high resolution electron tomography

The three-dimensional architecture of syncytial-type cell plates in the endosperm of Arabidopsis has been analyzed at approximately 6-nm resolution by means of dual-axis high-voltage electron tomography of high-pressure frozen/freeze-substituted samples. Mini-phragmoplasts consisting of microtubule clusters assemble between sister and nonsister nuclei. Most Golgi-derived vesicles appear connected to these microtubules by two molecules that resemble kinesin-like motor proteins. These vesicles fuse with each other to form hourglass-shaped intermediates, which become wide (approximately 45 nm in diameter) tubules, the building blocks of wide tubular networks. New mini-phragmoplasts also are generated de novo around the margins of expanding wide tubular networks, giving rise to new foci of cell plate growth, which later become integrated into the main cell plate. Spiral-shaped rings of the dynamin-like protein ADL1A constrict but do not fission the wide tubules at irregular intervals. These rings appear to maintain the tubular geometry of the network. The wide tubular network matures into a convoluted fenestrated sheet in a process that involves increases of 45 and 130% in relative membrane surface area and volume, respectively. The proportionally larger increase in volume appears to reflect callose synthesis. Upon fusion with the parental plasma membrane, the convoluted fenestrated sheet is transformed into a planar fenestrated sheet. This transformation involves clathrin-coated vesicles that reduce the relative membrane surface area and volume by approximately 70%. A ribosome-excluding matrix encompasses the cell plate membranes from the fusion of the first vesicles until the onset of the planar fenestrated sheet formation. We postulate that this matrix contains the molecules that mediate cell plate assembly.     

Low and high voltage electron microscopy of mitosis and cytokinesis in maize roots

The structure and distribution of cytoplasmic membranes during mitosis and cytokinesis in maize root tip meristematic cells was investigated by low and high voltage electron microscopy. The electron opacity of the nuclear envelope and endoplasmic reticulum (ER) was enhanced by staining the tissue in a mixture of zinc iodide and osmium tetroxide. Thin sections show the nuclear envelope to disassemble at prophase and become indistinguishable from the surrounding ER and polar aggregations of ER. In thick sections under the high voltage electron microscope the spindle is seen to be surrounded by a mass of tubular (TER) and cisternal (CER) endoplasmic reticulum derived from both the nuclear envelope and ER, which persists through metaphase and anaphase. At anaphase strands of TER traverse the spindle between the arms of the chromosomes. The octagonal nuclear pore complexes disappear by metaphase, but irregular-shaped pores persist in the membranes during mitosis. It is suggested that these form a template for pore-complex reformation during telophase. Phragmoplast formation is preceded by an aggregation of TER across the spindle at anaphase. Evidence is presented to suggest that the formation of the desmotubule of a plasmodesma is by the squeezing of a strand of endoplasmic reticulum between the vesicles of the cell plate.Abbreviations CER cisternal endoplasmic reticulum - ER endoplasmic reticulum - HVEM high voltage electron microscope - TER tubular endoplasmic reticulum - ZIO zinc iodide/osmium tetroxide     

杨树顶芽细胞内质网与其他膜系统的结构联系及其在休眠过程中的变化

杨树（Ｐｏｐｕｌｕｓ ｄｅｌｔｏｉｄｅｓ Ｂａｒｔｒ．ｅｘ Ｍａｒｓｈ）顶芽分生组织细胞经一种改良的高锰酸钾固定法固定后,显示出一种十分清晰的内膜结构,尤其展现了内质网与其他膜系统存在一种结构上的密切联系。一些与核膜相连接的内质网伸展到细胞质中与线粒体、质体及高尔基体发生联系,可延伸到质膜。还有些内质网的一端与一个细胞的核膜相连结,其另一端穿过胞间连丝与邻近的另一个细胞的核膜相连结,在两个相邻的细     

Electron tomographic analysis of post-meiotic cytokinesis during pollen development in<Emphasis Type="Italic"> Arabidopsis thaliana</Emphasis>

