ALBERT

Notes: In exconjugants of the hypotrich Keronopsis a large, highly polyploid macronuclear anlage is formed from which condensed chromatin bodies are passed into the cytoplasm where they are thought to give rise to the numerous small macronuclei of the vegetative cell. Electron microscopy shows that the chromatin bodies within the macronuclear anlage are separated from each other by sheets of low contrast lamellar material. The anlage appears therefore as a composite nucleus containing prepacked units which are extruded into the cytoplasm following condensation.

Notes: The hypotrichous ciliate Keronopsis rubra has ∼10 micronuclei and ∼100 small macronuclei. DNA synthesis proceeds synchronously in all macronuclei in the 2nd half of the cell cycle which takes about 24 hr at room temperature. A G2 phase is virtually absent, each nucleus dividing as soon as the replication band has passed over it. The micronuclear S phase falls within macronuclear G1 and is followed by immediate division. Comparative cytophotometric measurements of Feulgen-stained preparations indicate that the DNA content of G1 macronuclei is scattered widely in a skewed normal distribution, with a peak corresponding to the DNA content of a G1 micronucleus. Measurements of dividing macronuclei indicate unequal distribution of DNA between daughter nuclei and lead to the conclusion that the units of assortment must be smaller than whole genomes unless the micronucleus is polyploid. After conjugation, a large macronuclear anlage with threads resembling split prophase chromosomes is formed. The threads condense and pass singly into the cytoplasm where they are thought to give rise to the numerous small macronuclei of the vegetative cells.

Notes: After 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.

Notes: Summary Two types of spherical forms of this normally flattened organism appear sporadically in our cultures. Hollow spheres have an outer wall of flagellated ventral epithelium. The large fiber cells protrude into the central cavity which can include a closed compartment of flagellated dorsal epithelium. Cells of the outer wall that withdraw their flagellum and leave the epithelium are phagocytozed by fiber cells. Solid spheres consist of an outer layer of dorsal epithelium and densely packed fiber cells in the interior that may also include a closed compartment of ventral epithelium cells. Closely apposed fiber cells may form special cell contacts or pores connecting the cells.

Notes: Summary All cilia emerge from ciliary pits supported along their circumference by 22–24 dense rodlets that are connected by filaments to a surrounding sheath of endoplasmic reticulum. The proximal part of the basal body is provided with two short lateral rootlets and one long terminal rootlet, all associated with microtubules. The lateral rootlets are in turn connected by fine fibrous material to the dense supporting rodlets which follow the contour of the ciliary pit and extend along the ciliary membrane beyond the level of the basal plate where the central pair of microtubules originates. The proximal part of the basal body has fine fibrous connections to the endoplasmic reticulum while its distal portion is surrounded by nine curved sheets. The terminal cell contactions are by belt desmosomes that are accompanied by a bundle of microfilaments which encircle the apical region of the cell and insert at the cell membrane. Tight junctions are lacking. Endocytosis was demonstrated by the uptake of cationized ferritin. The structures associated with the ciliary pits are probably associated with the firm anchorage of the ciliary base since Trichoplax adheres to the substrate as it moves propelled by its ventral cilia. The marginal bundle of microfilaments may be involved in folding of the organism during feeding.

Notes: Summary The cytoskeleton of Trichoplax adhaerens fiber cells was studied after chemical fixation, freeze-substitution, lysis of attached cells with nonionic detergents and by immunofluorescence. Cytoskeletal elements present in the cell bodies and reaching into the extensions include microtubules, intermediate filaments, 6–7 nm and 2–3 nm microfilaments. The latter seem to interconnect other cytoskeletal elements. Actin-like microfilaments are found both as networks and parallel strands. Immunofluorescence with antiactin shows the presence of actin in the cell body, underneath the plasmalemma and within the extensions. Both the results of immunofluorescence and the identification of 6–7 nm actin-like microfilaments support the concept of contractility of the fiber cells as the cause of the rapid shape changes of Trichoplax. Anti-tubulin fluorescence corresponds to the location of microtubules in the extensions as well as the cell bodies of the fiber cells. The extensions are withdrawn upon depolymerization of the microtubules by colchicine.

Notes: Summary The female accessory glands include the tubular poison gland, the paired, lemon-shaped uterus glands, and Dufour's gland, an unbranched tubular organ. They consist essentially of a single layer of epithelium cells surrounded by a basement membrane. The lumen is lined by cuticle. The proteinaceous secretion of the poison gland is released into intracellular ducts provided with microvilli, each connected to a channel lined with cuticle which leads to the central lumen of the gland. The channel is formed by special canal cells. Nerve endings are interspersed among the gland cells. The uterus gland consists of four cell types derived from a single type of precursor cell found in newly hatched wasps. Type I cells are covered by type II cells and are thus without contact to the luminal surface of the gland. They contain stacks or whorls of mitochondria and smooth cisternae in an alternating arrangement. Vesicles with a secretory product are found in cells of types II and III. Deep anastomosing infoldings of the plasmalemma, stabilized by microtubules and dense material at the branchings, are characteristic for type II cells. Most secretory vesicles are found in type III cells, the prevalent cell type which is thought to be the source of the lipoprotein secretion. Coated vesicles are present at deep infoldings of the plasmalemma. The greatly enlarged apical surface area of type IV cells and the presence of mitochondria in slender outgrowths is suggestive of an osmoregulatory function. In Dufour's gland, two cell types appear in succession, the first with a very dense cytoplasm, the second with dense inclusions and many seemingly empty vesicles of smooth endoplasmic reticulum. The secretion products, lecithin and a cholesterol ester, are thought to be formed by the second cell type. The dense inclusion might be lecithin, which reacts with osmium tetroxide. The cholesterol ester could have been washed out of the “empty” vesicles by the embedding procedure.

