Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “GOLGI APPARATUS”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 847 records · Page 47Linked to original sources
PubMed1997

Rotavirus is released from the apical surface of cultured human intestinal cells through nonconventional vesicular transport that bypasses the Golgi apparatus.

Rotaviruses are nonenveloped viruses that infect enterocytes of the small intestine and cause severe infantile gastroenteritis. It was previously thought that rotavirus exits cells by lysis, but this behavior does not match the local pathogenesis of the virus. In this study, we have investigated the release of the simian rotavirus strain (RRV) from the polarized intestinal Caco-2 cells. We found that RRV is released almost exclusively from the apical pole of Caco-2 cells before any cells lyse. Using confocal laser scanning microscopy and drugs that inhibit vesicular transport, we studied the RRV transport route from the endoplasmic reticulum (ER) to the apical side of intestinal cells. We demonstrated that RRV exits from the ER through a carbonyl cyanide m-chlorophenylhydrazone-sensitive vesicular transport. RRV staining was never found within the Golgi apparatus or lysosomes, suggesting that the RRV intracellular pathway does not involve these organelles. This finding was confirmed by treatment with monensin or NH4Cl, which do not affect release of RRV. Electron microscopic analysis revealed RRV containing small smooth vesicles in the apical area and free virions outside the cell in the brush border, consistent with a vesicular vectorial transport of virus. These results may provide, for the first time, a cellular explanation of the pathogenesis of rotavirus.

Biological Transport↗
PubMed2003

Suggested role of the Golgi apparatus and endoplasmic reticulum for crucial sites of hepatitis C virus replication in human lymphoblastoid cells infected in vitro.

Iacovacci et al. [(1997a) Research in Virology 148:147-151] described that the euploid diploid cells, of the normal human bone marrow-derived lymphoblastoid B-cell line TO.FE., are susceptible to hepatitis C virus (HCV) infection and support long term virus production. Transmission electron microscopy described some steps of HCV replication cycle in this in vitro infected cellular system [Serafino et al. (1997) Research in Virology 148:153-159]. In the present study, in order to identify the intracellular sites involved in HCV replication, the ultrastructural changes associated with infection in TO.FE. cells were correlated with the subcellular localisation of structural and nonstructural viral proteins. Transmission electron microscopy and confocal microscopy data indicate that these viral proteins appeared located in the Golgi apparatus and endoplasmic reticulum, suggesting an active involvement of these compartments in viral assembly and morphogenesis. Furthermore, transmission and scanning electron microscopic observations on cultures infected chronically support the hypothesis that these cellular compartments may serve as starting sites of the morphological changes associated to viral infection and replication, leading to cell-cell fusion, syncytia formation, and finally lysis of infected cells and virus release.

B-Lymphocytes↗
PubMed1987

Codistribution of galactosyl- and sialyltransferase: reorganization of trans Golgi apparatus elements in hepatocytes in intact liver and cell culture.

The intracellular distribution of galactosyl- and sialyltransferase was investigated in rat hepatocytes of intact liver, primary monolayer cultures of freshly isolated hepatocytes, in a nontumorigenic hepatocyte cell line and in a hepatoma cell line. The two glycosyltransferases were detected by immunofluorescence using affinity-purified rabbit antibodies. Indirect double immunofluorescence showed that both terminal glycosyltransferases were identically codistributed in the same cell. This codistribution was always observed regardless of the cell type investigated, and in both stationary and migrating cells. The immunofluorescence pattern for both galactosyl- and sialyltransferase was found to be different in hepatocytes in vivo compared to hepatocytes grown in vitro. In hepatocytes of intact liver a spot-like cytoplasmic fluorescence was observed, whereas in cultured normal hepatocytes a perinuclear fluorescence from which an extensive tubular network radiated far into the cytoplasm existed. Cultured hepatoma cells also exhibited an extensive cytoplasmic fluorescence, which in contrast to the normal hepatocytes was rather diffuse. We conclude that (a) galactosyl- and sialyltransferase are codistributed in rat hepatocytes, and (b) a reorganization of (trans) Golgi apparatus elements containing both terminal glycosyltransferases occurs under conditions of in vitro growth and malignant transformation.

Animals↗
PubMed1984

A pathway of plasma membrane biogenesis bypassing the Golgi apparatus during cell division in the green alga Cylindrocapsa geminella.

