The control of anthozoan cilia by the basal apparatus

The ciliary basal apparatus in the pharynx of the sea anemone, Calliactis parasitica (Couch), is composed of two centrioles, a single striated rootlet at least 20 microns long, and a basal foot, to the tip of which is attached a bundle of microtubules leading to the rootlet. When the basal apparatus is sectioned in the plane of the ciliary power-stroke, the distal centriole, with which the cilium base is continuous, is rarely found to be erect. The orientation of the distal centriole is determined by bending in the basal apparatus. Bending occurs only in the plane of the ciliary power-stroke towards the side from which the basal foot projects, and it is closely correlated with membrane buckling in the belt desmosome region of the cell apex. Associated with the belt desmosome, but not directly with the basal apparatus, are bundles of filaments. These filaments are of two size classes, 5-6 and 10 nm in diameter. A model is presented in which the 5-6 nm filaments form the basis of a contractile system which mediates membrane buckling in the region of the belt desmosome. This action effectively shortens the cell apex and thus forces the apparatus to bend. The precise reorientation of the distal centriole is a result of the mechanical properties of the basal apparatus.     

Ultrastructural study of sensory cells of the proboscidial glandular epithelium of Riseriellus occultus (Nemertea,Heteronemertea)

Only one sensory cell type has been observed within the glandular epithelium of the proboscis in the heteronemertine Riseriellus occultus. These bipolar cells are abundant and scattered singly throughout the proboscis length. The apical surface of each dendrite bears a single cilium enclosed by a ring of six to eight prominent microvilli. The cilium has the typical 9×2 + 2 axoneme arrangement and is equipped with a cross-striated vertical rootlet extending from the basal body. No accessory centriole or horizontal rootlet was observed. Large, modified microvilli (stereovilli) surrounding the cilium are joined together by a system of fine filaments derived from the glycocalyx. Each microvillus contains a bundle of actin-like filaments which anchor on the indented inner surface of a dense, apical ring situated beneath the level of the ciliary basal body. The tip of the cilium is expanded and modified to form a bulb-like structure which lies above the level where the surrounding microvilli terminate. In the region where the cilium emerges from the microvillar cone, the membrane of the microvillar apices makes contact with a corresponding portion of the ciliary membrane. At this level microvilli and cilium are apparently firmly linked by junctional systems resembling adherens junctions. The results suggest that these sensory cells may be mechanoreceptors. © 1996 Wiley-Liss, Inc.     

The ciliary rootlet maintains long-term stability of sensory cilia

The striated ciliary rootlet is a prominent cytoskeleton originating from basal bodies of ciliated cells. Although a familiar structure in cell biology, its function has remained unresolved. In this study, we carried out targeted disruption in mice of the gene for rootletin, a component of the rootlet. In the mutant, ciliated cells are devoid of rootlets. Phototransduction and ciliary beating in sensory and motile cilia initially exhibit no apparent functional deficits. However, photoreceptors degenerate over time, and mutant lungs appear prone to pathological changes consistent with insufficient mucociliary clearance. Further analyses revealed a striking fragility at the ciliary base in photoreceptors lacking rootlets. In vitro assays suggest that the rootlet is among the least dynamic of all cytoskeletons and interacts with actin filaments. Thus, a primary function of the rootlet is to provide structural support for the cilium. Inasmuch as photoreceptors elaborate an exceptionally enlarged sensory cilium, they are especially dependent on the rootlet for structural integrity and long-term survival.     

Organization of the ciliary basal apparatus in embryonic cells of the sea urchin,Lytechinus pictus

Summary The basal apparatus of embryonic cells of the sea urchin Lytechinus pictus was examined by transmission electron microscopy and compared with the basal apparatus of other metazoan cells. The basal apparatus in these cells is associated with a specialized region of the apical cell surface that is encircled by a ring of microvilli. The basal apparatus includes several features that are common to all ciliated cells, including a basal body, basal foot, basal foot cap, and striated rootlet. However, a component not seen in the basal apparatus of other species has been observed in these cells. This structure is continuous with the striated rootlet, and its ultrastructure indicates that it is composed of the same components as the rootlet. This structure extends from the junction of the basal body and striated rootlet to the cortical region that surrounds the basal body. Based on its morphology and position, this structure is referred to as a striated side-arm. The striated side-arm is always aligned in the plane of the basal foot. Thus, both of these structures extend from the basal body in the plane of the effective stroke. It is suggested that the striated side-arm serves to stabilize the basal apparatus against force exerted by the cilium.     

