The microtubule cytoskeleton in developing cysts of the green algaAcetabularia: Involvement in cell wall differentiation

Summary Cysts of the green algaAcetabularia develop a unique lid structure to enable the release of gametes. This lid is separated from the rest of the thick cellulose cell wall by a circular fault line formed within the fibrillar texture of the wall. By immunofluorescence microscopy, we show that, prior to the first division of the single cyst nucleus, the radially symmetrical, perinuclear microtubule system which is a remnant carried over from previous developmental stages of cyst morphogenesis transforms into a circular microtubule band (CMB) around the nucleus. This band consisting of only a few bundled microtubules beneath the plasma membrane encircles the cyst nucleus at a distance of 75 to 100m. In a previous fine structural study, a lid-forming apparatus (LFA) was described as a circular band of rod-like structures in the plane of the plasma membrane, demarcating the contour of the future lid. Both the CMB and the LFA are superimposed on the rim of the lid. We therefore propose that the microtubule band is a component of the LFA identical with the rod-like structures. Formation of the CMB and, hence, lid formation are blocked by the microtubule-specific herbicide Oryzalin but not by the actin filament-disrupting inhibitor cytochalasin D. Upon recovery from Oryzalin treatment, the nuclei but not the prospective sites of the CMBs serve as nucleation centers, indicating that the CMB is not formed by a pre-existing template in the plasma membrane. This suggests that the dynamic behavior of the microtubules within the perinuclear microtubule cytoskeleton gives rise to the CMB. Since the stage of CMB assembly marks the beginning of cell wall formation, it is proposed that the CMB determines the position of the lid by spatially controlling cell wall deposition. On the basis of current hypotheses, two scenarios for the role of the LFA/CMB in lid formation are discussed.Abbreviations CMB circular microtubule band - EGTA ethylene glycol bis-(-aminoethyl ether) N,N,N,N-tetraacetic acid - FITC fluorescein isothiocyanate - LFA lid-forming-apparatus - MAP microtubule-associated protein - MT microtubule - MTOC microtubuleorganizing center Dedicated to the memory of Professor Oswald Kiermayer     

Ultrastructure of mitosis inAmoeba proteus

Summary The fine structure ofAmoeba proteus nuclei has been studied during interphase and mitosis. The interphase nucleus is discoidal, the nuclear envelope is provided with a honeycomb layer on the inside. There are numerous nucleoli at the periphery and many chromatin filaments and nuclear helices in the central part of nucleus.In prophase the nucleus becomes spherical, the numerous chromosomes are condensed, and the number of nucleoli decreases. The mitotic apparatus forms inside the nucleus in form of an acentric spindle. In metaphase the nuclear envelope loses its pore complexes and transforms into a system of rough endoplasmic reticulum cisternae (ERC) which separates the mitotic apparatus from the surrounding cytoplasm; the nucleoli and the honeycomb layer disappear completely. In anaphase the half-spindles become conical, and the system of ERC around the mitotic spindle persists. Electron dense material (possibly microtubule organizing centers—MTOCs) appears at the spindle pole regions during this stage. The spindle includes kinetochore microtubules attached to the chromosomes, and non-kinetochore ones which pierce the anaphase plate. In telophase the spindle disappears, the chromosomes decondense, and the nuclear envelope becomes reconstructed from the ERC. At this stage, nucleoli can already be revealed with the light microscope by silver staining; they are visible in ultrathin sections as numerous electron dense bodies at the periphery of the nucleus.The mitotic chromosomes consist of 10 nm fibers and have threelayered kinetochores. Single nuclear helices still occur at early stages of mitosis in the spindle region.     

