The fiber cells of the middle layer of Trichoplax adhaerens are interconnected by slender extensions. Newly formed connections after mechanical disruption of the tissue studied in ultrathin sections revealed cytoplasmic continuity between the cell bodies, suggesting a syncytial organisation of the fiber cell layer. The slender extensions connecting the cell bodies are traversed by microtubules and microfilaments. The structure of rare osmiophilic cell contacts suggests a stage in the fusion of adjacent cell membranes.
In the heterotrichous ciliate Nyctotherus ovalis, a meshwork of 10 nm filaments present in the cell cortex surrounds cytostome and cytoproct and is connected by thin strands to a perinuclear coat. Like intermediate filaments (If) of higher organisms the filaments are insoluble in non-ionic detergent, and paracrystals can be formed after dissolution in 8 M urea. The cross-reactions of these filaments with monoclonal anti-vimentin and anti-cytokeratin antibodies suggest that they share common epitopes with both types of If proteins.
The fate of intracellular membranes stained by the osmium ferricyanide (OsFeCN) procedure was followed from premeiotic interphase to interkinesis inDysdercus intermedius. During diakinesis the centrioles forming primary cilia attach temporarily with their proximal ends to the nuclear envelope which is stretched from pole to pole. Breakdown of the nuclear envelope is preceded by deep indentations with microtubules from growing asters. Vesicles of smooth endoplasmic reticulum which accumulate gradually in the course of prophase contribute to the ensheathment of the chromosomes with membranes. When the nuclear envelope breaks down, the polar parts of the formerly perinuclear membranes follow the ingrowth of the spindle microtubules towards the cell equator where the seven bivalents are arranged in a circle with the X1X2 sex chromosomes in the centre. The metaphase I spindle thus contains longitudinally oriented membranes between the poles, membranous envelopes around all chromosomes and radial connections from the autosomes to the sex chromosomes in the centre. At anaphase the homologues leave their common sheath and a microtubular stembody surrounded by membranes appears between the receding dyads. In the interkinetic nucleus the gonosomes are separated from the autosomes by a common membranous sheath which may be instrumental in their joint assignment to only one pole in the second meiotic division. Calcium sequestering sites visualized by oxalate precipitation are the Golgi lamellae and vesicles derived from them that surround the whole spindle body.
ANosema-like microsporidian with transovarian passage was found in most organs, including muscle and nervous tissue, of newly hatched and adult wasps. The mitotic apparatus of the dikaryotic meronts includes spindle-pole bodies with intra- and extranuclear components and small kinetochores with single microtubules. Cytokinesis in stages with four nuclei involves budding. The spore wall reacts with Calcofluor White, a fluorochrome specific for β-1,4 glycosidic linkages such as those found in chitin.
Abstract Flageilated swarmers representing an alternative mode of asexual reproduction are budded off as hollow spheres at the dorsal surface. They are covered by cells of the dorsal epithelium and the interior cavity is lined by ventral epithelium. Fiber cells are in the interspace. Early primordia in the interspace between both epithelia of the mother animal are cavities filled with granular material and surrounded by non-flagellated epithelium cells. No mitoses are seen during growth of the bud whose interior becomes densely flagellated as is characteristic for the ventral epithelium. Fiber cells are passively included. The bud at first forms a bulge at the dorsal surface which is then pinched off. The flattened shape of Trichoplax is attained via cup-shaped and C-shaped intermediate stages after opening of the hollow sphere.
ABSTRACTAfter a thin membranous envelope surrounding the cell body and cilia of Colpoda steinii has been formed, the main mass of the proteinaceous cyst wall is deposited without exocytosis. It can be composed of two layers, the denser and wrinkled ectocyst and the smooth‐walled endocyst; however, the ectocyst may be missing. Evidence is presented that ecto‐ and endocyst are formed from vesicles derived from abundant rough endoplasmic reticulum which appears at the time of wall formation. The cilia are retained and become embedded in the peripheral cytoplasm. Synthesis of RNA and protein is required as actinomycin C and cycloheximide block cyst formation. Calcium is required during a sensitive phase prior to encystment.
