Cytokinesis: relative alignment of the cell division apparatus and the mitotic spindle

The cell division apparatus is assembled at different stages of the cell cycle in different eukaryotic organisms. Mechanisms exist in all organisms, however, to ensure that the cell division apparatus and the mitotic spindle are aligned perpendicular to each other. Such an alignment ensures that each daughter cell receives a nucleus and that the cell division apparatus does not cleave and destroy the genetic material. The interaction(s) of astral microtubules with the cell cortex appears to play an important role in establishing perpendicularity between chromosome segregation and cell division machinery.     

Ultrastructure of the Giant Amoeba Pelomyxa palustris*

SYNOPSIS. The ultrastructure of the herbivorous amoeba Pelomyxapalustris was studied. Nuclear division is not understood in this amoeba, and evidence for the method of nuclear division was sought. This species typically has many spheroidal nuclei which are similar within a given cell. However, some amoebae from our collections differed from this common type in both the number and structure of their nuclei. This suggested stages associated with nuclear division. One current hypothesis of nuclear division in this organism is that of nuclear budding. Our evidence is more in accord with this method than with mitosis. The cytoplasm contained no mitochondria, Golgi bodies, contractile vacuoles or crystals. Most amoebae had 2 types of bacteria (bacteroids or endosymbionts) in their cytoplasm; a separate vesicle enclosed each of these. Characteristically, only 1 type of bacterium (B_n) surrounded the nucleus. Another type (B) was found elsewhere in the cytoplasm. Also in the cytoplasm were the following: food vacuoles enclosing various algae, relatively clear vacuoles and vesicles, glycogen, various electron-opaque particles, and occasional microtubules. The plasmalemma was smooth, lacking the external fringe which characterizes other large fresh-water amoebae.     

Electron Microscope Studies of Nuclear Changes in Saccharomyces Cerevisiae During Bud Formation

Ultraviolet photolysis was used to obtain transparent yeast cells when observed with the electron microscope. Following proper irradiation the yeast nucleus was easily discernable and appeared to be in various stages of division. Spindle-like figures were seen which give weight to the concept of a mitotic process for the division of the yeast nucleus. Distinct chromosomes were not observed.     

Three new species of dicyemid mesozoans (phylum Dicyemida) from Amphioctopus fangsiao (Mollusca: Cephalopoda), with comments on the occurrence patterns of dicyemids

Three new species of dicyemid mesozoans are described from the renal appendages of Amphioctopus fangsiao, collected off Akashi, in Harima Nada, and from Osaka Bay. Dicyema akashiense n. sp. is a small species that reaches about 900 microm in length. The vermiform stages are characterized as having 15-17 peripheral cells, a conical calotte, and an axial cell that extends to the base of the metapolar cells. Infusoriform embryos consist of 37 cells; two nuclei are present in each urn cell, and the refringent bodies are solid. Dicyema helocephalum n. sp. is a small species that reaches about 800 microm in length. The vermiform stages are characterized as having 22 peripheral cells, a disc-shaped calotte, and an axial cell that extends to the base of the propolar cells. Infusoriform embryos consist of 37 cells; a single nucleus is present in each urn cell, and the refringent bodies are solid. Dicyema awajiense n. sp. is a small species that reaches about 300 microm in length. The vermiform stages are characterized as having 22 peripheral cells, a conical calotte, and an axial cell that extends to the middle of the propolar cells. Infusoriform embryos consist of 37 cells; a single nucleus is present in each urn cell, and the refringent bodies are solid. In A. fangsiao various occurrence patterns of dicyemid species were observed, including instances where different dicyemid species were found in the renal appendage on each side. This suggests that dicyemids infect each renal appendage independently. The prevalence, reproductive traits, calotte shapes, and co-occurrence patterns of dicyemids are briefly discussed.     