The Fine Structure of Acanthamoeba astronyxis,with Special Emphasis on Encystment1

ABSTRACT. The fine structure of the trophozoite, encysting cells, and the cyst of Acanthamoeba astronyxis has been examined. In the trophic form a microtubule organizing center was associated with a well developed Golgi complex. During encystment the organelles of the amoeba changed considerably. The profiles of rough endoplasmic reticulum elongated and were often arranged in circles of multilayered concentric systems, enclosing mitochondria, the nucleus, or other inclusions. The mitochondria showed a tendency toward elongation and constriction. One or two nucleolus-like bodies appeared in the nucleus. Lipid droplets increased considerably in amount and were distributed individually or as aggregates. The mature cyst was star-shaped and surrounded by an almost circular exocyst and an endocyst that was closely apposed to the cell membrane. Both walls differed in their thickness and granulation. The exocyst was continuous over the entire cyst, while the endocyst was interrupted by gaps, ostioles. in the region of the rays. Within the ostioles was a bell-shaped structure, the operculum. The latter was composed of a granular material comparable in electron density to that of the endocyst.     

Multinucleate cyst formation in Acetabularia mediterranea after x-irradiation

Cells of Acetabularia mediterranea were irradiated with increasing doses of X-rays (64.5–258·10^-4 kC kg^-1). The cells are radioresistant up to 193.5·10^-4 kC kg^-1 in terms of growth and progression through he life cycle but the morphogenesis of whorls, caps, and cysts is accompanied by morphological alterations. Microscopical examination of cyst bearing caps in irradiated cells has shown the presence of giant cysts neighboring particularly small ones. Photographic recording of cyst development showed that the multinucleate cap cytoplasm partitions into multinucleate portions rather than uninucleate ones as in the control cells. After complete cleavage a cyst wall is deposited onto the multinucleate cytoplasm. In contrast to uninucleate cysts with one lid the wall contains multiple lids. Their number appears to correspond to the number of nuclei in the cytoplasm compartment during cleavage. The results indicate that X-rays preferentially inhibit the synthesis of a factor which plays a role in establishing the normal spatial morphogenetic pattern necessary for cyst formation.     

Characterization of zoospore and cyst surface structure in saprophytic and fish pathogenicSaprolegnia species (oomycete fungal protists)

Summary The importance of the surface structure and chemistry in zoospores and cysts of oomycetes is briefly reviewed and the organelle systems associated with encystment described. The surface structure and chemistry of primary and secondary zoospores and cysts ofSaprolegnia diclina (a representative saprophytic species) andS. parasitica (a representative salmonid fish pathogen) were explored using the lectins concanavilin A (Con A) and wheat germ agglutinin (WGA) and monoclonal antibodies (MAbs) raised against a mixed zoospore and cyst suspension ofS. parasitica. The binding of lectins and antibodies to spores was determined using immunofluorescence microscopy with fluorescein isothiocyanate-labelled probes and with electron microscopy with gold-conjugated probes applied to spore suspensions post-fixation. In both species Con A, which is specific for glucose and mannose sugars, bound to both the surface of primary and secondary zoospores (the surface glycocalyx) and their cyst coats and readily induced zoospore encystment. The binding to the cysts appeared to be mainly associated with the matrix material released from the primary and secondary encystment vesicles and which appeared to diminish with time. No binding to germ tube walls was observed with this lectin. The MAb labelling showed a generally similar binding pattern to the primary and secondary cysts to that observed with Con A, although the binding to zoospores was more variable. Primary zoospores bound the antibodies but secondary zoospores appeared less reactive. It is suggested that the MAbs share a common epitope with one or more of the Con A-binding components. In both species WGA, which is specific for amongst other things the sugar N-acetyl glucosamine, bound to localised apical patches on the primary zoospores. This lectin also binds to the ventral groove region of secondary zoospores ofS. diclina, which were induced to encyst by this lectin. In contrast secondary zoospores ofS. parasitica were not induced to encyst by the addition of WGA and showed a patchy dorsal binding with this lectin. WGA also binds to both the inner wall of discharged primary cysts and the young germ tube walls of both species. These observations are discussed both in relation to other oomycete spores and to their possible functional and ecological significance.Abbreviations BSA bovine serum albumin - Con A Concanavalin A - DBA Dolichos biflorus agglutinin - ELISA enzyme-linked immunosorbent assay - EM electron microscope - EV encystment vesicles - FCS foetal calf serum - FITC Fluorescein isothiocyanate - FV peripheral fibrillar vesicles - G+F 0.2% glutaraldehyde and 2.0% formaldehyde primary fixative solution - 2G 2% glutaraldehyde primary fixative - LM light microscopy - MAbs monoclonal antibodies - LPV large peripheral vesicles - PBS phosphate buffered saline - PCV flattened peripheral cisternae - PEV primary encystment vesicle - PIPES piperazine-N,N1-bis(2-ethane sulfonic acid) - PNA Ricinus communis agglutinin - RAM-FITC/Au_10–20 Fluorescein isothiocyanate/gold (10 or 20 nm) labelled rabbit anti-mouse immunoglobulin - RCA Ricinus communis agglutinin - SEM scanning electron micrograph - SBA soybean agglutinin - SEV secondary encystment vesicles - TEM transmission electron micrograph - UEA I Ulex europaeus agglutinin - WGA wheat germ agglutinin     

