Numerical and structural changes in dictyosomes during zygospore germination ofClosterium ehrenbergii

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

Light, a nitrogen-depleted medium and cell-cell interaction in the conjugation process of Closterium ehrenbergii Meneghini

The conjugation process of heterothallic Closterium ehrenbergiiMeneghini consisted of the first cell division, cell aggregation,the second cell division, papilla formation and zygospore formation.Light irradiation and the depletion of nitrogen in the culturemedium were required for the first cell division and for cellaggregation, but the second cell division, papilla formationand zygospore formation took place independent of the presenceof light and of the depletion of nitrogen. The presence of twopartner strain cells was indispensable for cell aggregationand papilla formation. The first and second cell divisions andzygospore formation took place in the absence of partner straincelb, but interaction between the two strains was indispensableto induce them. (Received January 27, 1979; )     

DETECTION AND EVALUATION OF A NOVEL SEXUAL PHEROMONE THAT INDUCES SEXUAL CELL DMSION OF CLOSTERIUM EHRENBERGII (CHLOROPHYTA)1

Mating type-plus (mt⁺; NIES-228) cells of Closterium ehrenbergii undergo a division to form gamete-shaped cells. This cell division is induced by a substance produced by mating type-minus (mt^?; NIES-229) cells. Light and the presence of mt⁺ cells enhanced production of the substance. The active substance is heat labile and has an apparent molecular mass of 20 kDa. From these results, we conclude that the substance is a novel, proteinaceous sexual pheromone involved in reproduction of Closterium ehrenbergii.     

Changes in osmotic pressure and structure at the initial stages of zygote formation inClosterium ehrenbergii

Summary The behavior of gamete cells ofClosterium ehrenbergii in hypertonic solutions was observed and the significance of changes in osmotic pressure of the protoplasts is discussed in relation to zygote formation. The osmotic pressure of fusing gamete protoplasts was calculated to be 0.063 Osm at the original cell volume. The osmotic pressure of immature gamete protoplasts was 0.24 Osm at incipient plasmolysis. This lowering of cell osmotic pressure may serve to protect the rupture of the plasma membrane during migration of protoplasts in the conjugation tube after dissolution of cell walls. During maturation of gamete cells, chloroplasts and dictyosomes differed greatly in their ultrastructure from those of vegetative cells. These structural changes may be induced by changes of the physiological condition including osmotic pressure in the cells.     

Cell expansion and microfibril deposition inClosterium ehrenbergii

Cells ofClosterium ehrenbergii expanded for about 4.5 hr after vegetative cell division and about 3 hr after sexual cell division. Short cells were produced by the sexual expansion. At the early stage of the vegetative and the sexual expansion, most microfibrils were deposited transversely to the cell axis on the inner surface of the new wall. Then, bundles of microfibrils, running in various directions, overlaid the transverse microfibrils already deposited, at the late stage of the both types of expansion. The pattern of microfibril deposition changed from the transverse to the bundled pattern about 4 hr after the vegetative and about 2 hr after the sexual cell division, respectively. The change of pattern of microfibril deposition seemed to be highly correlated with cessation of cell expansion.     

Intercellular Communication During Sexual Reproduction of Closterium (Conjugatophyceae)

Closterium were reviewed. In the case of Closterium peracerosum-strigosum-littorale complex, two sex-specific pheromones and their receptors were involved in sexual reproduction. These pheromones were glycoproteins and the expression of corresponding genes was critically regulated by the sex and environmental conditions. In the case of Closterium ehrenbergii, chemotactic and sexual cell division-inducing activities for mating-type plus cells were detected and characterized. Although many processes remain to be elucidated, the present results will be helpful for understanding not only the mode of sexual reproduction in Closterium but also the variety of intercellular communication in the plant kingdom especially during sexual reproduction. Received 10 June 2000/ Accepted in revised form 6 July 2000     