The ultrastructure of primary cilia in quiescent 3T3 cells

Electron microscopy was used to investigate primary cilia in quiescent 3T3 cells. As in the case of primary cilia of other cell types, their basal centriole was found to be a focal point of numerous cytoplasmic microtubules which terminate at the basal feet. There are also intermediate filaments which appear to converge at the basal centriole. Cross-striated fibers of microtubular diameter, reminiscent of striated rootlets of ordinary cilia, appear associated with the proximal end of the basal centriole. Usually less than nine cross-banded basal feet surround the basal centriole in a well-defined plane perpendicular to the centriolar axis. The ciliary shaft was found to be entirely enclosed in the cytoplasm of fully flattened cells. In rounded cells, it could be found extending to the outside of the cell. Periodic striations along the entire shaft were observed after preparing the cells in a special way. The tip of the shaft showed an electron-dense specialization. Several unusual forms of primary cilia were observed which were reminiscent of olfactory flagella or retinal rods.Using tubulin antibody for indirect immunofluorescence, a fluorescent rod is visible in the cells [18] which we demonstrate is identical with the primary cilium.     

Muciliary transport in newt lungs: the ultrastructure of the ciliary apparatus in isolated epithelial sheets and in functional triton-extracted models

High voltage and conventional electron microscopy were used to investigate the ultrastructure of the ciliary apparatus in intact and in Triton-extracted, reactivated sheets of mucociliary epithelium isolated from newt lung. Each long (about 13 microns) ciliary axoneme terminates on a barrel-shaped basal body which is anchored in the apical cytoplasm by a variety of accessory structures. A basal foot is associated with the midpoint of each basal body and acts as a focal point for numerous microtubules (MTs). In many cases MTs can be seen to interconnect the feet of neighbouring basal bodies. Attached to the proximal end of each basal body and extending in a direction opposite the basal foot is a large 'ciliary root'. Each ciliary root is associated with a distinct bundle of 6-7 nm microfilaments which appear to stain with the specific F-actin probe NBD-phallacidin. A single 3-4 microns long striated rootlet inserts into each ciliary root and extends toward the cell nucleus through an extensive network of microfilaments. At the level of the basal plate 'Y-shaped' structures appear to connect each axonemal outer doublet MT to the plasma membrane. All of these ciliary accessory structures are present in the same relationship in Triton-extracted models. Their morphology and distribution indicates that they serve to anchor the cilia in the apical cytoplasm. In addition some of these structures appear to be responsible for maintaining the structural and functional integrity of the ciliary field in the demembranated and reactivated models.     

Comparative ultrastructure of the epidermal ciliary rootlets and associated structures in species of the Nemertodermatida and Acoela (Plathelminthes)

 The fine morphology of epidermal ciliary structures in four species of the Nemertodermatida and four species of the Acoela was studied, with emphasis on Meara stichopi (Nemertodermatida). The cilium of M. stichopi has a distal shelf and is proximally separated from the basal body by a cup-shaped structure. The bottom of the cup consists of a bilayered dense plate, or basal plate. The basal body consists of peripheral microtubule doublets continuous with those of the cilium. In the upper part of the basal body, the doublets are set at an angle and are anchored to the enclosing cell membrane by Y-shaped structures. The lower part of the basal body tapers eventually. The striated main rootlet arises on the anterior face of the basal body, initially like a flattened strap, and continues along the basal body shaped as a tube which further down becomes solid. The hour-glass-shaped posterior rootlet arises on the posterior face of the basal body. Contrary to the main rootlet, the striations in the proximal part of the posterior rootlet run parallel to the microtubule doublets of the basal body. A pair of microtubule bundles lead from the posterior rootlet to the two main rootlets in the hind ciliary row, and follow these to their lower tip. In the other species of the Nemertodermatida studied, the structure of the ciliary basal body and the ciliary rootlets is similar to that of M. stichopi. Structural differences in the species of the Acoela are that the lowermost end of the basal body is narrow and bent forwards, the proximal part of the main rootlet is trough-shaped, the main rootlet is accompanied by a pair of lateral rootlets and the posterior rootlet with associated microtubule bundles is thin. The epidermal ciliary structures in species of the Nemertodermatida and Acoela have a number of shared characters which are unique within the Plathelminthes. However, almost all of these characters are found in Xenoturbella bocki (Xenoturbellida), and some even in species of other ”phyla” of the ”lower” Metazoa. Hence, these characters cannot be considered apomorphic for the Acoelomorpha. A character seemingly present only in species of the Nemertodermatida and Acoela is the bilayered dense plate. This feature might represent an autapomorphic character state for the Acoelomorpha. Accepted: 7 March 1997     

