Centrioles in flies: The exception to the rule?

Centrioles and basal bodies are MT based structures that present a highly conserved ninefold symmetry. Centrioles can be found at the core of the centrosome where they participate in PCM recruitment and organization, contributing to cytoplasmic MT nucleation. Basal bodies are normally located closely to the plasma membrane where they are responsible for axoneme assembly to form structures such as cilia or flagella. While it is well accepted that these organelles have important roles in cell and tissue organization, their contribution to certain phases of animal development is still not entirely established. Here we review the role of centrosomes and cilia in Drosophila melanogaster and briefly discuss the implications of these findings to other model organisms.     

Modes of the formation of elementary bodies and their isolation from the cell in L-form bacteria

Elementary bodies are formed on the cell surface and inside the cell body in all cell types characteristic of L-form cultures, i. e. spherical cells, large bodies and filament structures. The following ways of elementary body formation are described: by budding on the cell surface, appearance immediately in the cytoplasm, in the vacuole, as a result of cytoplasmic fragmentation accompanied by the lysis of the cell, as well as in cases of the separation of cytoplasmic areas surrounded by the membrane or the myelin-like structure. The release of elementary bodies from the cell occurs as a result of the lysis or death of the mother cell, the thinning of the vacuole wall, and possibly due to small transient defects in the membrane, not accompanied by the death of the mother cell. The scheme of the formation and release of elementary bodies from the cell is presented.     

Electron Microscopic Observations on the Structure of the Envelopes of Mature Elementary Bodies and Developmental Reticulate Forms of Chlamydia psittaci

Purified suspensions of Chlamydia psittaci were prepared from L cells. Thin sections of intact elementary bodies and intact developmental reticulate bodies and of their purified envelopes were observed by electron microscopy. In both intact organisms and partially purified envelopes, two membranous structures, each appearing in electron micrographs as two darkly stained layers, were observed. In the elementary body sections, the outer membrane was round, apparently rigid, and was not soluble in 0.5% sodium dodecyl sulfate. The inner layer was irregular in shape and was completely removed by detergent treatment. We interpret these results to indicate that the outer rigid layer of the envelope is the cell wall and the inner layer is the cytoplasmic membrane. When the fragile reticulate body envelopes were similarly studied, the outer cell wall was clearly visible, and some evidence of an inner membrane was seen. After treatment with nucleases and detergent, all evidence of inner or cytoplasmic membrane was removed, but the outer cell wall remained. Thus, it appears that the cell wall of this organism is continuous throughout the growth cycle and that the fragility and lack of rigidity of the reticulate body cell is due to changes in chemical composition or structure of the cell wall.     

Assembly of the Paraflagellar Rod and the Flagellum Attachment Zone Complex During the Trypanosoma brucei Cell Cycle

Trypanosomes possess a single flagellum that is attached to their cell body via the flagellum attachment zone (FAZ). The FAZ is composed of two structures: a cytoplasmic filament complex and four microtubules situated next to it. There is a complex transmembrane crosslinking of this FAZ to the paraflagellar rod (PFR) and axoneme within the flagellum. We have partially purified the FAZ complex and have produced monoclonal antibodies both against the FAZ and the paraflagellar rod. The two antibodies against the FAZ (L3B2 and L6B3) recognise the cytoplasmic filament in immunofluorescence and in immunoelectron microscopy. On western blot, they detect a doublet of high molecular weight (M(r) 200,000). Two anti-PFR antibodies (L13D6 and L8C4) recognise the paraflagellar rod in immunofluorescence, but show a difference on Western blot: L13D6 recognises both major PFR proteins, whereas L8C4 is specific for only one of them. Using these new antibodies we have shown that although the growth of both cytoplasmic FAZ filament and external PFR are related, their growth initiates at different time points during the cell cycle and the two structures elongate at distinct rates.     

Immunofluorescence microscopic demonstration of vimentin filaments in asteroid bodies of sarcoidosis. A comparison with electron microscopic findings

Asteroid bodies in multinucleate giant cells from sarcoid granulomas were investigated by immunofluorescence and electron microscopy. The following points have been established: 1. Asteroid bodies are made up of individual components of the so-called cytoskeleton, predominantly vimentin filaments. Microtubules are involved in smaller amounts in the formation of the asteroid bodies. 2. They arise within the area of the cytosphere. The body of the asteroid includes the centrioles while the arms of the asteroid usually extend into the Golgi area and occasionally up to the cell nuclei. 3. Asteroid bodies result from aggregation of the flexible filamentous and microtubular systems of the centrosphere. The processes of aggregation probably result from local fluid shifts and sol-gel transformations. 4. The stellate form of the aggregations is determined by the preexistent radial arrangement of the elements of the cytosphere. 5. The prevailing specific environment of the underlying granulomatous disease, together with the internal characteristics of the structure and function of the giant cells, in particular in states of exhaustion may play a part in their development.     