Cell division in Cylindrocapsa geminella, in particular the mode of septum membrane biogenesis, has been studied with the transmission electron microscope. Septum formation takes place in a narrow layer of cytoplasm separating post-mitotic nuclei. First, each daughter nucleus develops a wide cytoplasmic pocket (invagination) containing numerous strands of rough endoplasmic reticulum (ER). Next, a proliferation of rough ER is observed in the equatorial zone of cytoplasm, which invariably contains a small number of widely scattered microtubules. The equatorially aligned cisternae of rough ER produce smooth-membraned vesicles, interpreted as smooth ER, which subsequently coalesce to form the membranous transverse septum. Thus, primary septum formation does not follow any of the two previously known basic cytokinetic patterns in green plants (i.e. plasma membrane furrowing and cell-plate formation), but instead represents a novel type of membrane flow, which effectively bypasses the Golgi apparatus. This pathway of membrane flow has remained largely ignored in current concepts of endomembrane structure and function in eukaryotes. However, it appears to be more widespread than has previously been recognized, especially in autospore-producing green algae and in red algae during the formation of tetraspores. It may represent an evolutionary intermediate type of cell division between the supposedly primitive method of plasma membrane furrowing and the more advanced cell-plate system.

Cell Division↗
PubMed2002

Altered sphingolipid metabolism in multidrug-resistant ovarian cancer cells is due to uncoupling of glycolipid biosynthesis in the Golgi apparatus.

Multidrug-resistant tumor cells display enhanced levels of glucosylceramide. In this study, we investigated how this relates to the overall sphingolipid composition of multidrug-resistant ovarian carcinoma cells and which mechanisms are responsible for adapted sphingolipid metabolism. We found in multidrug-resistant cells substantially lower levels of lactosylceramide and gangliosides in sharp contrast to glucosylceramide, galactosylceramide, and sphingomyelin levels. This indicates a block in the glycolipid biosynthetic pathway at the level of lactosylceramide formation, with concomitant accumulation of glucosylceramide. A series of observations exclude regulation at the enzyme level as the underlying mechanism. First, reduced lactosylceramide formation occurred only in intact resistant cells whereas cell-free activity of lactosylceramide synthase was higher compared with the parental cells. Second, the level of lactosylceramide synthase gene expression was equal in both phenotypes. Third, glucosylceramide synthase (mRNA and protein) expression and activity were equal or lower in resistant cells. Based on the kinetics of sphingolipid metabolism, the observation that brefeldin A does not restore lactosylceramide synthesis, and altered localization of lactosylceramide synthase fused to green fluorescent protein, we conclude that lactosylceramide biosynthesis is highly uncoupled from glucosylceramide biosynthesis in the Golgi apparatus of resistant cells.

ATP Binding Cassette Transporter, Subfamily B↗
PubMed1981

Development of the cell wall in Tetraselmis: role of the Golgi apparatus and extracellular wall assembly.

The green algal flagellate, Tetraselmis, is a key transition organism in the phylogeny of green algae. It has been proposed that the cell wall of Tetraselmis arose evolutionarily from the fusion of scales and that this event secondarily caused the alteration of some cytoplasmic processes such as mitotic and cytokinetic mechanisms. Ultrastructural and developmental studies of the cell wall were performed with several strains of Tetraselmis. Two major wall types are reported. The wall of type 1 cells consists of a thick inner region covered by a layer of regularly repeating subunits of 26 nm, comparable to the subunits found in the median W2-W6 layer of Chlamydomonas. The more elaborate type 2 cell wall consists of a thick median wall layer, homologous to the type 1 inner wall, with additional inner and outer strata of hairs, grains and scales. Development of the cell wall begins in the endomembrane system, particularly the Golgi apparatus, where fibrillar tufts and electron-dense droplets are synthesized, modified and transported to the outside. Here, the tufts and droplets are displaced around the protoplast and assemble in several steps to yield the intact wall. Edge-growth assembly of the wall occurs here synchronously with cytoplasmic developments to yield the characteristic anterior flagellar pit. Models explaining various aspects of this development are discussed. When released from the cell, the wall subunits are not completely comparable to stellate scales, but appear to correspond to developmental stages of scales in green flagellates possessing body scales.

Biological Evolution↗
PubMed1992

Tetrahymena gene encodes a protein that is homologous with the liver-specific F-antigen and associated with membranes of the Golgi apparatus and transport vesicles.