The mechanism of cell wall formation after male meiosis was studied in microsporocytes of Arabidopsis thaliana (L.) Heynh. by means of thin-section and immuno-electron microscopy and dual-axis electron tomography of high-pressure-frozen/freeze-substituted cells. The cellularization of four-nucleate microsporocytes involves a novel type of cell plate, called a post-meiotic-type cell plate. As in the syncytial endosperm, the microsporocyte cell plates assemble in association with mini-phragmoplasts. However, in contrast to the endosperm cell plates, post-meiotic type cell plates arise simultaneously across the entire division plane. Vesicles are transported along mini-phragmoplast microtubules by putative kinesin proteins and, prior to fusion, they become connected together by 24-nm-long linkers that resemble exocyst complexes. These vesicles fuse with each other to form wide tubules and wide tubular networks. In contrast to endosperm cell plates, the wide tubular networks in microsporocytes completely lack callose and do not appear to be constricted by dynamin rings. The most peripheral wide tubular networks begin to fuse with the plasma membrane before the more central cell plate assembly sites become integrated into a coherent cell plate. Fusion with the parental plasma membrane triggers callose synthesis and the wide tubular domains are converted into convoluted sheets. As the peripheral convoluted sheets accumulate callose and arabinogalactan proteins, they are converted into stub-like projections, which grow centripetally, i.e. toward the interior of the syncytium, fusing with the wide tubular networks already assembled in the division plane. We also demonstrate that the ribosome-excluding cell plate assembly matrix is delivered to the mini-phragmoplast with the first vesicles, and encompasses all the linked vesicles and intermediate stages in cell plate formation.Abbreviations AGP Arabinogalactan protein - MT Microtubule     

Nuclear envelope formation by chromatin-mediated reorganization of the endoplasmic reticulum

The formation of the nuclear envelope (NE) around chromatin is a major membrane-remodelling event that occurs during cell division of metazoa. It is unclear whether the nuclear membrane reforms by the fusion of NE fragments or if it re-emerges from an intact tubular network of the endoplasmic reticulum (ER). Here, we show that NE formation and expansion requires a tubular ER network and occurs efficiently in the presence of the membrane fusion inhibitor GTPgammaS. Chromatin recruitment of membranes, which is initiated by tubule-end binding, followed by the formation, expansion and sealing of flat membrane sheets, is mediated by DNA-binding proteins residing in the ER. Thus, chromatin plays an active role in reshaping of the ER during NE formation.     

Syncytial-type cell plates: a novel kind of cell plate involved in endosperm cellularization of Arabidopsis

Cell wall formation in the syncytial endosperm of Arabidopsis was studied by using high-pressure-frozen/freeze-substituted developing seeds and immunocytochemical techniques. The endosperm cellularization process begins at the late globular embryo stage with the synchronous organization of small clusters of oppositely oriented microtubules ( approximately 10 microtubules in each set) into phragmoplast-like structures termed mini-phragmoplasts between both sister and nonsister nuclei. These mini-phragmoplasts produce a novel kind of cell plate, the syncytial-type cell plate, from Golgi-derived vesicles approximately 63 nm in diameter, which fuse by way of hourglass-shaped intermediates into wide ( approximately 45 nm in diameter) tubules. These wide tubules quickly become coated and surrounded by a ribosome-excluding matrix; as they grow, they branch and fuse with each other to form wide tubular networks. The mini-phragmoplasts formed between a given pair of nuclei produce aligned tubular networks that grow centrifugally until they merge into a coherent wide tubular network with the mini-phragmoplasts positioned along the network margins. The individual wide tubular networks expand laterally until they meet and eventually fuse with each other at the sites of the future cell corners. Transformation of the wide tubular networks into noncoated, thin ( approximately 27 nm in diameter) tubular networks begins at multiple sites and coincides with the appearance of clathrin-coated budding structures. After fusion with the syncytial cell wall, the thin tubular networks are converted into fenestrated sheets and cell walls. Immunolabeling experiments show that the cell plates and cell walls of the endosperm differ from those of the embryo and maternal tissue in two features: their xyloglucans lack terminal fucose residues on the side chain, and callose persists in the cell walls after the cell plates fuse with the parental plasma membrane. The lack of terminal fucose residues on xyloglucans suggests that these cell wall matrix molecules serve both structural and storage functions.     

