Mitosis and cytokinesis inTribonema regulare (Tribophyceae,Chrysophyta)

Summary The ultrastructure of mitosis and cytokinesis of the uninucleateTribonema regulare has been investigated by employing transmission electron microscopy. Prophase is characterized by settlement of a pair of centrioles at the presumptive poles of the spindle, metaphase by equatorial bulging of the nucleus, anaphase by non-synchronous separation of the chromosomes, and telophase by a persistent, strongly elongated, interzonal spindle. Throughout mitosis, at each pole dictyosomes are associated with the polar gaps of the nuclear envelope that otherwise remains intact. Cytokinesis does not immediately follow mitosis; from the static images it can be concluded that it is necessary for the daughter nuclei to approach each other before cytokinesis is initiated by complete division of the protoplast via plasma membrane cleavage. Afterwards, a ring of cell wall material is deposited close near the lateral wall in the plane of protoplast separation followed by a simultaneous or centripetal development of a single integral partitioning septum. Once the septum is completed, the cylindrical portion of the H-shaped segment is manufactured. The phylogenetic position ofTribonema amongst those algae, which may have evolved from unicells into filaments, is discussed.     

Cytokinesis in brown algae: studies of asymmetric division in fucoid zygotes

Summary. The mechanism of cytokinesis was investigated during the first asymmetric division in fucoid zygotes. A plate of actin assembled midway between daughter nuclei where microtubules interdigitated and defined the cytokinetic plane. A membrane was then deposited in islands throughout the cytokinetic plane; the islands eventually fused into a continuous partition membrane and cell plate material was deposited in the intermembrane space. All of these structures matured from the center of the cell outward (centrifugal maturation). Pharmacological agents were used to investigate the roles of microtubules, actin, and secretion in cytokinesis. The findings indicate a mechanism of cytokinesis that may be unique to the brown algae.Correspondence and reprints: Department of Biology, University of Utah, 257 South 1400 East, Salt Lake City, UT 84112-0840, U.S.A.     

Rab11A-Controlled Assembly of the Inner Membrane Complex Is Required for Completion of Apicomplexan Cytokinesis

The final step during cell division is the separation of daughter cells, a process that requires the coordinated delivery and assembly of new membrane to the cleavage furrow. While most eukaryotic cells replicate by binary fission, replication of apicomplexan parasites involves the assembly of daughters (merozoites/tachyzoites) within the mother cell, using the so-called Inner Membrane Complex (IMC) as a scaffold. After de novo synthesis of the IMC and biogenesis or segregation of new organelles, daughters bud out of the mother cell to invade new host cells. Here, we demonstrate that the final step in parasite cell division involves delivery of new plasma membrane to the daughter cells, in a process requiring functional Rab11A. Importantly, Rab11A can be found in association with Myosin-Tail-Interacting-Protein (MTIP), also known as Myosin Light Chain 1 (MLC1), a member of a 4-protein motor complex called the glideosome that is known to be crucial for parasite invasion of host cells. Ablation of Rab11A function results in daughter parasites having an incompletely formed IMC that leads to a block at a late stage of cell division. A similar defect is observed upon inducible expression of a myosin A tail-only mutant. We propose a model where Rab11A-mediated vesicular traffic driven by an MTIP-Myosin motor is necessary for IMC maturation and to deliver new plasma membrane to daughter cells in order to complete cell division.     

Universal rules for division plane selection in plants

Coordinated cell divisions and cell expansion are the key processes that command growth in all organisms. The orientation of cell divisions and the direction of cell expansion are critical for normal development. Symmetric divisions contribute to proliferation and growth, while asymmetric divisions initiate pattern formation and differentiation. In plants these processes are of particular importance since their cells are encased in cellulosic walls that determine their shape and lock their position within tissues and organs. Several recent studies have analyzed the relationship between cell shape and patterns of symmetric cell division in diverse organisms and employed biophysical and mathematical considerations to develop computer simulations that have allowed accurate prediction of cell division patterns. From these studies, a picture emerges that diverse biological systems follow simple universal rules of geometry to select their division planes and that the microtubule cytoskeleton takes a major part in sensing the geometric information and translates this information into a specific division outcome. In plant cells, the division plane is selected before mitosis, and spatial information of the division plane is preserved throughout division by the presence of reference molecules at a distinct region of the plasma membrane, the cortical division zone. The recruitment of these division zone markers occurs multiple times by several mechanisms, suggesting that the cortical division zone is a highly dynamic region.     