Rootletin,a novel coiled-coil protein,is a structural component of the ciliary rootlet

The ciliary rootlet, first recognized over a century ago, is a prominent structure originating from the basal body at the proximal end of a cilium. Despite being the largest cytoskeleton, its structural composition has remained unknown. Here, we report a novel 220-kD protein, designated rootletin, found in the rootlets of ciliated cells. Recombinant rootletin forms detergent-insoluble filaments radiating from the centrioles and resembling rootlets found in vivo. An mAb widely used as a marker for vertebrate rootlets recognizes an epitope in rootletin. Rootletin has a globular head domain and a tail domain consisting of extended coiled-coil structures. Rootletin forms parallel in register homodimers and elongated higher order polymers mediated by the tail domain alone. The head domain may be required for targeting to the basal body and binding to a kinesin light chain. In retinal photoreceptors where rootlets appear particularly robust, rootlets extend from the basal bodies to the synaptic terminals and anchor ER membranes along their length. Our data indicate that rootlets are composed of homopolymeric rootletin protofilaments bundled into variably shaped thick filaments. Thus, rootletin is the long-sought structural component of the ciliary rootlet.     

The ciliated epidermis of Xenoturbella bocki (Platyhelminthes, Xenoturbellida) with some phylogenetic considerations

The epidermis of Xenoturbella bocki Westblad was studied by scanning and transmission electron microscopy. Two cell types predominate in the epidermis: multiciliated epidermal cells and non-ciliated or monociliated gland cells. A conspicuous feature is the dense ciliary coverage and the numerous gland cell openings. Xenoturbella has a characteristic pattern of axonemal filament termination in the distal tips of their cilia. Each epidermal cilium has the typical 9 + 2 patten through the major part of its shaft. Near the tip there is a shelf at which doublets 4–7 terminate. Doublets 1, 2, 3, 8 and 9 continue into the thinner distal part of the cilium. A similar shelf in cilia is known only from the turbellarian orders Nemertodermatida and Acoela, and hence may be an apomorphic feature which indicates a close relationship between Xenoturbellida, Nemertoder-matida and Acoela. The basal body is provided with a so-called basal foot which has a cross-striated appearance and an expanded distal plate that seems to act as a microtubule organizing center. Approximately 15–25 microtubuli radiate from the endplate of the basal foot to the basal bodies caudally. The arrangement of basal foot and ciliary rootlets in Xenoturbella differs from that of Acoela and related orders in that there are two striated rootlets only (an anterior and a posterior one), rather than one main rootlet and two lateral rootlets.     

ULTRASTRUCTURE OF THE FLAGELLAR APPARATUS OF PYROBOTRYS (CHLOROPHYCEAE)1

The flagellar apparatus of Pyrobotrys has a number of features that are typical of the Chlorophyceae, but others that are unusual for this class. The two flagella are inserted at the apex, but they extend to the side of the cell toward the outside of the colony, here designated as the ventral side. Four basal bodies are present, two of which extend into flagella. Four microtubular rootlets alternate between the functional and accessory basal bodies. In each cell, the two ventral rootlets are nearly parallel, but the dorsal rootlets are more widely divergent. The rootlets alternate between two and four microtubules each. A striated distal fiber connects the two functional basal bodies in the plane of the flagella. Two additional, apparently nonstriated, fibers connect the basal bodies proximal to the distal fiber. Another striated fiber is associated with each four-membered rootlet near its insertion into the flagellar apparatus. A fine periodic component is associated with each two-membered rootlet. A rhizoplast-like structure extends into the cell from each of the functional basal bodies. The arrangement of these components does not reflect the 180° rotational symmetry that is usually present in the Chlorophyceae, but appears to be derived from a more symmetrical ancestor. It is suggested that the form of the flagellar apparatus is associated with the unusual colony structure of Pyrobotrys.     