TRIM5 alpha cytoplasmic bodies are highly dynamic structures

Tripartite motif (TRIM)5 alpha has recently been identified as a host restriction factor that has the ability to block infection by certain retroviruses in a species-dependent manner. One interesting feature of this protein is that it is localized in distinct cytoplasmic clusters designated as cytoplasmic bodies. The potential role of these cytoplasmic bodies in TRIM5 alpha function remains to be defined. By using fluorescent fusion proteins and live cell microscopy, we studied the localization and dynamics of TRIM5 alpha cytoplasmic bodies. This analysis reveals that cytoplasmic bodies are highly mobile, exhibiting both short saltatory movements and unidirectional long-distance movements along the microtubule network. The morphology of the cytoplasmic bodies is also dynamic. Finally, photobleaching and photoactivation analysis reveals that the TRIM5 alpha protein present in the cytoplasmic bodies is very dynamic, rapidly exchanging between cytoplasmic bodies and a more diffuse cytoplasmic population. Therefore, TRIM5 alpha cytoplasmic bodies are dynamic structures more consistent with a role in function or regulation rather than protein aggregates or inclusion bodies that represent dead-end static structures.     

The LIM domain protein UNC-95 is required for the assembly of muscle attachment structures and is regulated by the RING finger protein RNF-5 in C. elegans

Here, we describe a new muscle LIM domain protein, UNC-95, and identify it as a novel target for the RING finger protein RNF-5 in the Caenorhabditis elegans body wall muscle. unc-95(su33) animals have disorganized muscle actin and myosin-containing filaments as a result of a failure to assemble normal muscle adhesion structures. UNC-95 is active downstream of PAT-3/beta-integrin in the assembly pathways of the muscle dense body and M-line attachments, and upstream of DEB-1/vinculin in the dense body assembly pathway. The translational UNC-95::GFP fusion construct is expressed in dense bodies, M-lines, and muscle-muscle cell boundaries as well as in muscle cell bodies. UNC-95 is partially colocalized with RNF-5 in muscle dense bodies and its expression and localization are regulated by RNF-5. rnf-5(RNAi) or a RING domain deleted mutant, rnf-5(tm794), exhibit structural defects of the muscle attachment sites. Together, our data demonstrate that UNC-95 constitutes an essential component of muscle adhesion sites that is regulated by RNF-5.     

X-ray microanalysis on the thyroid follicle of the hagfish, Eptatretus burgeri and lamprey, Lampetra japonica.

Intracellular dense bodies, cytoplasmic matrices, and luminal colloids in the thyroid follicle of cyclostomes, hagfish and lamprey were examined to identify the distribution of iodine using an energy dispersive X-ray microanalyzer attached to a scanning transmission electron microscope. High level of iodine was detected only in the large dense body of the hagfishes, while it was too slight in quantity to detect by this method in the cytoplasmic matrix as well as in the luminal colloid. In the adult lamprey thyroid, an appreciable amount of iodine was detected in a few large dense bodies. In mice and rats, it is very hard to detect iodine in the luminal colloid, intracellular colloid droplet, and in the lysosomal dense granule by this method, though these structures have been well known to contain a fairly large amount of iodine which is combined with thyroglobulin. These facts mean that intracellular dense bodies in the thyroid follicular epithelium of the cytoclostome, especially of the hagfish have an extremely larger amount of iodine. These bodies are considered to be secondary lysosomes or residual bodies containing reabsorbed colloid materials which are highly condensed.     

Goblet cell membrane differentiations in the midgut of a lepidopteran larva.

So-called goblet cells are present in the midgut of lepidopteran larvae. They are thought to be involved in the active transport of potassium out of the haemolymph and into the gut lumen. A number of plasma membrane differentiations within the goblet cell cavity has been investigated using conventional staining, lanthanum tracer and freeze-etch techniques. Of particular interest are junction-like inter- and intra-membrane differentiations found on the villus-like cytoplasmic projections present at the apical tip of the goblet cell cavities. These cytoplasmic projections appear to act as a valve; in some cases they seem to close off the top of the goblet cell cavity, so isolating it from the gut lumen, while in other cases they are spread apart leaving a wide channel from the cavity into the lumen. The junction-like structures on these cytoplasmic projections are different in structure from the septate-type junctions which seal the midgut cells together at their apical borders, and the 2 types are present on the same plasma membrane, often within one micron of each other. The need for a different type of junction may possibly be related to the fact that it occurs between 2 areas of the same plasma membrane. The morphology of this unusual junction-like structure is discussed and 2 diagrams are presented to illustrate our interpretation of its structure.     