Characteristics of Isolated Microsporocytes from Liliaceous Plants for Studies of the Meiotic Cell Cycle in Vitro

Experiments were aimed at obtaining a culture system for studyof the differentiation of meiotic cells, and techniques weresuccessfully established for the in vitro culture of premeioticmicrosporocytes isolated from various species of liliaceousplants. The problems concerning the culture system were whethermicrosporocytes could be isolated at each stage of the premeioticinterphase and whether they would survive and undergo cell divisionat high frequencies in culture media. In the species tested,the range of stages permitting isolation from anthers and thecytological features of the extruded cells were examined insome detail. Culture conditions are defined for the survivaland the induction of cell division in explanted cells, and somefactors influencing these frequencies are discussed. Explantedcells at premeiotic Gi and S phases underwent a mitotic division,whereas once cells entered the leptotene stage they completedmeiosis. Cells explanted at the G2 phase showed abnormal typesof meiotic division. The culture system developed makes possiblecytological, physiological and biochemical analyses of premeioticcells cultured under various conditions. (Received September 5, 1980; Accepted November 5, 1980)     

Meiotic division and fusion of nuclei in multinucleate cells from the induced fusion of meiotic protoplasts of liliaceous plants

Multinucleate protoplasts were produced from meiotic cells at the zygotene and pachytene stages in a lily andTrillium, and their meiotic divisions were followed during subsequent culture. In each multinucleate, a complete synchrony of nuclear division was maintained throughout the meiotic process, and chromosome behavior appeared normal up to the metaphase stage. In most dinucleates, chromosome segregation movement was organized in a common spindle, and the daughter nuclei at the telophase appeared to envelope each other in the newly formed nuclear membrane. The cell was divided into two daughter cells by a common cell plate. Trinucleates were similarly converted to two cells with a hexaploid number of chromosomes. Some of the di- and trinucleates subsequently completed the second meiotic division with the formation of typical tetrad configurations. In giant cells with more than several nuclei, chromosomes separated at random but reaggregated into one giant resting nucleus, with no later cytokinesis. The rate of meiotic development in multinucleates was relatively slower in cells which contained greater numbers of nuclei.     

Three-dimensional reconstructions of the mitotic spindle and dense plaques in three species of Leishmania

The ultrastructure of the mitotic nucleus in Leishmania braziliensis braziliensis, L. mexicana and L. donovani was studied by serial thin sections and three-dimensional reconstructions of each divisional stage. The structures of the interphase and four stages of dividing nuclei were described. Attention was paid to dense plaques and spindle microtubules. At the beginning of the nuclear division, a set of six dense plaques was found in association with spindle microtubules in the vicinity of the equatorial region of the nucleus. The number of the plaques was the same in the three species examined. Each plaque was divided into two, forming hemiplaques at the elongational stage of the division; these two sets then migrate to the poles. The plaques appeared to correspond with centromeres of metazoan cells and play an important role in the process of nuclear division.     

Enterocytozoon salmonis n. sp.: an intranuclear microsporidium from salmonid fish

The developmental stages of a recently described microsporidian from the nucleus of hematopoietic cells of salmonid fish were found to be unique among the Microsporida. All observed stages, including meronts, sporonts, and spores were in direct contact with the host cell nucleus (principally hematopoietic cells) of chinook salmon (Oncorhynchus tshawytscha). There is no parasitophorous vacuole and sporogony does not involve formation of a pansporoblastic membrane as with other members of the suborder Apansporoblastina. The extrusion apparatus differentiates prior to division of sporogonial plasmodia. The spores are ovoid (1 x 2 microns) and uninucleate, and possess a coiled polar tube with 8-12 turns. Developmental stages of the salmonid microsporidian are similar to those described for Enterocytozoon bieneusi as found in the intestinal mucosa of human AIDS patients. However, the intranuclear development, different cell types, and host infected clearly separate the salmonid and human parasites. Accordingly, the intranuclear parasite of salmonids is given the name Enterocytozoon salmonis n. sp. within the suborder Apansporoblastina.     