Cytoskeleton Structure and Composition in Choanoflagellates

The structure and composition of the cytoskeleton has been studied in Monosiga ovata (Protozoa: Order Choanofiagellida Kent 1880) using a combination of methods in association with light and electron microscopy. Supplementary observations are included for Desmarella moniliformis. The basal body of the single anterior flagellum is subtended proximally and at right angles by a second, non-flagellar basal body. The edges of the two basal bodies are connected by a fibrillar bridge. A long, narrow, striated, fibrillar rootlet extends posteriorly from the lower edge of the non-flagellar basal body towards the Golgi apparatus. It is associated throughout most of its length with the surface of a flattened sac. Rootlet microtubules pass radially from a ring of electron dense material which encircles the distal end of the flagellar basal body. These microtubules extend outwards for about one-third of the length of the cell. Within each collar tentacle is a longitudinal bundle of microfilaments composed of actin as illustrated by rhodamine-phalloidin staining for fluorescence microscopy. The base of each microfilament bundle is associated with one or more rootlet microtubules by fine fibrillar bridges. The attachment between microtubules and tentacle microfilaments is further demonstrated by their coordinated displacement when the cytoskeleton becomes dislodged. The role of the cytoskeleton in maintaining the position of the collar tentacles during interphase and cell division is discussed.     

Choanocyte ultrastructure in Halisarca dujardini (Demospongiae,Halisarcida)

Understanding poriferan choanocyte ultrastructure is crucial if we are to unravel the steps of a putative evolutionary transition between choanoflagellate protists and early metazoans. Surprisingly, some aspects of choanocyte cytology still remain little investigated. This study of choanocyte ultrastructure in the halisarcid demosponge Halisarca dujardini revealed a combination of minor and major distinctive traits, some of them unknown in Porifera so far. Most significant features were 1) an asymmetrical periflagellar sleeve, 2) a battery of specialized intercellular junctions at the lateral cell surface complemented with an array of lateral interdigitations between adjacent choanocytes that provides a particular sealing system of the choanoderm, and 3) a unique, unexpectedly complex, basal apparatus. The basal apparatus consists of a basal body provided with a small basal foot and an intricate transverse skeleton of microtubules. An accessory centriole, which is not perpendicular to the basal body, is about 45°. In addition, a system of short striated rootlets (periodicity = 50–60 nm) arises from the proximal edge of the basal body and runs longitudinally to contact the nuclear apex. This is the first flagellar rootlet system ever found in a choanocyte. The accessory centriole, the rootlet system, and the nuclear apex are all encircled by a large Golgi apparatus, adding another distinctive feature to the choanocyte cytology. The set of distinct features discovered in the choanocyte of H. dujardini indicates that the ultrastructure of the poriferan choanocyte may vary substantially between sponge groups. It is necessary to improve understanding of such variation, as the cytological features of choanocytes are often coded as characters both for formulation of hypotheses on the origin of animals and inference of phylogenetic relationships at the base of the metazoan tree. J. Morphol., 2009. © 2008 Wiley‐Liss, Inc.     