The fate of cytoplasmic membranes has been traced from spermatogonial interphase to the telophase of the second meiotic division with the aid of osmium ferrocyanide staining. At prophase, the nuclear membrane is indented to a funnel shape due to microtubules which radiate from the poles. These indentations open at prometaphase admitting microtubules to the nuclear interior. Since the nuclear envelope widens at prometaphase with only few lateral defects, organelles are excluded and the spindle is essentially intranuclear. Membranes surround the spindle until late anaphase. There are no cytoplasmic vesicles within the spindle, but there is an abundance of vesicles around the spindle poles following the radiating microtubules. At telophase, small vesicles around the chromosomes contribute to the new nuclear envelope while others fuse into large blisters which disappear gradually. A new vesicle system is formed in the daughter cells. At meiosis, the nuclear membrane is not indented by the polar microtubules which follow the contour of the nucleus together with vesicles. At late diakinesis, the bivalents are individually ensheathed by vesicles and lamellae which separate them from the remainder of the nucleus. In building up the metaphase configuration, the chromosomal sheaths become interconnected with the X1/X2-pseudotetrad in the center. Lamellae from these sheaths extend to the poles following the kinetochore microtubule bundles. In contrast to mitosis, the spindle body is pervaded by a structural framework of membranes with mitochondria in between. Essentially the same spindle architecture exists in the second meiotic division with the exception of the tandemly joined X1 and X2 chromosomes which are ensheathed by endoplasmic reticulum but lie outside the autosome group as they form spindle connections to one pole only. Since the chromosomes form a holocentric group at mitosis but behave as individual telocentrics at meiosis, their ensheathment by membranes during meiosis is thought to be essential for their orientation and orderly disjunction in the absence of true kinetochore plates.
Abstract Filamentous intranuclear crystals consisting of two sets of intersecting sheets are common in the intestinal epithelium. They are closely associated with chromatin granules but electron microscope cytochemistry using a modified Feulgen procedure did not show DNA in the crystal itself. Light microscope histochemical tests indicated that the crystals consist of protein.
Vegetative cells and spores of a yeast-like micro organism found in various tissues in both sexes of the ichneumonid wasp Pimpla turionellae are passed to the offspring by infection of the oocytes. Because of their intranuclearspindles with spindle pole bodies associated with the nuclear envelope as pole structures, the microorganisms are thought to be yeasts or closely related to yeasts. The high vitality and fertility of the wasps seem to exclude a pathogenic infection. Both the passage of vesicles from the microorganisms to the host cytoplasm and their transmission to the next generation by the oocytes point to anendosymbiotic relationship
Abstract During micronuclear metaphase, the decondensed chromosomes bear conical kinetochores connected by a single microtubule to the tapered ends of the spindle-shaped nucleus. Using Bernhard's procedure for the pre ferential staining of ribonucleo-protein-containing struc tures, the whole kinetochore cone retains its electron density. Chromosome separation at anaphase involves shortening of the kinetochore microtubule and elongation of the interpolar region.
Microtubule bundles anchored at the kinetosomes of cilia of the adoral zone of membranelles at the oral apparatus of Nyctotherus ovalis seem to represent a fairly rigid framework. No evidence for gliding or any other form of motility of these bundles could be obtained. Food uptake at the base of the funnel-shaped oral tube is accompanied by the appearance of electron-translucent discoidal vesicles between microtubule ribbons and nemadesmata-like microtubules inserting at the kinetosomes of cilia of the paroral membrane. After incubation with peroxidase, label is found in large food vacuoles but not in these discoidal vesicles. Such vesicles are still found after exposure of ciliates to cycloheximide. Colchicine, nocodazole, and low concentrations of vinblastine do not visibly affect the cytopharyngeal microtubules, and the transport of the vesicles appears undisturbed. However, vesicles are absent from microtubule ribbons in ciliates exposed to high concentrations of vinblastine or low temperature, although the microtubules themselves are morphologically unchanged. The presumably exocytotic membrane flow continues after treatment with D2O, implying that the discoidal vesicles are not transported along the bundles by attachment to ‘treadmilling’ microtubules [35]. Since we observed microfilaments in the cytoplasm of vinblastine-treated ciliates and at the cytopharynx of Triton-lysed cell models these filaments may be involved in vesicle movement and food uptake. However, cytochalasin B has no immediate effect on the distribution of the discoidal vesicles. The appearance of fine filaments in the vicinity of the presumed endocytotic site of the cytopharynx and the attachment of these filaments to microtubules and membranes rather suggests that microfilaments together with the microtubules could cooperate in food vacuole formation. Evidence for a direct participation of microtubules in the transport of the discoidal vesicles was not obtained.
After 5 h in 10(-3)M vinblastine the most obvious effects upon Vicia faba L. cells are seen in the spindle apparatus. These include the microtubules themselves as indicated by c-type metaphases and the pole regions of otherwise intact spindles, leading to multipolar anaphases and to telophases with more than two daughter nuclei. Vesicle transport may be undisturbed and new cell walls can be formed, although cases of disturbed cell plate and cell wall formation were noted occasionally, accompanied by myelinizations in phragmosomes. After 24 h in the same concentration of vinblastine, divisions are no longer observed and the plasma membranes are severely affected. They show extensive myelinizations, accumulations of lipids and dehiscence from the cell walls which are frequently thickened and irregularly formed. Of the other cellular membranes, the nuclear envelope and, more frequently, the tonoplast may be affected. Electron-dense deposits appear in the vacuoles. Comparable, though less severe, changes including multipolar anaphases and myelinizations result from treatment with lumicolchicine, but not with colchicine. Vinblastine leads to the appearance of filament bundles both in cytoplasm and karyoplasm, lumicolchicine to morphologically identical filaments in the cytoplasm alone. The nature of these filaments is unknown.