The role of kinetosome-associated organelles in the attachment of encysting secondary zoospores ofSaprolegnia ferax to substrates

Summary Electron and fluorescence microscopy were used to identify organelles involved in attachment of secondary zoospores ofSaprolegnia ferax as they were transformed into secondary cysts. When secondary zoospores were exposed to 1.0% peptone in the absence or presence of a substrate, they began to encyst. If substrates were present when encystment was induced, the groove surface of the secondary zoospores adhered to them. The first event in attachment was secretion of contents of the kinetosome-associated organelle (K-body), which was typically oriented with the tubule-filled cavity positioned toward the cell surface of the groove region in the zoospore. The tubules which contained carbohydrates became coarsely granular, the matrix became more fibrous, and the shell remained along the membrane concavity that was formed as the K-body fused with the plasma membrane.Five minutes later, a cyst coat appeared, and cysts were not readily dislodged from a substrate. The concavity was no longer found, presumably because it had evaginated; but a layered pad of adhesion material was between the cyst coat and substrate. The layers of the adhesion pad corresponded to the structure of the matrix of K-bodies. As with the tubules of the K-body, the coarsely granular portion at the edge of the pad stained for carbohydrates. Similarly, the lectins WGA and GS-II labeled with fluorescein stained the rim of the adhesion pad on cysts, indicating the presence of glycoconjugates containing N-acetylglucosamines. Because globular areas near the kinetosomes and groove of zoospores (where K-bodies were located) also bound WGA and GS-II, K-bodies contained the same carbohydrates as the adhesion pad. We conclude that K-bodies function in the attachment of encysting zoospores to substrates as the cell differentiates. The tubular portion of the K-body matrix contains carbohydrates which might assist in the adhesion process.Abbreviations D dictyosome - EV encystment vesicle - F flagellum - C cyst - CC cyst coat - Con A concanavalin A - GS-II Griffonia simplicifolia lectin - K K-body - Kt kinetosome - M mitochondria - N nucleus - NB nuclear beak - PC peripheral cisterna - PV peripheral vesicle - S shell region of K-body matrix - SBA soybean agglutinin - R 3 anteriorly directed triplet rootlet - R 8 posteriorly directed octet rootlet - WEV water expulsion vacuole - WGA wheat germ agglutinin     

Light and electron microscopical studies onHafniomonas gen. nov. (Chlorophyceae,Volvocales), a genus resemblingPyramimonas (Prasinophyceae)

Based on light and electron microscopical studies ofPyramimonas reticulata the genusPyramimonas is shown to contain a number of unrelated flagellates.P. reticulata andP. montana are transferred to the new genusHafniomonas, cells of which differ fromPyramimonas in shape, in the absence of scales and hairs on the body and flagellar surfaces, in details of the chloroplast, the position of the nucleus, the Golgi apparatus, the internal structure of the flagellar apparatus, and in cell division. The prasinophytePyramimonas contains a characteristic association of a large microbody and a rhizoplast, situated on the nuclear surface. A similar association is being found in an increasing number of prasinophycean flagellates, but is absent inHafniomonas, which is considered related to chlorophycean rather than prasinophycean flagellates. The phylogenetic position ofHafniomonas is discussed, based in particular on details of the unique flagellar apparatus.     