The structure and distribution of the cross-striated fibril and associated membranes in guinea pig photoreceptors

Examination of longitudinally and transversely sectioned photoreceptor cells of the guinea pig retina revealed an aggregate of thin filaments forming a single cross-striated fibril coursing through the full length of the non-receptor portion of the cell. The fibril begins as the ciliary rootlet from the region of the basal body of the connecting cilium. From the basal body it passes between the mitochondria of the ellipsoid and along the Golgi zone of the myoid region of the inner segment, narrowing from an irregularly shaped bundle to a ribbon-shaped aggregate. The fibril separates into discrete strands, each curving along the nucleus, reuniting into a single bundle to pass down the cell's axon, and terminating deep within the synaptic terminal. The fibril is flanked by two separate membranous saccules, each continuous along nearly its full length. The fibril's extensive course in the guinea pig and its association with continuous membranes necessitates a reexamination of earlier proposals for the function of cross-striated filamentous structures in photoreceptor cells.     

A fine structural study of the testicular wall and spermatogenesis in the crinoid,Florometra serratissima (Echinodermata)

The testicular wall and the process of spermatogenesis in the crinoid, Florometra serratissima, has been studied at the fine structural level. The testicular wall is composed of three layers: a perivisceral layer consisting of nerve processes, muscle fibers, and epithelial cells; a haemal sinus containing haemal fluid, collagen-like fibers, and haemocytes; and a germinal layer consisting of germinal and interstitial cells. The germinal layer is elaborated into numerous folds that project into the lumen of the testis and a branch of the haemal channel extends through the core of each fold. Evidence suggesting that nutrients are carried to the testis and germinal cells via the haemal system is presented. Spermatogonia are concentrated around the base of each fold and spermatocytes line the more distal regions. Spermatids occur at the luminal surface of the germinal layer and spermatozoa fill the testicular lumen. Interstitial cells phagocytize spermatozoa and may also transfer nutrients to spermatids. The nucleus of spermatogonia is large and contains one or two nucleoli. The cytoplasm contains numerous organelles, lipid granules, and a distal and proximal centriole, each with a satellite complex. A striated rootlet extends from the distal centriole. During first meiotic prophase, the distal centriole loses its striated rootlet and produces a flagellum, the proximal centriole loses its satellite complex, the nucleolus disappears, and proacrosomal vesicles are synthesized by the Golgi complex. During spermiogenesis, most of the mitochondria appear to fuse to form a single, large mitochondrion, the nuclear chromatin condenses, and superfluous cytoplasm is lost by autophagocytosis. The formation and definitive positioning of the acrosomal vesicle and periacrosomal material at the apex of the nucleus is described in detail.     

Identification of a 195 kDa protein in the striated rootlet: its expression in ciliated and ciliogenic cells

Little is known about the molecular composition of the ciliary rootlet. We raised monoclonal antibodies to a crude preparation of striated rootlets isolated from the human oviduct, and obtained a clone (R4109) that specifically labeled the ciliary rootlets. Rootlets associated with the solitary cilium in secretory cells and fibroblasts were also labeled. R4109 identified a 195-kDa protein by immunoblotting. Ciliogenic cells in the oviduct epithelium of young mice were labeled in the globular and/or granular pattern by R4109 by immunofluorescence microscopy. Immunoelectron microscopy showed that they corresponded to fibrogranular complex and dense granule, respectively. The result demonstrated that the 195-kDa protein is a component common to the striated rootlet and dense granule, and thus suggested that dense granules are involved in the rootlet formation.     

The ultrastructure of primary cilia in the endocrine and excretory duct cells of the pancreas of mice and rats

A primary cilium was frequently observed in the endocrine alpha, beta and delta cells, as well as in the excretory duct cells of the pancreas of normal mice and rats. The characteristic components of the cilium including the basal body, axoneme (shaft), and terminal part were clearly recognizable. The basal body or distal centriole surrounded by Golgi vesicles was perpendicularly oriented to the proximal centriole, and a dense striated band was seen filling the gap between them. The microtubules of the basal body consisted of nine peripheral triplets exhibiting a 9 + 0 pattern, an appearance similar to that of the proximal centriole. Rootlets, basal feet and alar sheets associated with the basal body were occasionally seen. The axoneme usually consisted of a 9 + 0 pattern of microtubule doublets, but other irregular patterns of 7 + 2, 7 + 3, and 8 + 1 were also seen. The microtubules in the terminal part of the cilium became fewer in number and had no peculiar arrangement. The cilium of the endocrine cells always projected into the intercellular canaliculus and was covered by the ciliary sheath, and occasionally, double cilia were visualized in the vicinity of beta cells. In the excretory duct cells, the cilium showed similar features, but it was slightly longer and always projected into the dense secretory content of duct lumen. On the other hand, no primary cilium was ever observed in the acinar cells of mouse and rat pancreas. In conclusion, the present study describes the morphology of primary cilia and its associated components in the endocrine and excretory duct cells of the pancreas of mice and rats. The findings suggest that the primary cilium should be considered as a constant intracellular organelle though its function and significance remain speculative.     