TIME LAPSE ANALYSES OF SEXUAL REPRODUCTION IN CLOSTERIUM EHRENBERGII (CONJUGATOPHYCEAE)1

The process of sexual differentiation was studied using heterothallic clones of Closterium ehrenbergii Meneghini. The first visible sign of sexual reproduction was agglutination of two or more cells in a group and this was followed by gametangiogenic division and conjugation of gametangial cells. Movements of gametangial cells were carefully studied. Gametangial cells occasionally participated again in gametangiogenesis instead of proceeding directly to the formation of conjugation papilla. The whole process of sexual differentiation from vegetative cell to zygospore was considered to be basically similar in both of the two closely related mating groups, A and B, of C. ehrenbergii. Nevertheless, there were some differences between the two groups in patterns of the sexual differentiation. In Group A, vegetative cell division was completely suppressed by mixing the two complementary mating type clones together into the same medium with high light illumination. This suppression was not caused by the nitrogen depletion in the medium, but by the presence of cells of opposite mating type. In Group B, vegetative cell division and sexual reproduction occurred side by side repeatedly for several days.     

Reactivation of the Cambium in Aesculus hippocastanum L.: A Transmission Electron Microscope Study

Changes taking place during cambial reactivation in Aesculushippocastanum have been studied using transmission electronmicroscopy. Cytoplasmic activity in the form of vesicle productionby dictyosomes and endoplasmic reticulum, and coated vesicleformation at the plasmalemma, was observed in samples collectedin mid-Feb. The first cell divisions occurred 1 month later,in cells to the phloem side of the cambium, and were of twotypes: penclinal divisions producing new phloem precursors,and oblique anticlinal divisions in phloem mother cells formedat the end of the previous growing season producing putativecompanion cell/sieve element pairs. The fusiform initial wasidentified as the cell adjacent to the boundary-layer of parenchymacells and was the last cell to divide, 2 weeks after the firstdivisions in phloem precursors. For the next 4 weeks phloemcells only were produced The first new differentiating xylemelements were formed in the middle of Apr., following a surgein the rate of cell division by the initial aRd its derivativexylem mother cells. These were a mixture of developing fibresand vessel elements. Some of the boundary-layer cells were converteddirectly to vessel elements without any division taking place,while others were derived from daughter cells of the fusiforminitial produced following its reactivation. Aesculus hippocastanum L., cambium, dormancy, reactivation     

Effects of Ions and Membrane Potential on the Elongation of the Unicellular Green Alga Closterium

Cells of the unicellular green alga Closterium ehrenbergii elongatedexclusively at septa and for 4–5 hours after cell division.Cell elongation was strongly inhibited by a decrease in eitherthe external concentration of Ca²⁺ or pH, and was also inhibitedby several competitive Ca²⁺ channel blockers. Changes in concentrationsof other external ions had no effect on the elongation. Theaverage concentrations of ions in the intracellular fluid ofthe interphase cell before cell division was as follows (inmM): K⁺=56.5, Na⁺=4.8, Ca²⁺=2.4, Mg²⁺=1.3, Cl^–=59.5; thepH was 7.4. The levels of K⁺, Na⁺ and Cl^– ions decreasedsignificantly with cell elongation, suggesting that this process,which proceeds with water uptake, surpasses ion absorption.The plasma membrane potential (Vm) in both the interphase cellsand in the elongating cells was in the range of –90 to–105 mV (interior negative). The Vm was entirely determinedby the simple diffusion of K⁺. A decrease in the external concentrationof Ca²⁺ caused depolarization, probably by an indirect effectof low Ca²⁺. Changes in the extracellular level of H⁺ and othercations barely affected Vm. Thus, external Ca²⁺ and H⁺ are concludedto affect cell elongation but not via a change in the Vm acrossthe plasma membrane. (Received February 29, 1988; Accepted June 8, 1988)     

Influence of spatial heterogeneity on predation by the flatworm Mesostoma ehrenbergii (Focke) on calanoid and cyclopoid copepods