DIVISION IN THE DINOFLAGELLATE GYRODINIUM COHNII (SCHILLER) : A New Type of Nuclear Reproduction

Dinoflagellates are of interest because their chromosomes resemble the nucleoplasm of prokaryotes both chemically and ultrastructurally. We have studied nuclear division in the dinoflagellate Gyrodinium cohnii (Schiller), using cells obtained from cultures undergoing phasic growth. Electron micrographs of serial sections were used to prepare three-dimensional reconstructions of nuclei and chromosomes at various stages of nuclear division. During division, a complex process of invagination of the intact nuclear envelope takes place at one side of the nucleus and results in the formation of parallel cylindrical cytoplasmic channels through the nucleus. These invaginations contain bundles of microtubules, and each of the bundles comes to lie in the cytoplasm of a cylindrical channel. Nuclear constriction occurs perpendicular to these channels without displacement of the microtubules. There are no associations between chromosomes and the cytoplasmic microtubules. In dividing cells most chromosomes become V-shaped, and the apices of the V's make contact with the membrane surrounding cytoplasmic channels. It is proposed that the membrane surrounding cytoplasmic channels in the dividing nucleus may be involved in the separation of daughter chromosomes. Thus, dinoflagellates may resemble prokaryotes in the manner of genophore separation as well as in genophore chemistry and ultrastructure.     

DNA SYNTHESIS,CELL DIVISION,AND CELL DIFFERENTIATION DURING LEAF DEVELOPMENT OF XANTHIUM PENNSYLVANICUM

Leaf growth consists of two basic processes, cell division and cell enlargement. DNA synthesis is an integral part of cell division and can be studied with autoradiographic techniques and incorporation of some labeled precursor. Studies were made on the synthesis of nuclear DNA through incorporation of ³H-thymidine in various parts of the lamina during the entire course of leaf development of Xanthium pennsylvanicum. The time course analysis of DNA synthesis was correlated with cell division and rates of cell enlargement. Significant differences in ³H-thymidine incorporation were found in various parts of the lamina. Cell division and DNA synthesis were highest in the early stages of development. Since no ³H-thymidine was incorporated after cessation of cell division (LPI 2.8) in the leaf lamina, it appears that DNA synthesis is not needed for enlargement and differentiation of Xanthium cells. Rates of cell enlargement were negligible in the early development and reached their maximum after cessation of mitoses, between plastochron ages (LPI) 3 and 4. Cells matured between LPI's 5 and 6. Enzymatic activity was correlated with cell division and cell differentiation at various stages of leaf development.     

Nuclear and division-plane positioning revealed by optical micromanipulation

The position of the division plane affects cell shape and size, as well as tissue organization. Cells of the fission yeast Schizosaccharomyces pombe have a centrally placed nucleus and divide by fission at the cell center. Microtubules (MTs) are required for the central position of the nucleus. Genetic studies lead to the hypothesis that the position of the nucleus may determine the position of the division plane. Alternatively, the division plane may be positioned by the spindle or by morphogen gradients or reaction diffusion mechanisms. Here, we investigate the role of MTs in nuclear positioning and the role of the nucleus in division-plane positioning by displacing the nucleus with optical tweezers. A displaced nucleus returned to the cell center by MT pushing against the cell tips. Nuclear displacement during interphase or early prophase resulted in asymmetric cell division, whereas displacement during prometaphase resulted in symmetric division as in unmanipulated cells. These results suggest that the division plane is specified by the predividing nucleus. Because the yeast nucleus is centered by MTs during interphase but not in mitosis, we hypothesize that the establishment of the division plane at the beginning of mitosis is an optimal mechanism for accurate symmetric division in these cells.     