Novel roles for saccharomyces cerevisiae mitotic spindle motors.

The single cytoplasmic dynein and five of the six kinesin-related proteins encoded by Saccharomyces cerevisiae participate in mitotic spindle function. Some of the motors operate within the nucleus to assemble and elongate the bipolar spindle. Others operate on the cytoplasmic microtubules to effect spindle and nuclear positioning within the cell. This study reveals that kinesin-related Kar3p and Kip3p are unique in that they perform roles both inside and outside the nucleus. Kar3p, like Kip3p, was found to be required for spindle positioning in the absence of dynein. The spindle positioning role of Kar3p is performed in concert with the Cik1p accessory factor, but not the homologous Vik1p. Kar3p and Kip3p were also found to overlap for a function essential for the structural integrity of the bipolar spindle. The cytoplasmic and nuclear roles of both these motors could be partially substituted for by the microtubule-destabilizing agent benomyl, suggesting that these motors perform an essential microtubule-destabilizing function. In addition, we found that yeast cell viability could be supported by as few as two microtubule-based motors: the BimC-type kinesin Cin8p, required for spindle structure, paired with either Kar3p or Kip3p, required for both spindle structure and positioning.     

A FINE STRUCTURE STUDY OF LIPID IN MOUSE LIVER REGENERATING AFTER PARTIAL HEPATECTOMY

The fine structure of liver 3½ to 72 hours after partial hepatectomy has been compared with that of liver from sham-operated animals; all animals were 60- to 90-day old male mice of the C3H strain. Numerous small bodies with diameters ranging from 300 to 1,000 A have been observed distributed randomly throughout the cytoplasm of the hepatic parenchymal cells at early intervals after partial hepatectomy. In material fixed in osmium tetroxide and embedded in methacrylate, they appear as uniformly electron-opaque bodies, but in permanganate-fixed liver, they display only a peripheral rim of electron-opaque material surrounding a clear core. Each of these cytoplasmic bodies appears to be located within a vesicle. A few of the opaque bodies are also present in sinusoids and in the spaces of Disse; these bodies are not located within vesicular structures. Fat droplets of various sizes are easily distinguished in regenerating liver; with the increase in number of these fat droplets at later postoperative intervals, there occurs a concomitant decrease in the number of cytoplasmic bodies. It is suggested that the cytoplasmic bodies contain some lipid component. Possible explanations of the origin, nature, and fate of the cytoplasmic bodies are discussed.     

Fine structure of degenerating abdominal motor neurons after eclosion in the sphingid moth,Manduca sexta

Summary Ultrastructural aspects of the natural degeneration of a group of six motor neurons in the fourth abdominal ganglion of Manduca sexta are described. These motor neurons innervate intersegmental muscles that degenerate and disappear immediately after adult eclosion. The first detectable changes in the cell bodies appear 12 h after eclosion and include disruption of the endoplasmic reticulum and an increase in the size and number of lamellar bodies. At 32 h the nuclear membranes rupture, and the membranous and granular cytoorganelles segregate in different parts of the cell. At that stage the surrounding glial cells participate in the digestion of material from the degenerating neurons. From 72 h onward the remaining neuronal structures become disrupted, and are finally transformed into a single, large lamellar body (residual body) within the glial profile. The degeneration pattern differs significantly from that of embryonic vertebrate neurons.     

Localization of mitochondrial ribosomal RNA on the chromatoid bodies of marine planarian polyclad embryos

Electron-dense cytoplasmic structures, referred to as chromatoid bodies, are observed in the somatic stem cells, called neoblasts, and germline cells in adult planarians. Although it has been revealed that the chromatoid bodies morphologically resemble germline granules in Drosophila and Xenopus embryos, what essential role it plays in the planarian has remained unclear. In the present study, to examine whether chromatoid bodies in planarian embryos are responsible for germline formation, the presence and behavior of chromatoid bodies during embryogenesis were examined. Mitochondrial large ribosomal RNA and mitochondrial small ribosomal RNA were used as candidate markers for components of the chromatoid body. Starting from the fertilized egg, extramitochondrial signals of both RNA (mtrRNA) were observed. At the ultrastructural level, mtrRNA were localized on the surface of the chromatoid bodies. At subsequent stages, the signals of mtrRNA were observed in certain restricted blastomeres that contribute to the formation of larval structures. The signals gradually decreased from the gastrula stage. These results suggest that the chromatoid bodies associated with mtrRNA in embryogenesis are not germline granules. The chromatoid bodies of blastomeres may be concerned with the toti- or pluripotency and cell differentiation as proposed in adult planarian neoblasts.     