Evolution and development of polarized germ cell cysts: new insights from a polychaete worm, Ophryotrocha labronica

Polarized oogenic cysts are clonal syncytia of germ cells in which some of the sister cells (cystocytes) differentiate not as oocytes, but instead as nurse cells: polyploid cells that support oocyte development. The intricate machinery required to establish and maintain divergent cell fates within a syncytium, and the importance of associated oocyte patterning for subsequent embryonic development, have made polarized cysts valuable subjects of study in developmental and cell biology. Nurse cell/oocyte specification is best understood in insects, particularly Drosophila melanogaster. However, polarized cysts have evolved independently in several other animal phyla. We describe the differentiation of female cystocytes in an annelid worm, the polychaete Ophryotrocha labronica. These worms are remarkable for their elegantly simple cysts, which comprise a single oocyte and nurse cell, making them an appealing complement to insects as subjects of study. To elucidate the process of cystocyte differentiation in O. labronica, we have constructed digital 3D models from electron micrographs of serially sectioned ovarian tissue. These models show that 2-cell cysts arise by fragmentation of larger “parental” cysts, rather than as independent units. The parental cysts vary in size and organization, are produced by asynchronous, indeterminate mitotic divisions of progenitor cystoblasts, and lack fusome-like organizing organelles. All of these characteristics represent key cytological differences from “typical” cyst development in insects like D. melanogaster. In light of such differences and the plasticity of female cyst structure among other animals, we suggest that it is time to reassess common views on the conservation of oogenic cysts and the importance of cysts in animal oogenesis generally.     

Structural differences between the cyst walls of Streptocephalus dichotomus (Baird), S. torvicornis (Waga), and Thamnocephalus platyurus (Packard) (Crustacea: Anostraca), and a comparison with other genera and species

The surface topography and layering of the cyst wall of the fairy shrimps Streptocephalus dichotomus, S. torvicornis and Thamnocephalus platyurus are described, based on scanning electron micrographs. Cyst walls are not invariably bilayered, but may be composed of up to four layers. They are species-specific in all taxa so far studied. Cyst wall structure within single cysts may differ according to the topographical location on the cyst.     

Fine structure of spermatozoa and spermatogenesis ofSchizomus palaciosi,Reddell and Cokendolpher, 1986 (Arachnida: Uropygi,Schizomida)

Summary Spermatogenesis ofSchizomus palaciosi occurs in cysts in paired tubular testes located ventrally in the opisthosoma. Only few germ cells comprise one cyst. In early spermiogenesis an acrosomal complex composed of a spherical vacuole and a short acrosomal filament is established opposite of which a 9×2+3 flagellum emerges from a flagellar tunnel. The latter, however, is only a short-lasting structure. A manchette of microtubules surrounds nucleus and part of the acrosomal vacuole. The alterations in the arrangement of the microtubules during spermiogenesis are described. The spermatid finally is an elongate cell with a slender acrosomal vacuole on top of the helical nucleus. A deep implantation fossa filled with dense material is encountered. The acrosomal vacuole is accompanied by an intricate paracrosomal lattice structure not known at present of otherArachnida. This structure disappears during final spermiogenesis. The acrosomal filament (perforatorium) reveals filamentous subunits arranged in a regular pattern. Large ovoid mitochondria do not establish a distinct middle piece. Finally the elongate spermatid is coiled to form the mature spherical spermatozoon.The results are discussed under functional and taxonomical aspects.     