The F-antigen is a prominent liver protein which has been extensively used in studies on natural and induced immunological tolerance. However, its intracellular localization and biological function have remained elusive. It has generally been assumed that the F-antigen is confined phylogenetically to vertebrates. Now we have cloned and characterized a gene from the ciliated protozoan Tetrahymena thermophila encoding a protein which clearly is homologous with the rat F-antigen. The coding region of the Tetrahymena F-antigen (TF-ag) gene specifies a 46,051 M(r) protein and is interrupted by three introns. In accordance with the predicted molecular mass of the TF-ag protein, antibodies raised against a cro-lacZ'-TF-ag fusion protein specifically recognized a 45,000 M(r) protein in Western blots of total T. thermophila protein. Immunoelectron microscopy demonstrated that the TF-ag is associated with membranes of the Golgi apparatus and transport vesicles pointing to a role of TF-ag in membrane trafficking. Transcription of the TF-ag gene, as determined by run-on analyses, was only detectable in growing cells, and following transfer to starvation condition pre-existing TF-ag mRNA was rapidly degraded. The abundance of the TF-ag protein, however, declined only moderately during prolonged periods of starvation demonstrating that extensive release of the TF-ag did not take place. In combination these results suggest that the TF-ag protein is a recycled constituent of the intracellular membrane network in T. thermophila.

Amino Acid Sequence↗
PubMed2002

Intracellular localization of the peanut clump virus replication complex in tobacco BY-2 protoplasts containing green fluorescent protein-labeled endoplasmic reticulum or Golgi apparatus.

RNA-1 of Peanut clump virus (PCV) encodes the proteins P131 and P191, containing the signature motifs of replication proteins, and P15, which regulates viral RNA accumulation. In PCV-infected protoplasts both P131 and P191 were immunodetected in the perinuclear region. Laser scanning confocal microscopy (LSCM) showed that P131 and P191 colocalized with neosynthesized 5-bromouridine 5'-triphosphate-labeled RNA and double-stranded RNA, demonstrating that they belong to the replication complex. On the contrary, the P15 fused to the enhanced green fluorescent protein (EGFP) never colocalized with the two proteins. In endoplasmic reticulum (ER)-GFP transgenic BY-2 protoplasts, the distribution of the green fluorescent-labeled ER was strongly modified by PCV infection. LSCM showed that both P131 and P191 colocalized with ER green fluorescent bodies accumulating around the nucleus during infection. The replication process was not inhibited by cerulenin and brefeldin A, suggesting that PCV replication does not depend on de novo-synthesized membrane and does not require transport through the Golgi apparatus. Electron microscopy of ultrathin sections of infected protoplasts showed aggregates of broken ER but also visualized vesicles, some of which resembled modified peroxisomes. The results suggest that accumulation of PCV during infection is accompanied by specific association of PCV RNA-1-encoded proteins with membranes of the ER and other organelles. The concomitant extensive rearrangement of these membranous structures leads to the formation of intracellular compartments in which synthesis and accumulation of the viral RNA occur in defined areas.

Arachis↗
PubMed1996

Metabolism of the "Swedish" amyloid precursor protein variant in neuro2a (N2a) cells. Evidence that cleavage at the "beta-secretase" site occurs in the golgi apparatus.

The 4-kDa beta-amyloid peptide (Abeta), a principal component of parenchymal amyloid deposits in Alzheimer's disease, is derived from amyloid precursor proteins (APP). To identify potential intracellular compartments involved in Abeta production, we expressed human APP-695 (APPwt) and APP-695 harboring the Swedish double mutation (APPswe) associated with familial early-onset Alzheimer's disease, in mouse N2a cells. We demonstrate that cells expressing APPswe secrete high levels of Abeta peptides and beta-secretase-generated soluble APP derivatives (APP s beta) relative to cells expressing APPwt. In addition, we observed a concomitant diminution in the levels of alpha-secretase-generated soluble APP derivatives (APP s alpha). Our interpretation of these findings is that beta-secretase cleavage occurs in an intracellular compartment and disables those substrates which would normally be cleaved by alpha-secretase. As anticipated, the levels of APPswe are diminished relative to the steady-state levels of surface-bound APPwt; moreover, surface-bound APPswe and APPwt molecules are released from the plasma membrane after cleavage by alpha-secretase, but not by beta-secretase. Finally, by examining the rate of appearance of specific APP metabolites generated by beta-secretase, we now unequivocally demonstrate that beta-secretase cleavage of APPswe occurs within the Golgi apparatus, as early as the medial compartment.