The fine structure of myelinated nerve cell bodies in the bulbus olfactorius of man

Summary In the bulbus olfactorius of man numerous myelinated nerve cell bodies occur in the stratum plexiforme internum and stratum granulosum internum. In many respects they resemble the neighbouring granule cells: small chromatin clumps border on more than half of the circumference of the nucleus, the thin cytoplasmic rim contains abundant polysomes and sometimes pigment complexes with numerous light vacuoles, the cells often show a process which extends up to the stratum glomerulosum, the perikarya are devoid of synaptic contacts whereas the proximal segment of the peripheral processes display rare contacts. The myelin sheath varies in thickness, consisting of 2 to 24 lamellae with distances between the major dense lines ranging from 9.3 to 11.3 nm. The myelin sheath may enclose the cell body completely or partially and accompany the proximal segment of the process arising from the perikaryon. On partially enveloped perikarya, the myelin lamellae end in formations like those of the node of Ranvier, though often less regularly. Within the compact myelin sheath all of its lamellae may be distended for a short distance by glial cytoplasm as in the Schmidt-Lanterman incisures of peripheral nerve fibres. Adjacent to the outermost myelin lamella myelinated axons and cell bodies, tentatively identified as oligodendrocytes, as well as granule cells may be closely joined.Supported by the Deutsche Forschungsgemeinschaft (Br. 634/1)     

Cellular differentiation in the kidneys of newborn mice studies with the electron microscope

The structure of the kidney of the Swiss albino mouse changes progressively during the first 2 weeks after birth. Cells proliferate to form new nephrons, cells differentiate by acquiring specialized membranous components, and certain cytological features which are present at birth diminish in abundance or disappear. The differentiation of the cells of the cortical tubules has been studied using the light and electron microscopes. The tubules are partially and variably differentiated at birth. During the first 2 weeks after birth the brush border develops in the proximal tubules by the accumulation of numerous microvilli on the apical cell margins. Basal striations develop in proximal and distal tubules as an alignment of mitochondria, the result of what appears to be progressive interlocking of adjacent fluted cells. The mitochondria increase in number and size, accumulate homogeneous matrix, and acquire small, very dense granules. The collecting ducts develop tight pleating of the basal cell membranes, and dark cells containing numerous small cytoplasmic vesicles and microvilli appear. At birth there are dense irregular cytoplasmic inclusions presumed to be lipide in renal cells, the cytoplasmic granules of Palade are abundant, and there are large round bodies in the cells of the proximal tubules. The lipide inclusions disappear a few days after birth, and the cytoplasmic granules of Palade diminish in abundance as the cells differentiate. The large round bodies in the proximal tubules consist of an amorphous material and contain concentrically lamellar structures and mitochondria. They resemble the cytoplasmic droplets produced in the proximal tubules of adult rats and mice by the administration of proteins. The large round bodies disappear from the proximal tubules of infant mice during the first week after birth, but the concentric lamellar structures may be found in adult mice.     

Relationship between Cell Wall, Cytoplasmic Membrane, and Bacterial Motility

High-resolution electron microscopy of polarly flagellated bacteria revealed that their flagella originate at a circular, differentiated portion of the cytoplasmic membrane approximately 25 nm in diameter. The flagella also have discs attaching them to the cell wall. These attachment discs are extremely resistant to lytic damage and are firmly bound to the flagella. The cytoplasm beneath the flagellum contains a granulated basal body about 60 nm in diameter, and a specialized polar membrane. The existence of membrane-bound basal bodies is shown to be an artifact arising from adherence of cell wall and cytoplasmic membrane fragments to flagella in lysed preparations. Based on structures observed, a mechanism to explain bacterial flagellar movement is proposed. Flagella are considered to be anchored to the cell wall and activated by displacement of underlying cytoplasmic membrane to which they are also firmly attached. An explanation for the membrane displacement is given.     