High-resolution immunogold localization of Giardia cyst wall antigens using field emission SEM with secondary and backscatter electron imaging

We describe here the ultrastructural localization of Giardia cyst antigens in the filaments associated with the outer portion of intact cysts and on developing cyst wall filaments in encysting trophozoites. Post-embedding immunogold labeling of thin sections of intact Giardia cysts with polyclonal and monoclonal antibodies specific for cyst wall antigens (major protein bands of approximately 29, 75, 88, and 102 KD on Western blots) showed strong labeling of the filamentous cyst wall, whereas no labeling was seen on the membranous portion. High-resolution field emission scanning electron microscopy (FESEM) of Giardia cysts revealed that the cyst wall-specific polyclonal rabbit antisera and monoclonal mouse antibody produced gold labeling of 20-nm filaments in the cyst wall as detected with secondary electron imaging (SEI) and backscatter electron imaging (BEI) at 10 kV, despite coating of the cells with platinum by ion sputtering. FESEM studies of encysting Giardia trophozoites demonstrated that immunostaining with antibodies to cyst wall antigens produced colloidal gold labeling of developing cyst wall filaments on the cell surface; however, the intervening membrane domains were unlabeled. Substitution of normal serum for cyst wall-specific antibodies, or preabsorption of specific antibodies with Giardia cysts, eliminated immunolabeling of the filaments.     

Ultrastructural characterization of asexual cysts ofGonyaulax polyedra Stein (Dinoflagellata)

Summary In the bioluminescent dinoflagellateGonyaulax polyedra, the formation of asexual cysts was elicited either by addition of 5-methoxytryptamine or by transfer to short-day conditions under lower temperature and decreased light intensity. The resulting changes were followed in vivo by light microscopy, and analysed ultrastructurally by electron microscopy. Irrespective of the method of cyst induction, theca and flagella shedding and the formation of a cyst wall can always be observed as essential steps in this process. Despite the extremely low level of bioluminescence emitted from the cysts, some scintillons persist. Encystment is accompanied by organelle and substructure rearrangement. Although cysts induced by 5-methoxytryptamine or by short days closely resemble each other, electron microscopy reveals typical differences. In cysts obtained by treatment with 5-methoxytryptamine most chloroplasts are of the expanded type, extending to the central region, whereas only a few are compact and peripherally positioned. Cysts induced by shortdays predominantly contain chloroplasts of the compact type and contain large amounts of stored starch and lipids. Their ultrastructural organization therefore resembles that of mastigote cells during darkness.     

Encystment Processes and the “Matrioshka-like Stage” in a Moss-dwelling and in a Limnic Species of Eutardigrades (Tardigrada)

Tardigrades have two forms of dormancy, namely cryptobiosis and encystment. The encystment is a form of diapause known for a limited number of species of tardigrades and still little studied. To increase the knowledge on encystment, two species of eutardigrades from Italy have been considered: the moss-dwelling Amphibolus volubilis (Eohypsibiidae), and the limnic Dactylobiotus parthenogeneticus (Murrayidae). Cysts have been collected in nature, or induced under laboratory conditions. In the latter case, it was possible to follow the several steps of encystment processes. Two different types of cyst (“type 1” and “type 2”) have been found in A. volubilis, while in D. parthenogeneticus only one type has been found. In general, the ovoid-shaped cysts are constituted by a series of cuticles surrounding the animals and resemble an onion or a Matrioshka Russian doll. In all three types of cyst, the encystment processes show both common and peculiar traits. Encystment begins with the discharging of the sclerified parts of the buccal-pharyngeal apparatus, as in the molting process, but without the loss of the old cuticle. Then, two or three new cuticles are serially synthesized, according to cyst type. In A. volubilis, the ultrastructure of these new cuticular involucra is similar to that of non-encysted animal cuticles, while in D. parthenogeneticus the ultrastructure of the new cuticular involucra differs from that of non-encysted animal cuticle. A modified buccal-pharyngeal apparatus has been observed both in “type 2” cyst of A. volubilis and in the D. parthenogeneticus cyst.     

