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At least 19 recordsLinked to original sources
PubMed1984

Autonomous cholesterol biosynthesis in murine hepatoma. A receptor defect with normal coated pits.

These studies indicate that autonomous cholesterol biosynthesis by hepatocellular carcinoma may result from absent or defective receptors for chylomicron remnants on the surface of the malignant hepatocytes. In vivo, DAB2 hepatoma or liver were perfused with chylomicron remnants labeled with tritiated palmitic acid. Normal liver had chylomicron remnant uptake/gm tissue that was ten times that of hepatoma. In vitro studies using isolated hepatocytes and cultured DAB2 hepatoma cells showed similar results. Uptake of chylomicron remnants labeled with 3H-palmitic acid by normal hepatocytes during a 4-hour period was ten times that of hepatoma cells. Both in vivo and in vitro differences were statistically highly significant (P less than 0.005). Since many surface receptors are related to the coated pits, the cellular membranes of both neoplastic and normal liver cells were examined by electron microscopy. Coated pits were present in both the hepatoma and normal liver cells and occupied 2.61% and 2.65% of the cell surface, respectively. The defective uptake of chylomicron remnants by DAB2 hepatoma appears to be related to the chylomicron remnant receptor and not to the coated pit-internalization mechanism.

Animals↗
PubMed1988

Structural properties of the proton translocating complex of the clathrin-coated vesicle.

The clathrin-coated vesicle proton pump is a representative member of the new class of endomembrane proton ATPases that share an inhibitor profile which distinguishes them from classic F1F0 and E1E2-type proton pumps. The coated vesicle proton pump is a large (530 kDa) heteroligomer composed of eight polypeptides with molecular masses of 116, 70, 58, 40, 38, 34, 33 and 17 kDa. The 200-fold purified enzyme catalyses ATP-generated proton pumping when reconstituted in liposomes composed of pure lipids. Subunit function has been determined by partial reaction analysis of subunit and subcomplex activities. The isolated 17 kDa subunit, when co-reconstituted with bacteriorhodopsin, forms a dicyclohexylcarbodiimide-inhibitable proton channel. Selective removal of the 116 kDa subunit transforms the proton ATPase from a Mg2+-activatable to a Ca2+-activatable ATPase. Subsequent dissociation and reconstitution of subunits reveals that the 70, 58, 40 and 33 kDa components are required, in composite, to form a functional ATP-hydrolytic core, and that no single subunit or subcomplex deficient in these subunits can catalyse ATP hydrolysis.

Adenosine Triphosphate↗
PubMed1983

The distribution of actin in cultured ovarian granulosa cells.

Ovarian granulosa cells grown on glass coverslips were split by a "sandwich" technique. Using this technique we describe a complex filamentous network in the cytoplasm of cultured granulosa cells that was composed of a branching and anastomosing lattice of filaments 20-40 nm in diameter. Since filament identification is impossible on the basis of size, split cells were decorated with S-1 fragments of rabbit skeletal muscle myosin. It was readily apparent that the major constituent of the filamentous lattice was actin. Actin was organized in large bundles in which individual filaments were longitudinally aligned. Actin was also observed organized in a loose network throughout the remainder of the cytoplasm. Actin appeared to be intimately associated with organelle and plasma membranes. Coated pits were also a site of actin-filament interaction. Filament polarity was generally away from the membrane with which filaments were associated.

Actins↗
PubMed1990

Three-dimensional analysis of erythrophagosomes in rat mesenteric lymph node macrophages.

Erythrocytes extravasated into the sinuses of the rat mesenteric lymph nodes as a result of short-term clamping of the portal vein were, although autologous, phagocytized markedly by the lymph node macrophages at 1 hr after reopening of the vein. The erythrophagosomes formed in the macrophages were exposed three-dimensionally by the cellular matrix maceration method and observed with a high-resolution scanning electron microscope. These results were compared to those obtained by conventional transmission electron microscopy. The process of degradation of an erythrocyte took about 6 hr. Coated pits were formed on the erythrophagosomal membrane at the early stage, and the erythrophagosomes were degraded by two different pathways: 1) the degradative pathway by invaginations of the phagosomal membrane, through which the erythrophagosome shrank and broke into secondary lysosomes, and 2) the hemolytic degradative pathway, by which it lost its content and formed a ghost.

Animals↗
PubMed1989

Teratogenic antibodies are directed against a coated-pit glycoprotein.