Through laboratory experiments, we analysed the influence ofspatial heterogeneity on predation by Mesostoma ehrenbergiion the calanoid Boeckella gracilis and the cyclopoid Acanthocyclopsrobustus in four horizontal and two vertical spatial arrangements.This spatial heterogeneity simulated that of Juncaceae stems,a major macrophyte in the natural environment of these zooplankton.Our results indicated that M. ehrenbergii preyed differentlyon these copepod species. The rate of predation of M. ehrenbergiion A. robustus females reached saturation in all the treatmentswith and without horizontal spatial heterogeneity, but lowerpredation rates were observed in the medium heterogeneity treatment.Predation rates of Mesostoma on B. gracilis increased with theincrease in prey abundance in the treatment without heterogeneity,while predation rates reached saturation in the treatments withhorizontal spatial heterogeneity. Mesostoma ehrenbergii consumedmales and females of B. gracilis in each of the arrangementstested. In natural habitats, interaction between extent of macrophytedevelopment and intensity of predation by M. ehrenbergii oncopepods species may be expected. We suggest that the structuralcomplexity given by macrophytes in Patagonian fishless habitatsprovide a bottleneck for M. ehrenbergii predation. This paper was presented at Plankton Symposium III, held atFiguera da Foz, Portugal between 17 and 20 March 2005, underthe auspices of the University of Coimbra and the Universityof Aveiro, and coordinated by Mário Jorge Pereira andUlisses M. Azeiteiro.     

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

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

INTRACELLULAR LOCALIZATION OF AN ENDOGENOUS CELLULOSE SYNTHASE OF MICRASTERIAS DENTICULATA (DESMIDIALES,CHLOROPHYTA) BY MEANS OF TRANSIENT GENETIC TRANSFORMATION1

The desmid Micrasterias denticulata Bréb. is useful for the study of streptophyte cell wall biology and morphology. However, no tools to analyze cell biological processes in vivo in this species are available. In the present study, transient gene expression under the control of the chl a/b–binding protein gene of the Closterium peracerosum–strigosum–littorale complex (CpCAB1) promotor was achieved for M. denticulata and illustrated by the intracellular localization of an endogenous cellulose synthase (MdCesA1). A transformation efficiency of 1/5,000 cells was achieved following microparticle bombardment. The free green fluorescent protein (GFP) signal was detected both in the nucleus and in the cytoplasm. The MdCesA1‐GFP fusion protein, on the other hand, occurred at the plasma membrane in particles concentrated at the lobe indentations, the lobe tips, and, to a lesser extent, along the lobe sides. Hence, the multipolar growth mechanism of the cell is reflected. In addition, the margins of cytoplasmic compartments, most likely dictyosomes, were labeled, in accordance with the known secretory pathway of cellulose synthase complexes. Besides intracellular localization studies, the utility of the system for overexpression phenotyping is discussed.     

Developmental Features of Cell Wall Formation in Sieve Elements of the Grass Aegilops comosa var. thessalica

In differentiating sieve elements of Aegilops comosa var. thessalicadictyosomes are abundant and they produce numerous smooth vesicles.Coated vesicles seem to bud from smooth ones. Since both kindsof vesicles appear both in the cytoplasm and in associationwith the plasmalemma, it is proposed that they move to and fusewith the plasmalemma transferring products for cell wall synthesis.During differentiation sub-plasmalemmal microtubules are initiallyscarce and randomly oriented but soon afterwards they becomenumerous and transversely oriented to the long axis. Cellulosemicrofibrils in the cell wall appear to run parallel to themicrotubules and the latter may regulate microfibril orientation. Root protophloem sieve elements develop wave-like wall thickenings,which are, during development, overlaid by microtubules perpendicularto the long axis. Just after maturation these thickenings progressivelybecome smooth and finally the walls appear uniform in thickness.The wave-like wall thickenings may function as stored wall material,utilized in later stages of development when wall material willbe needed and its synthesis will be impossible because of theabsence of a synthesizing mechanism in the highly degraded protoplastsof mature sieve elements. It is suggested that in this way thethickenings may enable root protophloem sieve elements to growand keep pace with the active clongation of the surroundingcells. Aegilops comosa var. thessalica, sieve elements. cell wall, microtubules, dictyosomes, coated vesicles, wave-like thickenings     

Re-formation of microtubules in Closterium ehrenbergii Meneghini after cold-induced depolymerization