Les phases précoces de la formation des cils et le problème de l'origine du corpuscule basal

Summary Developing cilia and non-ciliated centrioles are both present in the rat choroid cell studied with the electron microscope. The development of the cilium begins with the appearance of a thick-walled vesicle close to the centriole or in contact with it. Very soon this vesicle lines the distal extremity of the centriole. The growing ciliary shaft invaginates the vesicle, forming a temporary ciliary sheath. This sheath isolates the growing cilium from its environment.Degenerating ciliary shafts are by no means rare. They are interpreted as evidence of a biological cycle of the cilia. Club-shaped forms are also frequent.Non-ciliated centrioles are sometimes in contact with striated structures consisting of alternating dark and light bands, separated by a distance of 600 Å. These formations (treillis) are different from the pericentriolar satellites. On morphological grounds, they might be considered as centriole precursors.     

Fine Structure and Function of Pharynx Cilia in Glossobalanus minutus Kowalewsky (Enteropneusta)

Two kinds of cilia have been observed in the pharynx of Glossobalanus minutus Kowalewsky. From the present study, a ciliary specialization can be found in order to move a determinate substance, i.e. mucus or water. Mucus-moving cilia (type I cilia) have a single basal centriole and poorly developed ciliary rootlets. Their tips are rounded, bearing an inner, asymmetrical cap attached to some tubules. Water-moving cilia (type II cilia) are exclusively located at lateral epithelia of branchial bars, giving rise to the water current through the gills. They have two basal centrioles, proximal and distal, and a complex system of ciliary rootlets made up of a principal rootlet, a secondary or accessory rootlet and a 'fan' rootlet. The tips of type II cilia have a long process with some tubules inside. All basal structures are precisely orientated in order to assure a good coordination of ciliary beat. The possible functional significance of ciliary substructure is also discussed. From these observations a model for mucus and water currents through gill slits is postulated.     

The ventral epithelium of Trichoplax adhaerens (Placozoa): Cytoskeletal structures,cell contacts and endocytosis

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.     

Fine structure of the epidermis in Gnathostomulida

Summary The fine structure of the epidermis in Haplognathia simplex, Haplognathia rosea, Pterognathia meixneri (Filospermoidea) and Gnathostomula paradoxa (Bursovaginoidea) has been investigated. The epidermis in the filospermoidean species is uniform, consisting of epidermal cells with a single locomotory cilium. The structure and development, including ciliogenesis, of these epidermal cells are described. In G. paradoxa additional epidermal elements have been found: mucous cells with a presumably apocrine secretion modus are scattered in a strip-like arrangement within the epidermis. Their deverlopment is separate from epidermal cells with locomotory function. Two further types of glandular cells with either a single cilium or a diplosome are located ventrally. It is assumed that they represent an adhesive system.Abbreviations (used in figures) ac accessory centriole - ap appendix of accessory centriole - ax axoneme - bb basal body - bf basal foot - bl basal lamina - c cilium - cA ciliary adhesive cell - ce centriole - cp ciliary pit - d diplosome - dy dictyosome - dA diplosomal adhesive cell - E epidermal cell with locomotory cilium, epidermis - ev epidermal vesicle (epitheliosome) - gv gland vesicle - m mitochondrion - ma microvillus of apical cell membrane - mp microvillus of ciliary pit - mv microvillus - n nucleus - ps prosecretory-vesicle - R receptor - r ciliary rootlet - rc caudal ciliary rootlet - rr rostral ciliary rootlert - sv secretory vesicle - v vesicle - v _i central vesicle of multivesicular body - v _o surrounding vesicle of multivesicular body - z cisternae