It has been well established that heterologous antibodies against certain tissue components may cause congenital abnormalities when injected into pregnant rats during the critical period of organogenesis. A glycoprotein antigen (gp340) of rat renal proximal tubules was isolated (C.C.K. Leung: (J. Exp. Med., 156:372-384, 1982); antibodies against gp340 were teratogenic. Indirect colloidal gold immunocytochemical method was utilized to study the ultrastructural localization of gp340. For comparative studies, both preembedding and postembedding immunostaining procedures were used. The results indicate that gp340 is a resident of coated pits and possibly also of coated vesicles of the rat renal proximal tubules and visceral yolk-sac (VYS) endodermal cells. It appears that gp340 may also be associated with the microvilli and some as-yet-unidentified cytoplasmic structures of the same tissues. However, gp340 is absent on the epithelium of the small intestine. It is hypothesized that the teratogenic antibodies may interact with gp340 on the coated pits and interfere with receptor-mediated endocytosis, causing yolk-sac placenta dysfunction which in turn causes abnormal embryonic development.

Animals↗
PubMed1992

The mechanism of receptor-mediated endocytosis: more questions than answers.

Receptor-mediated endocytosis occurs via clathrin-coated pits and is therefore coupled to the dynamic cycle of assembly and disassembly of the coat constituents. These coat proteins comprise part, but certainly not all, of the machinery involved in the recognition of membrane receptors and their selective packaging into transport vesicles for internalization. Despite considerable knowledge about the biochemistry of coated vesicles and purified coat proteins, little is known about the mechanisms of coated pit assembly, receptor-sorting and coated vesicle formation. Cell-free assays which faithfully reconstitute these events provide powerful new tools with which to elucidate the overall mechanism of receptor-mediated endocytosis.

Adaptor Protein Complex alpha Subunits↗
PubMed1990

Clathrin-coated membrane: a distinct membrane domain in acetylcholine receptor clusters of rat myotubes.

We have used antibodies to clathrin light chains in immunocytochemical studies of acetylcholine receptor (AChR) clusters of cultured rat myotubes. Immunofluorescence and ultrastructural experiments show that clathrin is present in coated pits and in large plaques of coated membrane. Coated membrane plaques are spatially and structurally distinct from AChR-rich membrane domains and the bundles of microfilaments that are also present in AChR clusters. Clusters contain a relatively constant amount of clathrin light chain protein, which is not dependent on the amount of AChR. Clathrin plaques remain after AChR domains are disrupted by azide, or after microfilament bundles are destabilized by cytochalasin D. Extraction of myotubes with saponin removes clathrin without disrupting AChR domains. Thus, clathrin plaques, microfilament bundles, and AChR-rich domains are independently stabilized.

Animals↗
PubMed1987

Initial junctions between developing parallel fibers and Purkinje cells are different from mature synaptic junctions.

During postnatal development of cerebellar cortex, junctions are formed between parallel fiber axons and the shafts of Purkinje cell dendrites. These shaft junctions resemble synaptic junctions on spines in thin sections, in that the axon contains a cluster of synaptic vesicles, and the pre- and postjunctional membranes are lined by electron-dense material. The shaft junctions do not have the aggregate of particles arrayed on the extracellular half of the postjunctional membrane that is characteristic of mature spine synaptic junctions in freeze-fractured preparations, however, and so presumably have a different protein composition. Shaft junctions are transient specializations, present only in developing tissue, but do not appear simply to be intermediates in the formation of mature spine synaptic junctions. In normal development, spines are formed by the Purkinje cell dendrite at sites not occupied by shaft junctions. Moreover, in certain neurological mutant mice, shaft junctions form in the absence of spine synapses, and in other systems spines develop in the absence of shaft junctions. We suggest that shaft junctions are a class of synapse formed during development which is distinguished by its capacity to dissociate, and that only some fraction of the parallel fibers forming shaft synapses with a developing Purkinje cell will have established spine synapses in the adult.

Aging↗
PubMed1991

Ultrastructure of the ganglion cells of the terminal nerve in the dwarf gourami (Colisa lalia).

In our previous light microscopic studies (Oka et al., Brain Res. 367: 341-345, '86; Oka and Ichikawa, J. Comp. Neurol. 300: 511-522, '90), we reported that there are at least two types of terminal nerve (TN) cells based on cell size and immunoreactivity: type I cells had large cell bodies, while type II cells had smaller cell bodies. Type I TN cells were immunoreactive to gonadotropin-releasing hormone (GnRH) and may be the major source of GnRH-immunoreactive fibers that are widely distributed throughout the brain. Type II TN cells, on the other hand, were not immunoreactive to GnRH. In the present paper, we examined the cytology and synaptology of these two types of TN cells with electron microscopy. Type I TN cell bodies were found to have morphological characteristics similar to those of other peptide-synthesizing neurons and are likely to be actively synthesizing GnRH. The frequent occurrence of coated vesicles close to the plasma membrane of the cell body was suggestive of membrane retrieval following exocytosis of the vesicular contents from the cell surface. Neighboring TN cells were either in direct juxtaposition with one another or made specialized "glomeruloid" cell-to-cell contacts; these specializations may be relevant for nonsynaptic intercellular communications among the TN cells. Within these glomeruloid complexes, the somatic processes of TN cells received inputs from two types of synaptic terminals: one containing only spherical synaptic vesicles and another containing a small number of dense-cored vesicles in addition to the spherical synaptic vesicles. Axosomatic synapses were rare on type I TN cell bodies. In contrast, type II TN cell bodies had morphological characteristics similar to those of neurons in other brain regions. These receive axosomatic inputs from synaptic terminals containing only spherical synaptic vesicles and those with a small number of dense-cored vesicles in addition to the spherical synaptic vesicles. Thus, each type of TN cell has unique fine structural characteristics which may correlate to their different functional roles.