Winding threads around plant cells: a geometrical model for microfibril deposition

In this paper, a geometrical model is put forward to account for the deposition orientation of plant cell wall microfibrils (CMFs). The model presupposes the insertion in the plasma membrane of CMF initiation complexes, which, once inserted, are moved through the fluid plane of the plasma membrane by the kinetic force of CMF synthesis, leaving CMFs in their wake. Deposition occurs in a limited space and the CMFs are linked to wall matrix molecules. CMF orientation is governed by the laws of geometry and, taking space-limiting conditions into account, therefore depends on (1) cell geometry, (2) the other wall molecules linked to the CMFs, and (3) the number of CMF initiation complexes inserted into the plasma membrane. The model does not exclude the idea that cortical microtubules may determine initial CMF orientation after cell division by determining the cell elongation direction.     

The Effect of Isoproterenol upon the Chemical Composition of Plasma Membranes in the Mouse Parotid Gland

A single intraperitoneal injection of isoproterenol induces resting cells from the acini of the mouse parotid gland to enter the proliferative cycle. Parotid plasma membrane from non-stimulated and isoproterenol-treated mice were prepared by differential centrifugation of the homogenates. Comparing the chemical composition of plasma membranes from non-stimulated and isoproterenol-treated mice, no variation in the phospholipid/protein ratio was observed. However, the levels of neutral sugars, hexosamines and sialic acid falls drastically in the early prereplicative phase. The decrease in neutral sugars and hexosamines in plasma membranes caused by isoproterenol is imitated by pilocarpine, which induces secretion but little or no increase in DNA synthesis. However, pilocarpine does not mobilize sialic acid from the plasma membrane. Moreover, dosis of isoproterenol that elicits secretion but not mitosis in the acinar cells, does not induce the movement of sialic acid from the plasma membrane. The mobilization of sialic acid from plasma membranes caused by isoproterenol was also demonstrated in an in vitro system. Treatment of the plasma membrane with chloro-form/methanol shows that around 60% of the sialic acid is present in the less polar phase. We conclude that the separation of sialic acid from the plasma membrane is one of the early steps in the sequence of events leading to DNA synthesis and cell division in the isoproterenol-stimulated parotid gland of mice.     

Membrane trafficking during plant cytokinesis

Plant morphogenesis is regulated by cell division and expansion. Cytokinesis, the final stage of cell division, culminates in the construction of the cell plate, a unique cytokinetic membranous organelle that is assembled across the inside of the dividing cell. Both during cell-plate formation and cell expansion, the secretory pathway is highly active and is polarized toward the plane of division or toward the plasma membrane, respectively. In this review, we discuss results from recent genetic and biochemical research directed toward understanding the molecular events occurring during cytokinesis and cell expansion, including data supporting the idea that during cytokinesis one or more exocytic pathways are polarized toward the division plane. We will also highlight recent evidence for the roles of secretory vesicle transport and cytoskeletal machinery in cell-plate membrane trafficking and fusion.     

Roles of the TRAPP-II Complex and the Exocyst in Membrane Deposition during Fission Yeast Cytokinesis

The cleavage-furrow tip adjacent to the actomyosin contractile ring is believed to be the predominant site for plasma-membrane insertion through exocyst-tethered vesicles during cytokinesis. Here we found that most secretory vesicles are delivered by myosin-V on linear actin cables in fission yeast cytokinesis. Surprisingly, by tracking individual exocytic and endocytic events, we found that vesicles with new membrane are deposited to the cleavage furrow relatively evenly during contractile-ring constriction, but the rim of the cleavage furrow is the main site for endocytosis. Fusion of vesicles with the plasma membrane requires vesicle tethers. Our data suggest that the transport particle protein II (TRAPP-II) complex and Rab11 GTPase Ypt3 help to tether secretory vesicles or tubulovesicular structures along the cleavage furrow while the exocyst tethers vesicles at the rim of the division plane. We conclude that the exocyst and TRAPP-II complex have distinct localizations at the division site, but both are important for membrane expansion and exocytosis during cytokinesis.     

