Über den Feinbau von Theka,Pusule und Golgi-Apparat bei dem DinoflagellatenGymnodinium spec.

Zusammenfassung Das Pusulensystem vonGymnodinium spec. besteht aus röhrenförmigen Invaginationen des Plasmalemmas mit zarten Rippen auf der Lumen-Seite, die eng von einer besonderen, perforierten Zisterne umschlossen sind. Die Degeneration der Pusule, der Feinbau der Theka mit Trichocysten-ähnlichen Vesikeln und Kragengruben, die Bildung der Cystenwand, das Stigma und seine Verbindung mit der Geißelbasis und der Golgi-Apparat mit verschiedenen Vesikeltypen werden beschrieben.

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Ultrastructural studies on the theca, pusule, and golgi apparatus in the dinoflagellateGymnodinium spec.

Summary The pusule system ofGymnodinium spec. consists of tubes formed by the invaginated plasmalemma which is ornamented with delicate ribs on its luminal surface and tightly surrounded by a special, perforated cisterna. The degeneration of the pusule, the fine structure of the theca with some peculiar structures (trichocyst-like vacuoles, collared pits, the developing cyst wall), the stigma and its connection with the flagellar base, and the Golgi apparatus with different types of vesicles are described.

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Wir danken Herrn Dr. W. Koch, Göttingen, für die Überlassung derGymnodinium-Kultur und der Deutschen Forschungsgemeinschaft für Sachbeihilfen.     

Zellteilung und Bildung von Innenschalen beiHantzschia amphioxys undAchnanthes coarctata

Before cytokinesis, the identically constructed chromatophores ofHantzschia amphioxys andAchnanthes coarctata are transformed into less effigurated bodies. In normal cytokinesis, the course of mitosis, chromatophore division, and cleavage furrowing are exactly synchronized. The division of the chromatophore appears as a passive process, i.e. intersection by the cleavage furrow. In inequal cell divisions before the formation of inner valves cytokinesis can take place without chromatophore division. Once chromatophore division without mitosis and cytokinesis was observed. InHantzschia there are three types of inner valve formation, inAchnanthes coarctata only two. The inner valves develop under unfavorable growth conditions, the cells possessing them, however, are not resting spores as in some other diatoms. InHantzschia, auxospore formation is suppressed under the cultural conditions used, the cells multiply intensely without diminution.

     

SEXUAL REPRODUCTION AND MEIOSIS IN PERIDINIUM INCONSPICUUM LEMMERMANN (DINOPHYCEAE)

In Peridinium inconspicuum Lemmermann, sexual reproduction occurs in both nitrogen-enriched and nitrogen-deficient media. In this homothallic strain, protoplasmic fusion begins between two thecate gametes; but zygote formation is completed in a space outside the fusing pair. This diploid cell can form a plated theca which is shed as the cell enlarges. This spherical zygote then forms a new non-plated theca. The process of ecdysis and the formation of a new non-plated theca is repeated several times. During this process the zygote gradually elongates and by cytoplasmic infurrowing becomes peanut-shaped. Eventually two cells are formed. The first and second meiotic divisions are greatly separated in time. The first meiotic division occurs in the spherical non-thecate zygote. The second meiotic division can occur in the peanut-shaped zygote before it completes cytokinesis. This meiotic division may not be synchronous, occasionally resulting in a trinucleate stage. Eventually four flagellated, haploid products are produced.     

Solitary Song and its inhibition in some Estrildidae

Summary In the species studied song appears to have two functions; an epigamic function = Display Song, and a contact function = Solitary Song. Solitary Song appears to be common to all the species studied. Its utterance indicates that the bird is unpaired or separated from another individual with which it has formed a bond. InUraeginthus bengalus, U. angolensis, andAmandava amandava Solitary Song is also uttered by the hen in similar circumstances. InLonchura punctulata, A. amandava, andEuodice malabarica song is usually but not completely inhibited by the presence of a mate, in whose absence Solitary Song will be uttered even when other individuals of the same species are present. In the species studied of the generaEstrilda, Lagonosticta, andUraeginthus Solitary Song is inhibited by the continued close proximity of another bird even though this may be of the same sex or of a different species and may elicit aggressive or fleeing reactions; but conditions of close association with a bird other than a suitable mate would presumably only occur under captive conditions. There appears to be a distance factor controlling such inhibition. There is evidence of the inhibition of song due to the presence of a mate in other passerine species.     