牡丹愈伤组织发生和分生结节形成的细胞组织学研究

以牡丹‘Golden era’叶柄薄层为外植体,结合外部形态,对愈伤组织发生,并分化为分生结节过程进行了细胞组织学研究,为进一步研究牡丹不定器官分化、实现离体微繁殖奠定基础。结果表明:愈伤组织的发生与发育经历了启动、分裂、形成和组织分化4个时期。启动期,外植体束中形成层最先去分化,表现为形态上变暗,内部细胞核变大,淀粉代谢增强。细胞旺盛增殖的分裂期,先后于外植体形成层和皮层薄壁细胞处产生了愈伤组织,形成了质地紧实的愈伤增殖团,愈伤细胞体积小、质浓、核位于中心,且具有绕核积累的淀粉粒。形成期,愈伤组织增殖速度减缓,只有接触培养基的周缘愈伤组织仍在增殖。组织分化期,愈伤组织内部分化出维管单元,可以作为生长中心,有的是以特化的薄壁细胞为生长中心,外围核大、质浓的薄壁细胞层形成了具有再生潜能的分生结节。     

Microtubule arrays and Arabidopsis stomatal development

Microtubule arrays in living cells were analysed during Arabidopsis stomatal development in order to more closely define stages in the pathway and contexts where intercellular signalling might operate. Arabidopsis stomata are patterned iteratively via the orientation of an asymmetric division in a cell located next to an existing stoma. It was found that preprophase bands of microtubules (PPBs) were correctly placed away from stomata and from two types of precursor cells. This suggests that all three cell types participate in an intercellular signalling pathway that orients the division site. These and other asymmetric divisions in the pathway were preceded by a polarized cytoplasm, with the PPB around the nucleus at one end, and the vacuole at the other. PPBs before symmetric divisions of guard mother cells (GMCs) were broader than those in asymmetric divisions, and the GMC division site was marked by unusual end-wall thickenings. This work identifies an accessible system for studying cytoskeletal function and provides a foundation for analysing the role of genes involved in stomatal development.     

A fixed amount of chromosome replication needed for premature division septation in Bacillus subtilis

Germinating spores of the temperature-sensitive DNA initiation mutant of Bacillus subtilis, TsB134, were allowed to undergo a single round of replication, at a high and low level of thymine, by shifting to the non-permissive temperature shortly after its initiation. The rate of replication at the low thymine level was approximately half that at the other, but there was no significant difference in the rate of cell mass increase. The round of replication in each case was blocked at various stages by 6-(p-hydroxyphenylazo)uracil and outgrown cells examined at a later time for the frequency of central division septation. It was found that the same average amount of replication (fraction of the round) was required in both cases for premature division septation to proceed.     

Nucleus-associated microtubules help determine the division plane of plant epidermal cells: avoidance of four-way junctions and the role of cell geometry

To investigate the spatial relationship between the nucleus and the cortical division site, epidermal cells were selected in which the separation between these two areas is large. Avoiding enzyme treatment and air drying, Datura stramonium cells were labeled with antitubulin antibodies and the three-dimensional aspect of the cytoskeletons was reconstructed using computer-aided optical sectioning. In vacuolated cells preparing for division, the nucleus migrates into the center of the cell, suspended by transvacuolar strands. These strands are now shown to contain continuous bundles of microtubules which bridge the nucleus to the cortex. These nucleus-radiating microtubules adopt different configurations in cells of different shape. In elongated cells with more or less parallel side walls, oblique strands radiating from the nucleus to the long side walls are presumably unstable, for they are progressively realigned into a transverse disc (the phragmosome) as broad, cortical, preprophase bands (PPBs) become tighter. The phragmosome and the PPB are both known predictors of the division plane and our observations indicate that they align simultaneously in elongated epidermal cells. These observations suggest another hypothesis: that the PPB may contain microtubules polymerized from the nuclear surface. In elongated cells, the majority of the radiating microtubules, therefore, come to anchor the nucleus in the transverse plane, consistent with the observed tendency of such cells to divide perpendicular to the long axis. In nonrectangular isodiametric epidermal cells, which approximate regular hexagons in section, the radial microtubular strands emanating from the nucleus tend to remain associated with the middle of each subtending cell wall. The strands are not reorganized into a single dominant transverse bar, but remain as a starlike array until mitosis. PPBs in these cells are not as tight; they may only be a sparse accumulation of microtubules, even forming along non-diametrical radii. This arrangement is consistent with the irregular division patterns observed in epidermal mosaics of isodiametric D. stramonium cells. The various conformations of the radial strands can be modeled by springs held in two-dimensional hexagonal frames, and by soap bubbles in three-dimensional hexagonal frames, suggesting that the division plane may, by analogy, be selected by minimal path criteria. Such behavior offers a cytoplasmic explanation of long-standing empirically derived "rules" which state that the new cell wall tends to meet the maternal wall at right angles. The radial premitotic strands and their analogues avoid taking the longer path to the vertex of an angle where a cross wall is already present between neighboring cells.(ABSTRACT TRUNCATED AT 400 WORDS)     