The Arabidopsis KNOLLE Protein Is a Cytokinesis-specific Syntaxin

In higher plant cytokinesis, plasma membrane and cell wall originate by vesicle fusion in the plane of cell division. The Arabidopsis KNOLLE gene, which is required for cytokinesis, encodes a protein related to vesicle-docking syntaxins. We have raised specific rabbit antiserum against purified recombinant KNOLLE protein to show biochemically and by immunoelectron microscopy that KNOLLE protein is membrane associated. Using immunofluorescence microscopy, KNOLLE protein was found to be specifically expressed during mitosis and, unlike the plasma membrane H⁺-ATPase, to localize to the plane of division during cytokinesis. Arabidopsis dynamin-like protein ADL1 accumulates at the plane of cell plate formation in knolle mutant cells as in wild-type cells, suggesting that cytokinetic vesicle traffic is not affected. Furthermore, electron microscopic analysis indicates that vesicle fusion is impaired. KNOLLE protein was detected in mitotically dividing cells of various parts of the developing plant, including seedling root, inflorescence meristem, floral meristems and ovules, and the cellularizing endosperm, but not during cytokinesis after the male second meiotic division. Thus, KNOLLE is the first syntaxin-like protein that appears to be involved specifically in cytokinetic vesicle fusion.     

Cell cycle-dependent targeting of a kinesin at the plasma membrane demarcates the division site in plant cells

Eukaryotic cells have developed different mechanisms to establish the division plane. In plants, the position is determined before the onset of mitosis by the preprophase band (PPB). This ring of microtubules surrounds the nucleus and disappears completely by prometaphase. An unknown marker is left behind by the PPB, providing the necessary spatial cues during cytokinesis. At the position of the PPB, cortical actin is removed or modified to generate an actin-depleted zone that was proposed to provide the structural means for phragmoplast guidance. Here, we identify a plasma membrane domain that emerges at the onset of mitosis and persists until the end of cytokinesis. The narrow band in the plasma membrane corresponds to the position of the PPB and is prevented from accumulation of a GFP-tagged kinesin GFP-KCA1; hence, it is called the KCA-depleted zone (KDZ). The KDZ demarcates the cortical division site independent from the mitotic cytoskeleton. Cell divisions in the absence of a KDZ resulted in misplaced cell plates, suggesting that the PPB transmits a signal to the plasma membrane required for correct cell plate guidance and vesicular targeting to the cortical division site.     

Microspore ofSecale cereale as a transfer cell type

Summary In the mature microspore ofSecale cereale, a set of wall ingrowths deposited as the first (outer) intine layer between exine and the microspore plasma membrane, are revealed by electron microscopy. The wall ingrowths form a girdle in the vicinity of the apertural region at the external pole of microspore which is in contact with the tapetum, so the microspore can be considered as a transfer cell which is polarized. After microspore division the second (inner) intine layer is deposited by the vegetative cell and forms a labyrinth of branched wall ingrowths. As a result, the periphery of a vegetative cell is also irregular and appears as very thin plasmatubules or evaginations delimited by plasma membrane and penetrating the pollen wall.The possible functions of the microspore as a transfer cell and the wall-membrane system of the vegetative cell are discussed.     

Callose deposition and breakdown, followed by phytomelan synthesis in the seed coat ofGasteria verrucosa (Mill.) H. Duval

Summary In the seed coat ofGasteria verrucosa the deposition of phytomelan takes place during seed development in three stages. Phytomelan is a black cell wall material which is chemically very inert. First the radial walls and part of the transverse cell wall of the outer epidermis of the outer integument become thickened by exocytosis of dictyosome vesicles. Callose is deposited at the tangential plasma membrane against those walls. After the callose deposition about two thirds of the original cell volume is filled with callose. During the second stage the callose is broken down, probably into glucose monomers or small polymers. At the same time cellulose is deposited at the outer tangential plasma membrane, forming a wall between the dissolving callose and the plasma membrane. In the third phase small granules appear in the solution of dissolved callose. which grow out and finally fuse to form a block of phytomelan, consisting of spherical 15-nm units. Remarkable is the function of the callose: it determines the size of the phytomelan block, and it probably functions as carbohydrate source for the phytomelan synthesis and/or for the cellulose inner layer. In this study transmission electron microscopy and cryo scanning electron microscopy are used to study the three developmental stages of the formation of the phytomelan layer.     