Freeze-fracture and cytochemical studies on the in vitro cyst form of reptilian Blastocystis pythoni

After cultured cysts are osmotically shocked by treating with distilled water, there is an exponential increase in the cyst form of Blastocystis pythoni; this was demonstrated by an immunofluorescence antibody assay against the culture organisms. In 11-day-old cultures of B. pythoni, 68.8% of the organisms (= 2.2 x 10(8) cysts/ml) were in the cyst form. Examination of thin sections of cysts revealed many similarities to the cyst forms of Blastocystis obtained from fecal samples in previous investigations. Freeze-fracture images of the plasma membrane of non-cyst cells also revealed a similar distribution of the intramembrane particles (IMP) when compared to non-cysts of B. hominis, while the plasma membrane of the cyst form showed practically no IMP. The size and morphology of particle-rich small depressions and smooth small protrusions observed on the P face and E face of non-cyst cells, respectively, were similar to endocytic sites reported for B. hominis. In the present study glycogen was cytochemically demonstrated at the ultrastructural level by an alkaline bismuth staining method in both cyst and non-cyst cells.     

The effect of penicillin on the encystment of Bdellovibrio sp. strain W

When penicillin, and other inhibitors of peptidoglycan synthesis were added to encysting cultures of Bdellovibrio strain W, the encysting process continued, resulting in the production of cysts which were spherical in shape. Transmission electron micrographs of these spherical bdellocysts revealed the absence of an outer cyst wall. These cysts, devoid of cyst wall, were capable of germination under appropriate condition with the emergence from the prey ghost of highly motile spheroplasts. Withdrawl of the antibiotics after encystment had begun led to the production of spherical cysts that were surrounded by an outer cyst wall.     

Spermatogenesis in the inseminating African butterflyfish Pantodon buchholzi (Teleostei: Osteoglossiformes: Pantodontidae) with the revision of residual bodies formation

The aim of this study was to analyse spermatogenesis in the African butterflyfish, Pantodon buchholzi, using transmission electron microscopy and scanning electron microscopy. P. buchholzi is the most basal teleost that exhibits insemination and produces a highly complex introsperm with the most elongate midpiece known in teleost fishes. Their early stages (spermatogonia and spermatocytes) do not differ greatly from those of other fishes, with the exception of Golgi apparatus degradation appearing as spindle-shaped bodies (SSBs). In round, early spermatids, the development of the flagellum begins after the migration of the centriolar complex towards the nucleus. Later, the elongation of the midpiece coincides with the displacement of the mitochondria and their fusion to produce nine mitochondrial derivatives (MDs). In these spermatids, the nucleus is situated laterally to the midpiece, with condensing chromatin in the centre of the nucleus. Within the midpiece, the flagellum is located within a cytoplasmic canal and is surrounded by a cytoplasmic sleeve containing fibres, MDs and a great amount of cytoplasm located on one side. During the next phase, nuclear rotation, the highly condensed chromatin is displaced to a position above the centriolar apparatus, whereas chromatin-free nucleoplasm is transferred to the cytoplasm. Later, this nucleoplasm, still surrounded by the nuclear membrane, is eliminated into the cyst lumen as the nucleoplasmic packet. Within the highly elongate spermatids, other excess organelles (SSBs, endoplasmic reticulum and mitochondria) are eliminated as residual bodies (RBs). Fully developed spermatozoa, which contain conical-shaped nuclei, eventually coalesce to form unencapsulated sperm packets (spermatozeugmata) that are surrounded by RBs at the level of the extremely elongate midpieces. Later, RBs are removed at the periphery of the cyst by means of phagocytosis by Sertoli cells.     

Wall formation in the saprolegniales

Summary The primary and secondary cysts of Saprolegnia ferax and the secondary cysts of Dictyuchus sterile have a two layered wall structure, the outer layer of which bears various types of spines. These spines, and the outer wall layer are derived from preformed structures (bars) found in the cytoplasm prior to encystment. Golgi derived vesicles appear to contribute to the inner layer of the primary cyst wall of S. ferax. The outer surface of the secondary cyst walls of this species has fibrils which are not embedded in matrix material.     