Immunofluorescence microscopy was used to examine the re-formation of microtubules (MT), after cold-induced depolymerization, in Closterium ehrenbergii. The C. ehrenbergii cells undergo cell division followed by semicell expansion in the dark period of daily light-dark cycles. Five types of MTs, namely the MT ring, hair-like MTs around the nuclei, spindle MTs, radially arranged MTs and transverse wall MTs, appeared and disappeared sequentially during and following cell division. The wall MTs were distributed transversely only in the expanding new semicells. When cells were chilled in ice water, wall MTs in expanding cells were fragmented, and then disappeared as did the other types of MTs, within 5 min. When cells were warmed at 20°C after 2 h chilling, wall MTs and the other types of MTs re-formed. At the early stage of wall-MT re-formation in expanding cells, small, star-like MTs were formed, and then randomly oriented MTs developed in both the expanding new and the old semicells. The MT ring was also re-formed at the boundary between the new and old semicells. There were no obvious MT-organizing centers in the random arrangement. As time passed, the randomly oriented wall MTs in the old semicells disappeared and those in the expanding new semicells gradually assumed a transverse orientation. These results indicate that wall MTs can be rearranged transversely after they have been re-formed and that nucleation of wall MTs is separable from the mechanism for ordering them.Abbreviations MT(s) microtubule(s) - MTOC(s) microtubule-organizing center(s)     

Light dependency of the mating process in Closterium acerosum

Sexual cell division and activation of gametangial cells forconjugation in Closterium acerosum were induced by light. L₂₀₀cells conjugated at maximum level under the following conditions;(i) a light intensity higher than 1,000 lux in a 16-hr lightand 8-hr dark regime and (ii) an illumination time longer than12 hr at 3,000 lux. L₂₀₀ cells also conjugated under continuousillumination at 3,000 lux. The action spectrum for the activation of gametangial cellshad peaks around 450, 611 and 665 nm. 3-(4'-Chlorophenyl)-l,l-dimethylurea (CMU) inhibited the accumulationof carbohydrates and sexual cell division at 10^–5 M andthe activation of gametangial cells for conjugation at 10^–4M. (Received August 15, 1977; )     

The Ultrastructure of Tetrasporogenesis in the Marine Red Alga Chondria tenuissima (Good, et Woodw.) (Ceramiales, Rhodomelaceae)

Tetraspore development has been studied in Chondria tenuissimausing light and electron microscopy. The transformation of tetrasporangialmother cells into mature tetrasporangia involves a series ofstructural changes, especially of dictyosomes and of the nucleus.The youngest stage of tetrasporogenesis consists of a uninucleatetetraspore mother cell with synaptonemal complexes present duringearly prophase of meiosis I. Mitochondria are aggregated aroundthe nucleus, dictyosome activity is low, and proplastids occurin the peripheral cytoplasm. The cleavage furrows are initiatedalmost concomitantly with commencement of meiosis. When thecleavage furrows are initiated, spherical bodies bounded bytwo membranes are found within the cytoplasm; they develop intovacuoles with fibrillar contents (fv₁), which increase in sizeduring tetraspore development by fusing with each other andwith Golgi vesicles. The Golgi vesicles and the vacuoles withfibrillar contents (fv₁) contribute material to the developingtetraspore wall. During the middle stage of tetraspore formationthe vacuoles with fibrillar contents (fv₁) are dominant, dictyosomeactivity increases, as well as the number of plastids and mitochondria;starch formation also increases. Stacked cisternae of the endoplasmicreticulum are found within the peripheral part of the nucleus.The same nuclear structures are also observed in tetrasporangiaof the marine red alga Gastroclonium clavalum. The final stageis characterized by the disappearance of vacuoles with fibrillarcontents (fv₁) and of the stacked ER within the nucleus, presenceof straight, large dictyosomes which produce cored vesicles,an abundance of starch grains and by the formation of fullydeveloped chlorqplasts. The cored vesicles contain Thiéry-positivematerial and contribute to the formation of vacuoles with fibrouscontents (fv₂) as they are dominant in the tetraspores beforetheir liberation. Rhodophlyla, Chondria, tetrasporogenesis, ultrastructure, Golgi apparatus     

Changes in Ultrastructure and Abscisic Acid Level, and Response to Applied Gibberellins inTaxus maireiSeeds Treated With Warm and Cold Stratification