Notes: Summary Fiber cells isolated by mechanical disruption of the tissue in Ca2+-free sea water attach firmly to the substrate by discrete adhesion plaques. They are capable of forming a lamellipodium and long, slender extensions while the cell bodies remain stationary. The extensions are slowly elongated but can suddenly be withdrawn by contraction. Bundles of F-actin are attached with their “plus” ends close to the tips of the extensions. Within the cell body, microfilaments form a loose cortical layer and criss-crossing patches. Microtubules are not present in newly formed extensions but seem to stabilize “older” extensions. In these, they are bundled by crossbridges and oriented lengthwise. In the cell body, 37 nm “macrotubules” are found as well as the prevailing microtubules 24 nm in thickness. They radiate from an organizing center close to the mitochondrial complex, which can, however, also give rise to normal microtubules. Since this organizing center seems to nucleate either macrotubules or microtubules, but never both at the same time, it is speculated that it may exist in two alternative states.

Notes: Summary Hollow swarmers are budded off at the dorsal surface ofTrichoplax and are covered by dorsal epithelium. Their inner cavity is lined with the flagellated cells of the ventral epithelium. There is no indication that the fiber cells included between the epithelia take any part either in morphologenesis or the separation of the bud from the mother animal. The early primordium forms in the interspace. A single layer of cells derived from both epithelia surrounds a cavity filled with granular matter that stains like proteins. The latter is used up during the floating phase of the swarmers that may last for a week. After settling at the bottom, the hollow sphere opens at one point. The concave ventral epithelium gradually flattens as more cells become incorporated in it. The latter form new flagella and flagellar pits. More frequently found than swarmers are small spherical forms that are unable to float and possess a distinct polarity. Their upper half is covered by dorsal epithelium and their lower half by ventral epithelium. Large fiber cells are in the center. Their site and mode of formation is unknown. Rarely observed are dorsal stolons whose bulbous end flattens upon touching the substrate. Since they are totally covered by the flat cells of the dorsal epithelium, they may have to undergo a transformation, like the hollow swarmers, to bring the ventral epithelium into contact with the substrate.

Notes: Abstract The first microtubules which appear in the prophase micronucleus of Colpoda steinii are located beneath the nuclear envelope and not connected to the chromosomes. Most microtubules of the metaphase spindle are connected to the tapered tips of the micronucleus and terminate singly at the chromosomes surrounded by a conical, RNA-containing kinetochore which disappears upon cold treatment. During anaphase, an interzonal stembody is formed which is maximally stretched at telophase before the daughter micronuclei are pinched off from its ends. The macronucleus, which also stretches parallel to the micronuclear stembody, has fewer microtubules which insert at the inner nuclear envelope but are not attached to the chromatin. Based upon the effects of depolymerizing factors different classes of microtubules can be distinguished. Kinetochore microtubules are sensitive to cold and vinblastine (VLB). In 2.5×10−5 M VLB their number is drastically reduced and the interzonal microtubules of early anaphase, which are also highly sensitive to nocodazole, become completely disassembled. The cross-bridged microtubules of the fully formed stembody of late anaphase display the highest resistance to depolymerization. They show signs of partial disassembly only after prolonged cold exposure and withstand higher concentrations of VLB or nocodazole than other micronuclear microtubules. Microtubules in the elongating macronucleus are fairly insensitive to cold but are depolymerized by 5×10−5 M VLB while 1.66×10−5 M nocodazole, which leaves only traces of stembody microtubules, merely reduces their number and length. All microtubules are fairly resistant to colchicine since high concentrations (5×10−2 M) are required to prevent assembly while fully formed stembodies are unaffected. Macronuclear microtubules are depolymerized at this concentration. Nocodazole, which depolymerizes all premetaphase microtubules at 6.6×10−6 M, leads to multipolar metaphase spindles with numerous microtubules, even at 1.66×10−5 M, an effect ascribed to the activity of the nuclear envelope as a microtubule organizing centre. At twice this concentration multipolar spindles are no longer found and the remaining microtubules show no apparent order. A stabilizing influence of the micronuclear envelope is indicated by the fact that whenever remnants of microtubules are found after depolymerizing treatments, they are located in its vicinity.