Protein synthesizing cell apparatus in malignant growth and pharmacotherapy by chlofiden

Data on the influence of a new antitumor preparation chlofiden on the general contents of rat liver ribosomes and sarcoma 45 and their division on free and membrane of membrane bound and decrease of free ribosomes during tumor growth supposed synthesis of specific proteins bound are given in the paper. It was shown that in the liver of tumor bearing rats total and membrane bound ribosomes decreased and the level of free ribosomes increased. High contents of free ribosomes in sarcoma 45 may testify increase of intracellular protein synthesis including processes of cell growth and division as well as the tendency for increase. Chlofiden normalized total contents, increased free and decreased liver membrane bound ribosomes contents, during tumor growth supposed synthesis of specific proteins. Increase of free ribosomes and decrease of their specific radioactivity in sarcoma 45 testified membrane damage by chlofiden and inhibition of intracellular protein synthesis which are essential in cell division.     

Differential localization of LTA synthesis proteins and their interaction with the cell division machinery in Staphylococcus aureus

Lipoteichoic acid (LTA) is an important cell wall component of Gram‐positive bacteria. In Staphylococcus aureus it consists of a polyglycerolphosphate‐chain that is retained within the membrane via a glycolipid. Using an immunofluorescence approach, we show here that the LTA polymer is not surface exposed in S. aureus, as it can only be detected after digestion of the peptidoglycan layer. S. aureus mutants lacking LTA are enlarged and show aberrant positioning of septa, suggesting a link between LTA synthesis and the cell division process. Using a bacterial two‐hybrid approach, we show that the three key LTA synthesis proteins, YpfP and LtaA, involved in glycolipid production, and LtaS, required for LTA backbone synthesis, interact with one another. All three proteins also interacted with numerous cell division and peptidoglycan synthesis proteins, suggesting the formation of a multi‐enzyme complex and providing further evidence for the co‐ordination of these processes. When assessed by fluorescence microscopy, YpfP and LtaA fluorescent protein fusions localized to the membrane while the LtaS enzyme accumulated at the cell division site. These data support a model whereby LTA backbone synthesis proceeds in S. aureus at the division site in co‐ordination with cell division, while glycolipid synthesis takes place throughout the membrane.     

Cortical division zone establishment in plant cells

Plant cell division is spatially organized to maintain a critical cell volume and to control growth directionality. The correct orientation of the separating cell wall is secured by means of specialized cytoskeletal structures that guide the newly formed cell plate toward a predefined cortical position. A ring of microtubules called preprophase band defines a cortical zone that corresponds to the future division plane. Coincident with the disappearance of the preprophase band microtubules, cortical actin is removed at the corresponding position, leaving an actin-depleted zone that persists throughout mitosis. Here, we review the spatial and structural organization of the cortical division zone and discuss evidence that implicate the plasma membrane in division plane establishment.     

On the biomechanics of cytokinesis in animal cells

The material properties of the cell membrane are discussed. Various theories concerning the mechanism of cytokinesis in animal cells are presented. The currently accepted mechanism is that of active muscle-like contraction of the furrow base itself. A mathematical model is developed based on this theory. The cell membrane is modelled as a spherical membrane of nonlinear, elastic material. The membrane undergoes large deformations under the action of a contractile ring force in its equatorial plane. The numerical procedure employed in the solution of the governing equations is explained. The numerical results are compared with the experimental observations available in the literature. It is concluded that the cell membrane stiffness increases during the early stages of cleavage and it, later, decreases. The cell membrane division is a biomechanical instability problem. The factors that may facilitate or block cleavage are discussed. The experimental evidences that support the conjectures of the model are pointed out.     