Studies in the embryology of compositae

The male and female gametophytes formation, fertilisation, and embryo development were observed inSolidago canadensis andConyza stricta. The anther is tetrasporangiate and its wall development conforms to the Dicotyledonous type. Simultaneous cytokinesis in pollen mother cells results in tetrahedral or isobilateral tetrads. Pollen polymorphism is a common feature inS. canadensis. The ovule is anatropous, unitegmic and tenuinucellate showing an ovular vascular strand. The female archesporium is hypodermal and single celled. The development of the embryo sac is of the Polygonum type. InS. canadensis seed set is completely absent and multiplication occurs by vegetative propagation. InC. stricta fertilisation is progamous. Triple fusion and syngamy occur more or less simultaneously. Endosperm is cellular and the embryogeny corresponds to the Asterad type. A part of the thesis accepted for the Ph. D. degree of the Andhra University.     

A STUDY OF CELL WALL AND FLAGELLA FORMATION DURING CELL DIVISION IN THE SCALY GREEN ALGA,SCHERFFELIA DUBIA (CHLOROPHYTA)1

The thecate green flagellate Scherffelia dubia (Perty) Pascher divides within the parental cell wall into two progeny cells. It sheds all four flagella before cell division, and the maturing progeny cells regenerate new walls and flagella. By synchronizing cell division, we observed mitosis, cytokinesis, cell maturation, flagella extension, and cell wall formation via differential interference contrast microscopy of live cells and serial thin‐section EM. Synthesis of thecal and flagellar scales is spatially and temporally strictly separated. Flagellar scales are collected in a pool during late interphase. Before prophase, Golgi stacks divide, flagella are shed, the parental theca separates from the plasma membrane, and flagellar scales are deposited on the plasma membrane near the flagellar bases. At prophase, Golgi bodies start to synthesize thecal scales, continuing into interphase after cytokinesis. During cytokinesis, vesicles containing thecal scales coalesce near the cell posterior, forming a cleavage furrow that is initially oriented slightly diagonal to the longitudinal cell axis but later becomes transverse. After the progeny nuclei have moved into opposite directions, resulting in a “head to tail” orientation of the progeny cells, theca biogenesis is completed and flagellar scale synthesis resumes. Progeny cells emerge through a hole near the posterior end of the parental theca with four flagella of about 8 μm long. The precise timing of flagellar and thecal scale synthesis appears to be an evolutionary adaptation in a scaly green flagellate for the thecal condition, necessary for the evolution of the phycoplast and thus multicellularity in the Chlorophyta.     

The effects of cytochalasin B and colchicine on the morphogenesis of mitochondria in Drosophila hydei during meiosis and early spermiogenesis

Summary Morphogenesis of mitochondria in male germ cells in cultivated cytocysts begins in early prophase I at which time mitochondria thicken and become ordered along the spindle apparatus during meiosis. At the end of the second meiotic division they aggregate to form the Nebenkern.In the presence of colchicine or cytochalasin B mitochondria are able to begin differentiation, although the correct course of meiosis is not guaranteed. In medium supplemented with colchicine they undergo normal thickening but do not aggregate, in a pattern known from untreated cultures. This may indicate that microtubules are involved in the aggregation process of mitochondria as colchicine is known to inhibit microtubule formation. Moreover, in cell cultures treated with cytochalasin B mitochondrial aggregation does occur; it is concluded that microfilaments, which are sensitive to cytochalasin B, do not play a detectable role in the aggregation of mitochondria.     

Manipulation of cytokinesis affects polar ionic current around the egg of Lymnaea stagnalis

Summary The fertilized egg of the mollusc Lymnaea stagnalis generates a polarized current pattern as measured with the vibrating probe. Here we investigated the basis of these polar ionic currents. Ionic currents were measured around eggs during the second meiotic division after interference with cytokinesis. Cytokinesis was either displaced by centrifugation or inhibited with cytochalasin or nocodazole. Furthermore, ectopic constrictions were induced with lectin treatment. It appeared that the inward current of the animal pole can be displaced by centrifugation and remains associated with the position of the meiotic apparatus. The influence of the meiotic apparatus on the polar current pattern seems to be directly related to membrane constrictions rather than to karyokinesis. This was demonstrated by a change in current density after induction of an ectopic constriction at the vegetal pole and by the abolishment of currents after cytochalasin treatment. Since the location of the outward current was not sensitive to centrifugation, it may be concluded that the vegetal outward current depends upon properties of the vegetal cortex. On the basis of these results, we conclude that the Lymnaea egg generates two types of ionic currents during the second meiotic division. The first is an inward current activated at the site of membrane constrictions. The second is an outward current associated with the vegetal cortex.     