Alzheimer Disease↗
PubMed1988

Immunoperoxidase localization of bile salts in rat liver cells. Evidence for a role of the Golgi apparatus in bile salt transport.

The mechanisms of intracellular transport of bile acids from the sinusoidal pole to the canalicular pole of the hepatocyte are poorly understood. There is physiological and autoradiographic evidence for a vesicular pathway. The purpose of this study was to determine the localization of natural bile acids in the liver using antibodies against cholic acid conjugates and ursodeoxycholic acid. An indirect immunoperoxidase technique was used on rat liver sections fixed either with paraformaldehyde (PF) and saponin, a membrane-permeabilizing agent that allows penetration of antibodies into the cell, or with PF alone. Retention of taurocholate in the liver after tissue processing was 26 +/- SD 15% of the bile acid initially present. When sections fixed with PF and saponin were incubated with the antibody against cholic acid conjugates, a granular cytoplasmic staining was observed by light microscopy in all hepatocytes. By electron microscopy, strong electron-dense deposits were observed mostly on vesicles of the Golgi apparatus (GA) and, sometimes, in the smooth endoplasmic reticulum (SER). After taurocholate infusion, the intensity of the reaction increased. When the liver was fixed with PF alone, almost no reaction was visible on light microscopy, but on electron microscopy the label was localized on the hepatocyte plasma membrane, mainly on the bile canalicular domain and to a lesser extent on the sinusoidal domain. With the antibody against ursodeoxycholic acid, no staining was observed in three of four livers, and a slight staining was observed in one. However, after infusion of ursodeoxycholic acid, staining of GA and SER vesicles was observed when the liver was fixed with PF and saponin. With PF alone, the reaction was intense on the canalicular membrane. These results support the view that, within the limits of the method, vesicles from the GA and possibly vesicles of the SER are involved in the intracellular transport of bile acids before canalicular secretion.

Animals↗
PubMed1988

Association of neuroactive peptides with the protein secretory pathway in identified neurons of Aplysia californica: immunolocalization of SCPA and SCPB to the contents of dense-core vesicles and the trans face of the Golgi apparatus.

The subcellular distribution of two molluscan neuropeptides, the small cardioactive peptides A and B (SCPA and SCPB), has been determined in two identified Aplysia buccal ganglion neurons, B1 and B2. These neurons were previously shown to synthesize and release these neuropeptides. B1 and B2, identified by their size and location within the ganglion, were labeled by intrasomatic injection of an electron-dense particulate marker (ferritin or Imposil) permitting the unequivocal identification of their somata and proximal processes in thin sections. The somatic cytoplasm of both neurons had a conspicuous population of large dense-core vesicles along with a smaller number of compound vesicles and small lucent vesicles. All three vesicle types are found in the neurites within the neuropil and proximal axon in the esophageal nerve. Immunoreactivity was localized on the surface of thin sections by the indirect immunogold method. The primary antiserum was shown to recognize both SCPA and SCPB after the neuropeptides had been immobilized on protein-coated nitrocellulose membranes by means of glutaraldehyde, the primary fixative used to immobilize SCPA and SCPB in situ. SCP immunoreactivity was present in the lumens of the dense-core vesicles distributed throughout the cytoplasm of B1 and B2 and in dense-core regions of the Golgi apparatus in the somatic cytoplasm. Taken together with biochemical evidence that B1 and B2 synthesize and release SCPs, these data suggest that the neuropeptides are sequestered into the protein secretory pathway of B1 and B2, a distribution that supports the notion that the SCPs function physiologically as neurotransmitters or neuromodulators.

Animals↗
PubMed2005

Comparison of Golgi apparatus and endoplasmic reticulum proteins from livers of juvenile and aged rats using a novel technique for separation and enrichment of organelles.