DIPLOIDY IN DNA CONTENT IN VEGETATIVE CELLS OF CLOSTERIUM EHRENBERGII (CHLOROPHYTA)1

DNA content of the nucleus in the placoderm desmid, Closterium ehrenbergii Meneghini was measured throughout the life cycle by epifluorescence microspectrophotometry after DNA specific dye [4′,6-diamidino-2-phenylindol (DAPI)] staining. Postulating a mean DNA content of gamete nuclei as 1C, the nucleus of a newly divided vegetative cell was 2C. Most vegetative cells in the stage of exponential growth had a DNA content from 2C to 4C, while most in stationary phase, with the highest frequency of zygote formation, were 2C. They became pre-gametes (2C) upon mixing two heterothallic strains. Four gametes were made by a DNA reduction division of each pre-gamete cell. Therefore, there was a nonmeiotic DNA reduction stage by one half. During germination, the zygote underwent meiosis to produce two gones, each of which contained one surviving nucleus (large nucleus) and one degenerating nucleus (small nucleus). The DNA content of these four nuclei was 1C basically. The DNA of the surviving nucleus duplicated to 2C and further quadruplicated to 4C without cell or nuclear division. These two 4C gones had different cell morphology from ordinary vegetative cells. After the first cell division following meiosis, each gone produced two vegetative cells in which the DNA content became 2C to 4C again.     

Nuclear functions in postconjugational development of Paramecium caudatum: Ability to form food vacuoles analyzed by nuclear elimination and implantation

Paramecium caudatum loses the ability to form food vacuoles at the crescent stage of the micronucleus from 5 to 6 hr after the initiation of conjugation and regains it immediately after the third division of the zygotic nucleus. To assess the micronuclear function in the development of the oral apparatus after coniugation, prezygotic micronuclei was removed from cells at various stages of conjugation, and their ability to form food vacuoles were examined. (1) When all of the prezygotic micronuclear derivatives were eliminated before the stage of formation of the zygotic nucleus, the exconjugant did not regain its ability. (2) When a zygotic nucleus or postzygotic nuclei were removed, in some cases the cell formed as many food vacuoles as did nonoperated cells after conjugation, while in other operated cells the number of food vacuoles was subnormal. (3) When a micronucleus from a cell at vegetative phase (G1) was transplanted into a cell of an amicronucleate mating pair at the stage between 8 and 9 hr after the initiation of conjugation, the implanted cell regained the ability to form food vacuoles. However, no cell regained the ability when the implantation was carried out within 1 hr after the separation of the mates. The results show that the micronucleus plays an indispensable role in the development of the oral apparatus at the stages of exchange of gametic nuclei and fertilization and that the micronucleus transplanted from asexual cells can fulfill this function. On the other hand, removal of the macronucleus from exconjugants showed that the maternal macronucleus also has an indispensable function in regaining the ability to form food Vacuoles. © 1992 Wiley-Liss, Inc.     