Functional characterization of the KNOLLE-interacting t-SNARE AtSNAP33 and its role in plant cytokinesis

Cytokinesis requires membrane fusion during cleavage-furrow ingression in animals and cell plate formation in plants. In Arabidopsis, the Sec1 homologue KEULE (KEU) and the cytokinesis-specific syntaxin KNOLLE (KN) cooperate to promote vesicle fusion in the cell division plane. Here, we characterize AtSNAP33, an Arabidopsis homologue of the t-SNARE SNAP25, that was identified as a KN interactor in a yeast two-hybrid screen. AtSNAP33 is a ubiquitously expressed membrane-associated protein that accumulated at the plasma membrane and during cell division colocalized with KN at the forming cell plate. A T-DNA insertion in the AtSNAP33 gene caused loss of AtSNAP33 function, resulting in a lethal dwarf phenotype. atsnap33 plantlets gradually developed large necrotic lesions on cotyledons and rosette leaves, resembling pathogen-induced cellular responses, and eventually died before flowering. In addition, mutant seedlings displayed cytokinetic defects, and atsnap33 in combination with the cytokinesis mutant keu was embryo lethal. Analysis of the Arabidopsis genome revealed two further SNAP25-like proteins that also interacted with KN in the yeast two-hybrid assay. Our results suggest that AtSNAP33, the first SNAP25 homologue characterized in plants, is involved in diverse membrane fusion processes, including cell plate formation, and that AtSNAP33 function in cytokinesis may be replaced partially by other SNAP25 homologues.     

Cellular Mechanism of Myelination in the Central Nervous System

A study of myelination with electron microscopy has been carried out on the spinal cord of young rats and cats. In longitudinal and transverse sections the intimate relationship of the growing axons with the oligodendrocytes was observed. Early naked axons appear to be embedded within the cytoplasm and processes of the oligodendrocytes from which they are limited only by the intimately apposed membranes of both elements (axon-oligocytic membrane). In a transverse section several axons are observed to be in a single oligodendrocyte. The process of myelination consists in the laying down, within the cytoplasm of the oligodendrocyte and around the axon, of concentric membranous myelin layers. The first of these layers is deposited at a certain distance (200 to 600 A or more) from the axon-oligocytic membrane. This and all the other subsequently formed membranes have higher electron density and are apparently formed by the coalescence and fusion of vesicles (of 200 to 800 A) and membranes found in large amounts within the cytoplasm of the oligodendrocytes. At an early stage the myelin layers may be discontinuous and some vesicular material may even be trapped among them or between the myelin proper and the axon-oligocytic membrane. Then, when the 8th to 10th layer is deposited, the complete coalescence and alignment of the lamellae leads to the characteristic orderly multilayered organization of the myelin sheath. Myelination in the central nervous system appears to be a process of membrane synthesis within the cytoplasm of the oligodendrocyte and not a result of the wrapping of the plasma membranes as postulated in Geren's hypothesis for the peripheral nerve fibers. The possible participation of Schwann cell cytoplasm in peripheral myelination is now being investigated.     

Endocytosis restricts Arabidopsis KNOLLE syntaxin to the cell division plane during late cytokinesis

Cytokinesis represents the final stage of eukaryotic cell division during which the cytoplasm becomes partitioned between daughter cells. The process differs to some extent between animal and plant cells, but proteins of the syntaxin family mediate membrane fusion in the plane of cell division in diverse organisms. How syntaxin localization is kept in check remains elusive. Here, we report that localization of the Arabidopsis KNOLLE syntaxin in the plane of cell division is maintained by sterol-dependent endocytosis involving a clathrin- and DYNAMIN-RELATED PROTEIN1A-dependent mechanism. On genetic or pharmacological interference with endocytosis, KNOLLE mis-localizes to lateral plasma membranes after cell-plate fusion. Fluorescence-loss-in-photo-bleaching and fluorescence-recovery-after-photo-bleaching experiments reveal lateral diffusion of GFP-KNOLLE from the plane of division to lateral membranes. In an endocytosis-defective sterol biosynthesis mutant displaying lateral KNOLLE diffusion, KNOLLE secretory trafficking remains unaffected. Thus, restriction of lateral diffusion by endocytosis may serve to maintain specificity of syntaxin localization during late cytokinesis.