Animals↗
PubMed1986

Intracellular pathway of interleukin 2 following receptor-mediated endocytosis.

Electron microscope autoradiography was used to examine the intracellular pathway of radioiodinated interleukin 2 [( 125I]IL2) following its receptor-mediated endocytosis in CTLL cells. Direct measurement of the kinetics of endocytosis showed that 60-70% of surface-bound [125I]IL2 was internalized after 1 h at 37 degrees C. [125I]IL2 was observed to enter cells through invagination of coated pits at the cell surface and to concentrate into lysosomal multivesicular bodies as early as 5-10 min after endocytosis. These results provide ultrastructural information on the intracellular pathway of IL2 and on its probable site of degradation within the cell.

Biological Transport↗
PubMed1982

Transferrin receptor and its recycling in HeLa cells.

The transferrin receptor is a 180 000-dalton protein which can be dissociated to two 90 000-dalton polypeptides under reducing conditions. It can be labelled by lactoperoxidase-catalysed iodination on the cell surface at 0 degree C. Trypsin digestion of labelled cells at 0 degree C can be used to degrade those receptors on the cell surface; they release a 70 000-dalton soluble fragment which binds to transferrin. When cells are labelled at 0 degree C, then warmed to 37 degrees C, the labelled receptors enter the cells and become trypsin resistant. These receptors enter the cells, probably via coated pits, with a half-life of approximately 5 min. Since there is about three times as much receptor inside cells as on the surface, this means that transit through the cell to the cell surface takes approximately 21 min, if all receptors are on the same cycling pathway.

Coated Pits, Cell-Membrane↗
PubMed1983

Distribution of ferritin receptors and coated pits on giant HeLa cells.

HeLa cells bind horse spleen ferritin when the two are incubated at 0 degrees C. Since the majority of this bound ferritin is located in coated pits, we conclude that the ferritin binds to a specific receptor which takes part in an endocytic cycle. When substrate-attached and well-spread giant HeLa cells are briefly labelled at 0 degrees C with ferritin, ferritin particles are found to be concentrated towards the cell periphery, where they exist largely outside coated pits. This peripheral concentration is a property of circulating (and not just newly synthesized) receptors because it is not affected by prior incubation of giant cells in cycloheximide. However, coated pits are themselves roughly uniformly distributed over the surface of these cells. These results provide evidence that the membrane internalised by coated pits on these cells is returned to the cell surface at the leading edge of the cell. Because of this separation of the sites of endocytosis and exocytosis, a flow of membrane must occur across the cell surface. This flow is composed of lipid plus receptors. The implications of this for capping and for cell spreading are discussed.

Biological Transport, Active↗
PubMed1984

Purification and properties of 100-kd proteins from coated vesicles and their reconstitution with clathrin.

Bullock brain coated vesicles contain a family of at least six 100-kd polypeptides which have the property of promoting clathrin assembly. These proteins have been purified from Triton X-100-extracted coated vesicles by a combination of gel filtration and chromatography on hydroxylapatite and DE-52 cellulose. Three major 100-kd species occur as complexes with a stoichiometric amount of a 50-kd polypeptide. On cross-linking these complexes, the chief products appear to contain two polypeptides of 100 kd and two of 50 kd. These 100-kd/50-kd complexes will polymerise with low concentrations of clathrin to give a relatively homogeneous population of coats predominantly of the 'barrel' size. In contrast, three other polypeptides of 100 kd lack the 50-kd protein but polymerise with clathrin under the same conditions to yield coats of a wide range of sizes including 'barrels', truncated icosahedra and particles of greater than 100 nm diameter. When clathrin cages are reassembled with a saturating amount of 100-kd/50-kd complexes and studied by electron microscopy, the additional proteins appear to follow the underlying geometry of the clathrin polyhedra, partially filling in the polygonal faces of the cage structures. Saturation appears to require approximately 3 molecules of 100-kd polypeptide per clathrin trimer.

Animals↗