The chrysophycean paleocyst flora of the bottom sediments of Kortowskie Lake (Poland) and its ecological significance

The postglacial sediments of Kortowskie Lake contain well preserved chrysophycean cysts. Forty-six morphotypes were identified by scanning electron microscopy (SEM) and light microscopy (LM). Ten new morphotypes of fossil chrysophycean cysts are described. Some of the most abundant taxa show clear trends associated with the increase in lake trophic status. The cyst flora is divided into four groups: eutrophic forms (Cysta globata, C. cortoviensis, C. vermicularis); oligotrophic forms (Cysta curvicollis); cool-water forms (Cysta carinifera, C. crassicollis, C. microspinosa, C. modica and Cysta stellata), and indifferent forms (all other morphotypes which did not show a clear tendency). The percentage abundance of chrysophycean cysts wa related to climate changes, showing a considerable increase during the Subboreal period. Two factors, lake fertility and water level changes affected the abundance of the fossil cyst flora in the sediments of Kortowskie Lake.     

Autonomous circular and radial motions of the nucleus inPleurenterium tumidum and their relation to cytoskeletal elements and the plasma membrane

Summary InPleurenterium tumidum the nucleus leaves its central position at the end of cell development and moves centrifugally towards the cortical cytoplasm of the isthmus area. It passes between the chloroplast lobes and starts to perform circular motions along the cell wall ring of the isthmus independently from other cell organelles and cytoplasmic streaming. This autonomous nuclear motion is a unique phenomenon in plant cells which is reported here for the first time. One turn of the nucleus which may occur either clockwise or counter-clockwise lasts an average of 60 minutes. The velocity of circular nuclear motion lies between 0.03 and 0.08 m per second and increases with increasing number of nuclear turns. The nucleus undergoes at least 12 but sometimes up to 70 turns and may change its direction of motion several times. When circular nuclear motion is finished the nucleus migrates centripetally towards the cell center where the next mitosis takes place.Ultrastructural studies demonstrate that a distinct arrangement of the plasma membrane forming a ring-shaped fold together with the adjacent isthmus system of microtubules (IS) serves as a hoop-like track for the nucleus during the stage of circular motion. The nucleus moves along this track by surrounding it in a deep furrow which develops parallel to its longitudinal axis at its cell wall facing side. The spatial arrangement of the plasma membrane fold and the nuclear furrow are only present during the stage of circular nuclear motion. No actin filaments seem to be involved in the nuclear circulations since the nucleus continues its circular motions after cytochalasin B (CB) treatment even at concentrations which arrest cytoplasmic streaming. Amiprophos-methyl (APM) leads to an inhibition of circular nuclear motion which resumes when the APM solution is removed. Microtubules appear to be primarily responsible also for both the radial nuclear motions as well as the anchoring of the nucleus in its central position. The meaning of circular and radial nuclear motions for thePleurenterium cell is not yet clear, a relation between the nuclear behavior and the inner cell architecture is discussed and compared to that of other desmids.     

The mitotic apparatus during unequal microspore division observed by a confocal laser scanning microscope

Summary The structure of the mitotic apparatus during the microspore division ofTradescantia paludosa, which has a distinctively unequal division of large vegetative and small generative cells, was studied using -tubulin immunofluorescence methods and confocal laser scanning microscopy. Mitotic apparatuses began to develop asynchronously during early prophase at the vegetative pole (VP) and during prometaphase at the generative pole (GP). Both, however, reached completion together at the same time during metaphase. At the VP from prophase to prometaphase, microtubules (MTs) did not converge on the pole, and there was a circular area containing only a few MTs. The prophase spindles on the VP side were in the form of domes or cones that lacked the top. In the metaphase, however, the MTs concentrated at the pole to form a representative cone-shaped half-spindle. At the GP from prometaphase to metaphase, the MTs did not concentrate, and a circular area existed that lacked MTs. The half-spindles formed truncated cones. When the phragmoplast developed and curved around the generative nucleus during the telophase. it first grew toward the long axis of the ellipsoidal-shaped microspore; and after it arrived at the inner membrane of the microspore, it again curved past the generative nucleus toward the short axis. In conclusion, it was found that the mitotic apparatus ofT. paludosa microspores with its asynchronous growth and asymmetrical spindle structure and with its three dimensional growth of phragmoplasts had a peculiar developmental manner related to unequal division.