Ultrastructure of the Harmful Unarmored Dinoflagellate Cochlodinium polykrikoides (Dinophyceae) with Reference to the Apical Groove and Flagellar Apparatus

ABSTRACT. The external and internal ultrastructure of the harmful unarmored dinoflagellate Cochlodinium polykrikoides Margalef has been examined with special reference to the apical groove and three‐dimensional structure of the flagellar apparatus. The apical groove is U‐shaped and connected to the anterior sulcal extension on the dorsal side of the epicone. The eyespot is located dorsally and composed of two layers of globules situated within the chloroplast. A narrow invagination of the plasma membrane is associated with the eyespot. The nuclear envelope has normal nuclear pores similar to other eukaryotes but different from the Gymnodinium group with diagnostic nuclear chambers. The longitudinal and transverse basal bodies are separated by approximately 0.5–1.0 μm and interconnected directly by a striated basal body connective and indirectly by microtubular and fibrous structures. Characteristic features of the flagellar apparatus are as follows: (1) a nuclear extension projects to the R1 (longitudinal microtubular root) and is connected to the root by thin fibrous material; (2) fibrillar structures are associated with the longitudinal and transverse flagellar canal; and (3) a striated ventral connective extends toward the posterior end of the cell along the longitudinal flagellar canal. We conclude, based on both morphological and molecular evidence, that Cochlodinium is only distantly related to Gymnodinium.     

The effect of drugs on diatom valve morphogenesis

Summary The effects of various drugs on cell wall (valve) morphogenesis was investigated in three species of diatoms (Pinnularia spp., Surirella robusta, andHantzschia amphioxys) using light microscopy (LM) and scanning electron microscopy (SEM). Treatment ofSurirella with the microtubule (MT) disrupting agent colchicine during early valve formation results in a characteristic malformation of the valve, whereby part of the normally circumferential raphe canal forms as an abnormal protruding lip on the valve surface, located up to 20 m from the edge of the valve. The position of this malformed lip coincides with the location of a microtubule center (MC) at the time of colchicine addition, suggesting that the MC may play a direct role in positioning the tip of the raphe canal during valve formation. The migration of this MC to the tip of the cell during early valve morphogenesis is reversibly inhibited by the metabolic inhibitor 2-4-dinitrophenol (DNP). The effect of colchicine onPinnularia valve formation is less severe, causing occasional malformation of the raphe, but little if any lateral displacement. InHantzschia, colchicine has no effect on the positioning of the raphe, but prolonged exposure causes fusion of the raphe canal with the valve face. Cochicine treatment also results in the absence of the normal curvature at the central interruption in the raphe, as well as abnormal pore formation in this central area. Addition of cytochalasin D during early valve formation inHantzschia causes the raphe canal to form in the center of the valve face, suggesting that the normal translocation of the raphe canal to the valve edge is actindependent. Comparison of valves from control and cytochalasintreatmentHantzschia suggest that the pore spacing within the valve is determined by the position relative to the raphe, and does not depend on whether to pores form on the side (mantle) or the face of the mature valve.Abbreviations DM diatom medium - DNP dinitrophenol - MT microtubule - MC microtubule center - PSS primary silicification site - SDV silica deposition vesicle     

MITOSIS, CELL WALL AND FLAGELLA SYNTHESIS IN SYNCHRONIZED CULTURES OF THE PRASINOPHYTE, SCHERFFELIA DUBIA

Cell division occurs within the parental cell wall, yielding two progeny cells. Since Scherffelia dubia sheds all four flagella prior to cell division, the maturing progeny cells must regenerate new cell walls and flagella during and/or after cytokinesis. To better understand these processes, we have synchronized cell division in cultures of S. dubia and observed all stages of mitosis, cytokinesis, and progeny cell maturation, including flagella and cell wall formation, via DAPI staining of fixed cells, DIC microscopy of live cells embedded in agarose and standard TEM. Microscopical observations revealed the following sequence of events: 1) Golgi stacks divide during late interphase and immediately begin producing theca scales; 2) deflagellation and release of the parental cell wall from the plasma membrane occurs during early prophase; 3) synthesis of theca and flagella scales within the Golgi and/or scale reticulum continues throughout mitosis; 4) during cytokinesis, a coalescence of vesicles containing theca scales at the posterior end of the cell results in a cleavage furrow slightly diagonal to the cells' longitudinal axis (40 min); 5) post-mitotic nascent basal body formation and flagella elongation at the inherited basal bodies (and later at the mature nascent basal bodies) occurs concurrently with continued cell wall synthesis; 6) the cleavage furrow rotates into a transverse position (35 min); 7) reorientation of the nuclei results in a "head to tail" orientation of the maturing progeny cells; and 8) matured progeny cells emerge from the posterior end of the parental theca not before 8 hrs after the onset of mitosis.     