Dense bodies and actin polarity in vertebrate smooth muscle

The arrangement of cytoplasmic dense bodies in vertebrate smooth muscle and their relationship to the thin filaments was studied in cells from rabbit vas deferens and portal vein which were made hyperpermeable (skinned) with saponin and incubated with myosin subfragment 1 (S-1). The dense bodies were obliquely oriented, elongated structures sometimes appearing as chains up to 1.5 microns in length; they were often continuous across the cell for 200 to 300 nm and were interconnected by an oblique network of 10-nm filaments. The arrowheads, formed by S-1 decoration of actins, which inserted into both the sides and ends of dense bodies, always pointed away from the dense body, similar to the polarity of the thin filaments at the Z- bands of skeletal muscle. These results show that the cytoplasmic dense bodies function as anchoring sites for the thin filaments and indicate that the thin filaments, thick filaments, and dense bodies constitute a contractile unit.     

The reserve proteins in the cells of mature cotyledons ofLupinus albus var.Lucky. I. Quantitative ultrastructural study of the protein bodies

Summary A quantitative ultrastructural study of the protein bodies of the lupin cotyledonary cells was performed to determine the protein content per cell. Two kinds of protein body were observed by transmission and scanning electron microscopy whatever the cellular type within the cotyledon. Some of these were conventional spherical structures, entirely filled with a dense protein matrix, others exhibited one large or several small light inclusions within the dense matrix. Even when a few light areas contained globoids, the majority remained of unknown nature, but could not be considered proteinaceous since they never reacted with specific protein dyes. The reserve protein content per cell was determined by image analysis on two seeds (L₁ and L₂) selected because they had a markedly different total protein content. The volume occupied by the dense matrix of the intracellular protein bodies was considered representative of the reserve protein quantity. The protein content per cell increased from the periphery to the centre of the cotyledon in the two seeds studied. The protein content per cell of the L₂ seed was generally found to be greater than the L₁ seed, in particular in the abaxial zone where it was markedly higher. The difference in total protein content of the two seeds was demonstrated to be primarily due to a differential alveolation of the protein bodies. These results will be used to study the relationship between the protein content of the cotyledonary cells and their nuclear DNA content.     

Electron microscopic study on the thyroid gland of the salamander Hynobius nebulosus in the breeding season

Summary The thyroid gland of adult salamanders, Hynobius nebulosus, in the breeding season was studied by electron microscopy. The follicular cells are different in cell height and fine structures; the taller cells with many cell organelles and granules and the lower cells with a few cell organelles and granules are both present in the same follicle. In the cytoplasm, three types of membrane-bounded granules, namely, cytosomes, colloid droplets, and vacuolar bodies and circular membrane complexes occur. The vacuolar bodies are subdivided into two types; the ordinary type having loosely distributed particles and the specific type containing tubules and/or closely packed filaments, crystalloid structures, except for the particles. The chromophobe colloids within the Bensley-cells correspond to extremely large, ordinary type vacuolar bodies, while the Langendorff-colloid cells possess increased numbers of granular cisternae of endoplasmic reticulum and a ribosome-rich, dense cytoplasmic matrix but not extremely large colloid. The intracytoplasmic circular membrane complexes appear in the Golgi area of cytosome-rich cells. It is suggested that they originate from the Golgi apparatus which was activated to produce many cytosomes. Intranuclear inclusions consisting of microtubules and filaments and tight junctions between two adjacent lateral plasma membranes are occasionally encountered.     

Ultrastructural studies of the nucleoids of the pleomorphic forms of Chlamydia psittaci 6BC: a comparison with bacteria.

The nucleoids of the various pleomorphic forms of Chlamydia psittaci have been examined by direct observation of infected cells and by observations on isolated particles. The fixation and staining methods used were the same as those routinely used for the examination of bacteria to facilitate the comparison of chlamydial fine structure with that of bacteria. The nucleoids of reticulate bodies were composed of fine fibrils which extended throughout these particles. The nucleoids of intermediate bodies are characterized by an electron-dense mass with which the fibrous elements are associated in a structurally coherent manner. As condensation of the intermediate bodies proceeds, the electron-dense mass becomes eccentrically located and the fibers form a distinct radiating structure. Large elementary bodies have a few fibers associated with their condensed electron-dense nucleoids but the more condensed mature elementary bodies have a very discrete and homogeneous electron-dense nucleoid which is separated from the cytoplasmic elements of these particles by a very distinct electron-transparent space. These highly condensed elementary body nucleoids are usually ovoid, but may be elongated or irregular, and a small number of these structures react very strongly with ruthenium red. While the nucleoid structure of reticulate bodies resembles that of the bacterial cell, both the condensation process and the nucleoid morphologies which result from it in intermediate and elementary bodies have no parallels among the bacteria. Thus we conclude that major differences in nucleoid organization exist between the chlamydia and the bacteria.