Simultaneous visualization of the actin plate and new cell partition membrane during cytokinesis in the brown alga Sphacelaria rigidula (Sphacelariales,Phaeophyceae)

In many brown algae, cytokinesis is accomplished through the centrifugal expansion of the membrane structure formed by the fusion of Golgi vesicles and flat cisternae. In contrast, it has been reported that cytokinesis in Sphacelaria rigidula progresses centripetally by adding Golgi vesicles and flat cisternae to cleaving furrows of the plasma membrane. The reason why this cytokinetic pattern was observed only in Sphacelaria species is unknown. In either cytokinesis pattern, a plate-like actin structure (the actin plate) coincides with the cytokinetic plane between the daughter nuclei. However, it is unclear how the actin plate is related to cytokinesis progression. In this study, we re-examined cytokinesis in the apical cells of S. rigidula using transmission electron microscopy. Double staining of the actin plate and the developing membrane was followed by fluorescence microscopy analysis to determine the relationship between these two formations. The results showed that cytokinesis in S. rigidula, as in many brown algae, was completed by centrifugal growth of the new cell partition membrane. A furrow of the plasma membrane was observed at the beginning of cytokinesis; however, further invagination did not occur. The actin plate arose at the center of the cytokinetic plane before membrane fusion and extended parallel to the expansion of the new cell partition membrane. When cytokinesis was slow due to insufficient Golgi vesicle supply to the cytokinetic plane in the cells under brefeldin A treatment, the extension of the actin plate was also suspended. In this study, the spatiotemporal relationship between the occurrence and expansion of the actin plate and the new cell partition membrane was revealed. These observations indicate that the actin plate might promote membrane fusion or lead to the growth of a new cell partition membrane.     

Cytochemical localization of cellulase activity in pollen mother cells of david lily during meiotic prophase I and its relation to secondary formation of plasmodesmata

Summary Cellulase activity was localized at the ultrastructural level in pollen mother cells (PMCs) of David lily [Lilium davidii var.willmottiae (Wilson) Roffill] at different stages of meiotic prophase I. The enzyme was observed to appear at the early leptotene stage and reached its highest level at the subsequent zygotene stage, and its subcellular distribution revealed by the presence of electron-dense deposits of reaction product was found to be restricted exclusively to the endoplasmic reticulum (ER), the vesicles derived from that, and the cell wall, especially at the sites of secondary plasmodesmata and cytoplasmic channels where the wall was being digested. Other cytoplasmic organelles, such as dictyosomes and Golgi vesicles, lacked such deposits of reaction product. After zygotene the enzyme activity decreased abruptly, and at the pachytene stage only very few deposits could be observed in the cell wall. Our results indicate that cellulase is synthesized on rough ER and secreted directly via the smooth ER and ER-derived vesicles into the cell wall by exocytosis, where it brings about local wall breakdown, leading to the secondary formation of plasmodesmata and cytoplasmic channels.     

Ca2+-homeostasis Differs Between Plant Species with Different Cold-tolerance at 4℃ Chilling

Electron microscopic observations revealed that the tissues of poplar (Populus deltoides Bartr. ex Marsh) apical bud cells, which were fixed by a modified procedure of potassium permanganate fixative, showed a distinct endomembrane organization, in particular, the structural associations of the endoplasmic reticulum (ER) with other membrane systems. The striking findings are that some ER elements were in connection with the nuclear envelopes of two adjacent cells through plasmodesmata, and many ER elements were also associated with mitochondria, plastids, Golgi bodies or the plasma membrane (PM), forming a bridge-like continuum among various endomembrane systems or between nucleus to nucleus. A great number of plasmodesmata existed between cells, indicating a perfectly integrated symplasmic structure in poplar apical bud meristem grown in a long day environment. During the short day-induced dormancy, ER contracted, leading to its disassociation between nuclei, and between the nucleus and organelles/plasmalemma in many cells. After dormancy broke and shoots growth resumed, contracted ER was no longer observed in the apical bud cells. The ER associations with other endomembrane systems and the intercellular communication channels were re-established similar to that of plants before dormancy induction. These observations suggest that ER may play an important role in linking-up between the nucleus and organelles, and between the nucleus and the nucleus (or cell-to-cell), and seemingly coordinating various physiological processes by the bridging-like associations. And the contraction of ER under short-day may result in the growth cessation and the development of dormancy in poplar.