Cellular energy dependent agglutination of rat ascites tumor cells mediated by concanavalin A and Ricinus communis agglutinin

Effect of various metabolic inhibitors on the agglutination of rat ascites tumor cells mediated by concanavalin A and Ricinus communis agglutinin was studied using a quantitative assay method for agglutination in which turbidity of cell suspension is measured. Cell agglutination was inhibited by low temperature, cytochalasin B and inhibitors of energy generating systems without affecting lectin binding, and agglutination was not affected by hydroxyurea, actinomycin D or cycloheximide. The inhibitors of energy generating systems decreased the cellular ATP level and inhibited macromolecular synthesis under the conditions where they inhibited the agglutinations. In contrast, cytochalasin B did not depress the cellular ATP level nor inhibit RNA and protein syntheses. These results suggest that the agglutination is associated with cellular energy dependent processes other than macromolecular synthesis; probably with some cellular surface movements participated by microfilament activity.     

Petiole elongation inRumex species during submergence and ethylene exposure: The relative contributions of cell division and cell expansion

Submergence ofRumex crispus L. andR. palustris Sm. stimulates elongation of the youngest petiole. InR. palustris this response can be mimicked partially by exposure to exogenous ethylene. In both species, petiole elongation induced by ethylene and/or submergence is distributed nearly equally over the whole petiole length and is almost completely attributable to increased cell expansion. InR. acetosa L., extension of the youngest petiole is inhibited by submergence of the whole plant. The strongest growth inhibition within the youngest petiole was observed in the most distal parts and is probably the result of reduced cell expansion.     

CORTICAL F‐ACTIN REORGANIZATION AND A CONTRACTILE RING‐LIKE STRUCTURE FOUND DURING THE CELL CYCLE IN THE RED CRYPTOMONAD,PYRENOMONAS HELGOLANDII1

Cortical F‐actin reorganization during the cell cycle was observed in Pyrenomonas helgolandii U. J. Santore (SAG 28.87) for the first time in Cryptophyta using fluorescein‐isothiocyanate (FITC)–phalloidin staining. In interphase, a number of F‐actin bundles were observed as straight lines running parallel to the long axis of the cell on the cell cortical region. They extended from an F‐actin bundle that runs along the margin of the vestibulum. Although the F‐actin bundles running parallel to the long axis of the cell disappeared during anaphase, they gradually reappeared in telophase. By contrast, the F‐actin bundle along the vestibulum margin remained visible during cytokinesis and dynamically changed following the enlargement of the vestibulum, suggesting that F‐actin was involved in the mechanism of vestibulum enlargement. F‐actins were not found in the cytoplasmic and nucleoplasmic regions throughout the cell cycle. In addition, a contractile ring‐like structure appeared at the cleavage furrow during cytokinesis. Treatment with cytochalasin B and latrunculin B significantly inhibited the formation of cleavage furrow, resulting in forming an abnormal cell with two nuclei, suggesting that cytokinesis in P. helgolandii is controlled by the contractile ring‐like structure constituted of F‐actin.     

Role of multipolar mitoses in the proliferation of multinucleate cells induced by cytochalasin B

A prolonged action of cytochalasin B results in the formation of numerous multipolar mitoses (26%) in Chinese hamster cell cultures. The transition to multipolar mitoses in the presence of cytochalasin B is not accompanied by K-mitotic delay. It is shown that a multipolar mitosis without cytoplasmic division is one of the main causes of multinucleation development in cytochalasin B-treated cultures. After stopping the drug action the cytochalasin B-induced multinucleate cells continue to divide by multipolar mitosis. In this case it completes with cytokinesis and, probably, leads to a decrease in the number of nuclei per cell. The origin of multipolar mitotic apparatus after the action of cytochalasin B is discussed in addition to the role of multipolar mitosis in formation and proliferation of multinucleate cells.     