Seeds ofTaxus maireiare known for their deep dormancy whichcan only be broken by a procedure involving warm stratificationfollowed by cold stratification. Treatments with alternatingtemperatures of 25/15 or 23/11 °C (12 h light) for 6 monthsfollowed by 5 °C for 3 months were successful in overcomingseed dormancy. After 6 months of warm stratification, cytologicalchanges observed included: enlargement of the embryo; a decreasein the number of lipid bodies; appearance of ER; and increasesin mitochondria, plastids, dictyosomes, vacuoles and microbodiesin the shoot apical meristem. Cold stratification followingthe warm treatment induced cell division, and one or two distinctnucleoli in the shoot apical meristem cells were observed. Bothwarm and cold stratification reduced endogenous ABA concentrationsfrom the original 8888 pg per freshly harvested seed to 392and 536 pg, respectively. Treatment with exogenous gibberellinsafter seeds had been warm-stratified showed that GA₄and GA₇wereeffective at promoting seed germination, but GA₃was not. Theseresults suggest that the strong seed dormancy ofT. maireicouldbe caused by a high ABA content and underdevelopment of theembryos in freshly shed seeds. We conclude that warm stratificationwith alternating temperatures increases the growth of embryosby cell expansion and enlargement and decreases ABA content,but seeds still remain ungerminated. Cold stratification mayinduce the response to GAs and initiate cell division resultingin release from physiological dormancy and subsequent germinationofT. maireiseeds.Copyright 1998 Annals of Botany Company Taxus mairei; ultrastructure; abscisic acid; gibberellin; seed dormancy; stratification; germination.     

Short-term Salinity and High Temperature Stress-associated Ultrastructural Alterations in Young Leaf Cells ofOryza sativaL.

Salinity and high temperature stresses adversely affect growthand development of rice plants. To investigate the responseof rice cells to these stresses, we have analysed short-termstress-induced subcellular alterations in undifferentiated leafcells of rice seedlings by transmission electron microscopy.Perturbations noted particularly with respect to plasma membrane,mitochondrial membranes, endoplasmic reticulum, polyribosomesand dictyosomes are highlighted. The subcellular changes evokedby both stresses after 4 h were lysis of the cytoplasm, accumulationof electron-dense granules in the cytoplasm, distension in theER membranes, enhanced association of ribosomes with the endoplasmicreticulum, reduction in the number of mitochondrial cristae,as well as disorganization of cell wall fibrillar material.Certain changes were found to be unique to either the salinityor high temperature stress. Plasmolysis and increased cytoplasmicvesiculation were seen only in response to salinity stress,while discontinuity in the plasma membrane with close associationof the osmiophilic granules were observed only in response tohigh temperature.Copyright 1997 Annals of Botany Company Electron dense granules; high temperature stress; leaf cells; Oryza sativaL.; rice; salinity; ultrastructure     

The formation and consumption of lipid droplets in the zygote and germinated cells ofClosterium ehrenbergii

The formation and consumption of lipid droplets was observed with an electron microscope in the zygote and the germinated cells of the green alga,Closterium ehrenbergii. The lipid droplets were formed in lysosomal vesicles during zygote maturation following conjugation. In the germinated cells, they were enclosed in ERs and gradually consumed in them. This consumption occurred in the cells at the early stages of expansion. The derivative substances may possibly be used for cell surface expansion.     

Differentiation of the Tapetum During Microsporogenesis in Tomato (Lycopersicon esculentum Mill.), with Special Reference to the Tapetal Cell Wall

During microsporogenesis and pollen maturation, the tapetumin anthers of tomato (Lycopersicon esculentum) underwent severalultrastructural changes and ultimately degenerated. The changesobserved related to the secretory function of the tapetum andto the transfer of materials from the cytoplasm to the surfaceof tapetal cells. Electron dense deposits, initially in thevacuoles, disappeared coincident with the appearance of orbiculeson the cell wall. The fibrillar wall of the tapetal cells loosened,presumably to facilitate transfer of materials through the wall.In Addition, membranous fragments were a consistent featurein the tapetum wall and may play a role in transport of materials.The cells of the inner tapetum (towards the connective) andouter tapetum (towards the epidermis) had different ultrastructuralfeatures. The cytoplasm of the outer tapetum was more electrondense and had a higher proportion of dictyosomes and mitochondriathan the inner tapetum, indicating the greater secretory natureof the outer tapetum. The plastids and mitochondria also differedin morphology between the two regions. Degenerations of thetapetal cytoplasm began by the vacuolate microspore stage. Atanthesis, cytoplasm was absent but the orbicular wall of thetapetum remained appressed to the wall of the middle layer ofthe anther.Copyright 1993, 1999 Academic Press Lycopersicon esculentum, microsporogenesis, pollen development, tapetum development, tomato, ultrastructure