The broad dynamic range of protein abundances, which can vary from about 10(6) for cells to 10(10) for tissues in complex proteomes, continues to challenge proteomics research. Proteome analysis, in particular organelle proteomics, using current approaches, requires extensive fractionation, separation, and enrichment. Over the years, organelle separation was achieved through the use of differential and density-gradient ultracentrifugation. However, the traditional fixed-volume process is a time-consuming and labor-intensive method, especially with large quantities of sample. Here, we present a novel tool for subcellular fractionation of biologically complex mixtures: continuous-flow ultracentrifugation of tissue homogenates to obtain both organelle separation and extensive organelle enrichment at the same time. In this study, rat liver tissues from two different age groups (3-8 wk and greater than 1 y old) were homogenized by blending. After removing nuclei, the resulting homogenates were further fractionated at the subcellular level by the use of sucrose gradient continuous-flow ultracentrifugation. Each organelle's enriched fractions were identified by Western blot analysis. To study the possible effects of aging on the endoplasmic reticulum and Golgi apparatus, we compared the organelle protein profiles of the two groups of rat liver tissues using two-dimensional gel electrophoresis, digitized imaging of two-dimensional gel electrophoresis, and mass spectrometry. Significant differences in the protein profiles of both organelles were observed between the two groups of rat tissues. The technique described here for fractionation and enrichment of organelles demonstrated a useful tool for proteomics research, including identification of low-abundance proteins and post-translational modifications.

Aging↗
PubMed2006

Presenilin mutations linked to familial Alzheimer's disease reduce endoplasmic reticulum and Golgi apparatus calcium levels.

Presenilin-1 and -2 (PS1 and PS2) mutations, the major cause of familial Alzheimer's disease (FAD), have been causally implicated in the pathogenesis of neuronal cell death through a perturbation of cellular Ca(2+) homeostasis. We have recently shown that, at variance with previous suggestions obtained in cells expressing other FAD-linked PS mutations, PS2-M239I and PS2-T122R cause a reduction and not an increase in cytosolic Ca(2+) rises induced by Ca(2+) release from stores. In this contribution we have used different cell models: human fibroblasts from controls and FAD patients, cell lines (SH-SY5Y, HeLa, HEK293, MEFs) and rat primary neurons expressing a number of PS mutations, e.g. P117L, M146L, L286V, and A246E in PS1 and M239I, T122R, and N141I in PS2. The effects of FAD-linked PS mutations on cytosolic Ca(2+) changes have been monitored either by using fura-2 or recombinant cytosolic aequorin as the probe. Independently of the cell model or the employed probe, the cytosolic Ca(2+) increases, caused by agonist stimulation or full store depletion by drug treatment, were reduced or unchanged in cells expressing the PS mutations. Using aequorins, targeted to the endoplasmic reticulum or the Golgi apparatus, we here show that FAD-linked PS mutants lower the Ca(2+) content of intracellular stores. The phenomenon was most prominent in cells expressing PS2 mutants, and was observed also in cells expressing the non-pathogenic, "loss-of-function" PS2-D366A mutation. Taken as a whole, our findings, while confirming the capability of presenilins to modify Ca(2+) homeostasis, suggest a re-evaluation of the "Ca(2+) overload" hypothesis in AD and a new working hypothesis is presented.

Adult↗
PubMed1992

Brefeldin A-resistant mutants of human epidermoid carcinoma cell line with structural changes of the Golgi apparatus.

We have isolated brefeldin A (BFA)-resistant cell lines, KB/BF-1 and KB/BF-2, from the human epidermoid carcinoma KB cell line. The BFA-resistant phenotypes have been stably maintained for more than 3 months in the absence of BFA. KB/BF-1 and KB/BF-2 showed 10-30-fold higher resistance to cytotoxicity of BFA but were 2-3-fold more sensitive to monensin and nigericin, than KB cells. KB/BF-1 showed aberrant structures of the Golgi complex with poorly developed cisternae surrounded by many small vesicles. Immunocytochemical studies were done with antibodies against a Golgi-specific antigen (chronic rheumatoid arthritis antigen) and a coatomer subunit (beta-subunit for coat proteins of non-clathrin-coated vesicles). Golgi-specific markers were distributed into the small vesicles which were localized diffusedly in cytoplasm of KB/BF-1 cells. Such Golgi markers were observed in a strictly confined perinuclear region of the parental KB cells, whereas in the mutant cells the markers were distributed more diffusedly in dot-like structures at perinuclear regions. In addition, when exposed to BFA, the mutant and parental cells showed a different distribution of these markers. Synthesis and maturation of low density lipoprotein receptor showed apparently slower rates in processing of low density lipoprotein receptor in KB/BF-1 and KB/BF-2 cells than those observed in their parental KB cells. Protein secretion in KB/BF-1 and KB/BF-2 cells was about 30% less than that in KB cells. Much less inhibition by BFA on the secretion was observed in KB/BF-1 and KB/BF-2 cells. A BFA-resistant mutation in BFA-resistant KB cell lines appears to affect assembly of the Golgi apparatus as well as some Golgi-specific functions.

Brefeldin A↗