Nuclear division of the vegetative cells, conchosporangial cells and conchospores of Porphyra yezoensis (Bangiales, Rhodophyta)

To confirm the position and timing of meiosis in Porphyra yezoensis Ueda, the nuclear division of vegetative cells, conchosporangial cells and conchospores was observed. An improved staining method using modified carbol fuchsin was introduced to stain the chromosomes of Porphyra. Pit‐connections between conchosporangial cells also stained well with this method. Leptotene, zygotene, pachytene, diplotene, diakinesis, metaphase, anaphase and telophase were observed in the conchosporangial cells. During the germination of conchospores, no characteristics of meiosis I were found. No difference between the nuclear division of vegetative cells and that of conchospores was observed, and 2–3 days were needed for the first cell division both in vegetative cells and conchospores. Therefore, the cell division that occurs during conchospore germination is not meiosis I. Our results indicate that the prophase of meiosis I begins during the formation of conchosporangial branches, and metaphase I, anaphase I and telophase I take place during the maturation of conchosporangial branches. Then the three‐bivalent nucleate sporangia complete cell division to form two individual conchospores, each with one three‐univalent nucleus. The conchospores released from the sporangia are at meiotic interphase. Meiosis II occurs at the first nuclear division during conchospore germination, which is a possible explanation for the observation of mosaic thalli in mutant germlings of P. yezoensis. The mosaic thalli might also arise from gene conversion/post meiotic segregation events, comparable to those in Sordaria fimicola (Roberge ex Desm.) Ces. & De Not. and Neurospora crassa Shear & B.O. Dodge.     

Control of F′lac Replication in Escherichia coli B/r

The timing of replication of an F'lac plasmid during the division cycle of Escherichia coli B/r lac(-)/F'lac was examined in relation to the timing of initiation of chromosome replication. This was accomplished by measuring the induction of beta-galactosidase and the incorporation of radioactive thymidine into cells at different ages in cultures growing exponentially at various rates. In cells growing with interdivision times of 27, 36, and 55 min, the F'lac replicated at various stages in the division cycle but always at approximately the same time as initiation of chromosome replication. In cells growing with an interdivision time of 85 min, the F'lac episome replicated midway through the division cycle, whereas chromosome replication initiated at the start of the cycle. Measurements of absorbance at 450 nm per cell suggested that the F'lac replicated when the cells reached a mass which was a constant multiple of the number of episomes per cell at each growth rate. In contrast, the mass per cell at initiation of chromosome replication in cells with an 85-min interdivision time was significantly lower than this constant value. A possible explanation for the apparent coupling between F'lac replication and initiation of chromosome replication at the higher growth rates, and the lack of coupling at the lowest growth rate, is discussed.     

Somatic nuclear division in the sporidia of Ustilago violacea. III. Ultrastructural observations.

The paper provides detailed ultrastructural observations on nuclear division in the smut fungus Ustilago violacea and is based on previous light-microscopic work outlining the division in living and stained cells. The division as in many other Basidiomycetes is not intranuclear, but occurs within a partially disrupted membrane. The division takes place after migration of most of the nucleus into the bud cell, after limited breakdown of the nuclear membrane, and after the formation of a spindle between two spindle-pole bodies (SPB). The remaining part of the nucleus containing the nucleolus is left behind in the parent cell and degenerates there. The SPB, as in other Basidiomycetes, is a dome-shaped relatively structureless body, quite distinct from the flat plaques of many Ascomycetes and the elaborat centrioles of Phycomycetes. The SPB divides shortly before migration into the daughter cell and invariably is located at the apex of the migrating nucleus. Nuclear division is completed when the two masses of chromatin clustered about each of the SPB's are separated as the spindle elongates. One daughter neculeus reforms in the bud and the other is reformed in the mother cell. Cells fixed and stained by conventional light-microscopic methods were examined in the light of the electron-microscopic observations to determine whether these procedures induce artefacts. Aceto-orcein and Giemsa when used cold were found to produce relatively artefact-free preparations. However, previous results in which the cells were warmed gently in these stains are now seen to contain artefacts in the form of contracted chromatinic granules often arranged in chains. These artefacts may provide useful information but clearly they must be interpreted cautiously until the nature of the changes induced by heating are known.