Cholesterol loading induces a block in the exit of VSVG from the TGN

Recent work from our laboratory demonstrated that increased cellular cholesterol content affects the structure of the Golgi apparatus. We have now investigated the functional consequences of the cholesterol-induced vesiculation of the Golgi apparatus and the role of actin for these changes. The results showed that cholesterol-induced vesiculation and dispersion of the Golgi apparatus is a reversible process and that reversal can be inhibited by cytochalasin D, an actin-disrupting reagent. Furthermore, electron microscopy revealed that jasplakinolide, which stabilizes actin filaments, prevented the dispersion, but not the vesiculation of the Golgi cisternae. Importantly, the different Golgi markers seemed to be separated even after vesiculation. To investigate whether transport through the different steps of the exocytic pathway was affected in cholesterol-treated cells, we visualized ER to plasma membrane transport by using ts045-VSVG-GFP. In COS-1 cells expressing ts045-VSVG-GFP increased cholesterol levels did not affect transport of VSVG into the vesiculated Golgi apparatus. However, increased levels of cholesterol resulted in retention of the nascent G protein in vesicles with the TGN-marker TGN46. Biotinylation of cell surface molecules to quantify arrival of VSVG at the plasma membrane confirmed that cholesterol treatment inhibited export of the VSVG protein. In conclusion, the data show that transport of VSVG into/through a vesiculated Golgi is feasible, but that cholesterol loading inhibits exit of VSVG from the vesicles containing TGN markers. Furthermore, the data illustrate the importance of actin filaments for Golgi structure.     

Hemmung der Cytokinese und Bildung von Riesenzellen beiPoterioochromonas malhamensis durch organische Bleiverbindungen und andere Agenzien

Summary Organic lead compounds inhibit cytokinesis of the chrysophycean flagellatePoterioochromonas malhamensis leading to giant, multinucleate cells. This action on cytokinesis is compared with the long-time effects of various compounds with better known subcellular activities.Calcium (10 mM), and cytochalasin B (up to 100 g/ml) do not visibly influence cytokinesis. Caffeine (1 mM) totally inhibits multiplication of the algae whereas calcium has only a slight and cytochalasin has no effect on this parameter.The other reference-compounds (colchicine, sodium cacodylate, deuterium oxide, local anesthetics, and sodium dodecylsulfate) all inhibit cell multiplication, simultaneously leading to giant multinucleate cells, obviously by inhibition of cytokinesis.The most potent inhibitor of cytokinesis is triethyl lead which was shown to be 250× more effective than colchicine in respect to the molar concentrations.The comparison of the effects of tetraethyl lead and triethyl lead with the reference agents leads to the conclusion that organic lead compounds might inhibit cytokinesis ofPoterioochromonas malhamensis by disintegrating peripheral microtubules and/or by interfering with structures and functions of membranes.

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Verwendete Abkürzungen im Text CB Cytochalasin B - KE Karminessigsäure - KV kontraktile Vakuole - LV Leukosinvakuole - MT Mikrotubuli - SDS Na-Dodecyl-sulfat - TEL Tetraäthylblei - TriEL Triäthylblei     

Golgi apparatus activity and membrane flow during scale biogenesis in the green flagellateScherffelia dubia (Prasinophyceae). II: Cell wall secretion and assembly

Summary Secretion of the cell wall (theca) in the scaly green flagellateScherffelia dubia (Prasinophyceae) has been examined by electron microscopy during cytokinesis. The bi-laminate wall forms by the extracellular amalgamation of two layers of scales produced in the Golgi apparatus (GA). Each mature GA cisterna contains ca. 12,000 scales of two distinct varieties arranged in two layers on the cisternal membrane. GA cisternae undergo turnover and one scale containing cisterna matures from thetransface of each dictyosome every 3–4 minutes. Cisternae then fuse with the plasma membrane at the anterior end of the cell releasing the scales onto the cell surface. The two layers of wall scales integrate on the cell surface in a time-dependent self-assembly process. The first scales deposited commence assembly at the cell posterior and the wall develops anteriorly by edge growth. The daughter cell wall is composed of ca. 1.2 million scales deposited in about 3 hours. Calculations of net membrane flow strongly indicate extensive endocytosis during wall deposition.     

Sequential protein-dependent steps in the cell cycle initiation and completion of division in vitamin B12 replenishedEuglena gracilis

Summary InEuglena gracilis Z, vitamin B₁₂ deficiency arrests cell divisions in S/G 2 phase. After the addition of vitamin B₁₂ to blocked cells, nuclear and cellular divisions begin to be induced between the 3rd and the 4th and between the 4th and the 5th hour respectively; the cell population is doubled after the 11th hour.Addition of cycloheximide either with vitamin B₁₂ or 2 to 6 hours later inhibits the resumption of divisions and blocks cell development in different stages between G 2, karyokinesis and cytokinesis. These results suggest that as a prerequisite protein-dependent steps are required at precise times during the reversibility of blocked cell divisions: at least sequential syntheses of protein concern a) formation of the mitotic spindle and replication of the pellicle; b) completion of the nuclear division; c) progression of the cleavage furrow.