Exposure of Nyctotherus ovalis to low temperatures or vinblastine caused similar reactions of “classes” of microtubules (mt) present in the mitotic micronucleus of this ciliate towards both treatments. However, differences of sensitivity between certain “classes” of mt at individual mitotic stages exist. Unlike the kinetochore mt (kmt) of most other eukaryotic cells, kmt in Nyctotherus completely disassemble after incubation at 6–8 ° C (60 min) and most disappear after prolonged exposure to vinblastine (10−5 M, 16 h). The depolymerization of kmt causes the collapse of the spindle and a dislocation of chromosomes at metaphase, yet the reduced number of kmt after vinblastine-treatment still allows an alignment of composite complexes at the spindle equator. The data suggest that three individual sets of mt exist in the interpolar spindle region during ana- and telophase: 1) interpolar mt (int mt), which are assembled during anaphase, are cold- and vinblastine sensitive; 2) manchette mt (ma mt), which are first observed underneath the nuclear envelope during mid-anaphase, are cold-stable and insensitive to vinblastine treatment (10−5 M); after prolonged treatment (16 h) they form spiral structures; 3) stembody mt (st mt), comprising the interpolar region of the nucleus during telophase, are cold- and vinblastine insensitive. Paracrystalline structures resembling a stembody are formed in telophase-like division stages after prolonged vinblastine exposure (16 h, 10−5 M). Since kmt and int mt possess the same sensitivity under depolymerizing conditions, they probably have a similar composition. Thus the idea that the int mt in this organism arise by elongation of kmt is supported. However, st mt apparently do not originate from an extension of preexistent int mt, but appear to represent a new set of stable mt. This is emphasized not only by their greater stability compared to the int mt but also by the distribution of cold-stable mt in late anaphase micronuclei. The ma mt may be an intermediary step in formation of st mt since their stability resembles that of the st mt. A comparison of the substructure of vinblastine-induced paracrystals in Nyctotherus with those observed in in vitro systems with known composition suggests that a turnover of MAPs may be responsible for the different stability of mt and thus could specify and regulate mt sensitivity and function. Another organelle, possibly involved in conferring stability to mt, is the nuclear membrane. The assumption that the nuclear envelope possesses an intrinsic property to nucleate mt and thus aid in the alignment of mt is supported.
Abstract Under culture conditions the egg nucleus of Trichoplax adhaerens reaches an unusually high and variable DNA content. Before the "fertilization membrane" is formed, the nucleus undergoes fragmentation. It is assumed that culture conditions differ from natural conditions by preventing the switch-over from the S-phase to the G 2-phase of the cell cycle: DNA replication continues and nuclear division is impossible. Cleavage ceases at an earlier or later stage.
The formation of homopolar doublets can be induced by the action of antibiotics which inhibit the growth of cytoplasmic bacterial symbionts whose cell cycle appears to be controlled by the host. They multiply during the macronuclear S-phase of the ciliate and become enclosed in vesicles and are largely destroyed just before cell division is completed. Since doublet formation is due to an incomplete cell division, and because experimental disturbances at the cell cortex of dividing ciliates also lead to doublets, the symbionts are thought to contribute some factor which is essential for normal cytokinesis of the host cell.
Trichoplax cells can be dispersed after the action of colchicine, vinblastine, and sea water free of divalent ions. Since lumicolchicine, which is inactive against microtubules can also cause disaggregation, the plant alkaloids are thought to be bound to components of the cell membrane involved in cellular adhesion. After washing, the cells reaggregate spontaneously forming first irregular lumps capable of fusion and then spherical aggregates in which the different cell types have sorted out. The rebuilding of tissues can, in principle, be understood on the basis of differential cellular adhesion.
The existence of an X1X2-mode of sex determination is confirmed by a study of all meiotic stages in the male cotton stainer (X1X2 and pertinent stages in the female (X1X1 X2X2). In the male, the X-chromosomes are heterochromatic and pair end-to-end in early meiotic prophase. At diakinesis, they disjoin and align side-by-side in the center of the spindle, forming a pseudotetrad. Anaphase I is equational for the sex chromosomes. At late anaphase or telophase, X1 and X2 join end-to-end but form spindle fiber connections to only one of the poles of the metaphase II spindle, leading to one daughter cell without X chromosomes and one with both X1 and X2. An attempt is made to explain sex chromosome pairing and orientation on the basis of a telocentric organization of meiotic chromosomes. The apparent differences in the kinetic organization of mitotic and meiotic chromosomes in Heteroptera are discussed.
