Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0038362 (stomatitis)
 8,852 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Membrane bound polysomes were prepared from HeLa cells infected with vesicular stomatitis virus (VSV), after pulse labeling with [3H]mannose for various times from 15 to 90 min. Oligosaccharides on nascent chains were released from peptides by treatment with endoglycosidase H and sized by high resolution Biogel P4 chromatography. Processing on some nascent chains proceeded to the removal of all three types of alpha-linked glucose and one alpha-1,2-mannose from the Glc3Man9GlcNAc precursor showing that the enzymes responsible were not only active on nascent chains but were present in the rough endoplasmic reticulum (RER). Incubation of cells for various times in cycloheximide, where chain elongation had ceased, made no difference to the profile of oligosaccharides on the nascent chains, and trimming proceeded no further than Man8GlcNAc2Asn . Carbonyl cyanide m-chlorophenylhydrazone (CCCP), an energy inhibitor reportedly able to block the transfer of glycoproteins from the RER, increases the amount of Man8-oligosaccharides on the nascent chains and also the amount of Glc3Man9GlcNAc precursor. On completed G protein in the RER fraction from which membrane bound polysomes were prepared, processing occurred to Man6 - but not to Man5GlcNAc sized oligosaccharides in the CCCP-treated cells. By contrast, processing to Man5GlcNAc oligosaccharides was observed in unfractionated control cells.
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PMID:Co-translational excision of alpha-glucose and alpha-mannose in nascent vesicular stomatitis virus G protein. 632 29

We have altered the structure of the COOH-terminus of the vesicular stomatitis virus (VSV) glycoprotein (G) by introducing deletions into a cDNA clone encoding G protein. We examined the effects of these deletions on intracellular transport of G protein after expression of the deleted genes in eucaryotic cells under control of the SV40 late promoter. To prevent readthrough of translation into vector sequences, we introduced synthetic DNA linkers containing translation stop codons at the site of the deletion. G proteins that lacked the cytoplasmic domain and most of the transmembrane domain were secreted slowly from the cells. Deletion mutants affecting the structure of the cytoplasmic domain fell into two classes. The first class completely arrested transport of the protein to the cell surface at a stage prior to acquisition of complex oligosaccharides. The second class showed severely reduced rates of complex sugar addition although the proteins were eventually transported to the cell surface. Indirect immunofluorescence microscopy suggested that mutant proteins in both classes may accumulate in the rough endoplasmic reticulum.
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PMID:Altered cytoplasmic domains affect intracellular transport of the vesicular stomatitis virus glycoprotein. 635 53

We have characterized two stable transformed mouse cell lines (CG1 and CTG1) that express either the normal vesicular stomatitis virus glycoprotein (G) or a truncated form of the G protein (TG) that lacks the COOH-terminal anchor sequences and is secreted from the cells. These cell lines were obtained using a hybrid vector consisting of the transforming DNA fragment of bovine papilloma virus linked to a segment of the SV40 expression vector pSV2 containing cloned cDNA encoding either the normal or truncated form of the vesicular stomatitis virus G protein. Using indirect immunofluorescence we have found that greater than 95% of the cells in each line express the G protein(s), although the level of expression within the population is variable. The normal G protein expressed in these cells obtains its complex oligosaccharides in less than 30 min and is transported to the cell surface. In contrast, the TG protein obtains its complex oligosaccharides with a half-time of about 2.5 h. Immunofluorescence data show an apparent concentration of the TG protein in the rough endoplasmic reticulum. These data together suggest that transfer of this anchorless protein from the rough endoplasmic reticulum to the Golgi apparatus is the rate-limiting step in its secretion. We observed, in addition to normal G protein, two smaller G-related proteins produced in the CG1 cell line. We suggest that these proteins could result from aberrant splicing from sites within the G mRNA sequence to the downstream acceptor in the pSV2 vector.
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PMID:Isolation of stable mouse cell lines that express cell surface and secreted forms of the vesicular stomatitis virus glycoprotein. 641 65

Mixed monolayers containing vesicular stomatitis virus-infected Chinese hamster ovary clone 15B cells (lacking UDP-N-acetylglucosamine transferase I, a Golgi enzyme) and uninfected wild-type Chinese hamster ovary cells were formed. Extensive cell fusion occurs after the monolayer is exposed to a pH of 5.0. The vesicular stomatitis virus encoded membrane glycoprotein (G protein) resident in the rough endoplasmic reticulum (labeled with [35S]methionine) or Golgi complex (labeled with [3H]palmitate) of 15B cells at the time of fusion can reach Golgi complexes from wild-type cells after fusion; G protein present in the plasma membrane cannot. Transfer to wild-type Golgi complexes is monitored by the conversion of G protein to an endoglycosidase H-resistant form upon arrival, and also demonstrated by immunofluorescence microscopy. G protein in the Golgi complex of the 15B cells at the time of fusion exhibits properties vis a vis its transfer to an exogenous Golgi population identical to those found earlier in a cell-free system (Fries, E., and J. E. Rothman. 1981. J. Cell Biol., 90: 697-704). Specifically, pulse-chase experiments using the in vivo fusion and in vitro assays reveal the same two populations of G protein in the Golgi complex. The first population, consisting of G protein molecules that have just received their fatty acid, can transfer to a second Golgi population in vivo and in vitro. The second population, entered by G protein approximately 5 min after its acylation, is unavailable for this transfer, in vivo and in vitro. Presumably, this second population consists of those G-protein molecules that had already been transferred between compartments within the 15B Golgi population, in an equivalent process before cell fusion or homogenization for in vitro assays. Evidently, the same compartment boundary in the Golgi complex is detected by these two measurements. The surprisingly facile process of glycoprotein transit between Golgi stacks that occurs in vivo may therefore be retained in vitro, providing a basis for the cell-free system.
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PMID:Transport of protein between cytoplasmic membranes of fused cells: correspondence to processes reconstituted in a cell-free system. 642 57

The biosynthesis of the erythrocyte anion transport glycoprotein, Band III (Mr 100,000), is of interest, as its N-terminal half is hydrophilic and faces the cytoplasmic surface; the C-terminal half spans the phospholipid bilayer several times. Band III is synthesized by erythroid precursor cells obtained from the spleens of anaemic mice. Newly synthesized Band III was inserted into rough endoplasmic reticulum membranes with an asymmetric orientation which resembled that of mature Band III in erythrocyte membranes: the N-terminal portion of the molecule facing the cytoplasm. Newly made Band III contained a high-mannose asparagine-linked oligosaccharide, which was susceptible to cleavage by endoglycosidase H. During the next 20-30 min, this oligosaccharide was processed to a form resistant to endoglycosidase H degradation, presumably in the Golgi complex. The processed Band III was subsequently expressed on the cell surface, at about 30-45 min after synthesis. To study the mechanism of insertion of Band III into microsomes, we used erythroid precursor cells from the spleens of anaemic mice as a source of messenger RNA for studies in vitro in the wheat germ and reticulocyte lysate cell-free system containing dog pancreatic microsomes. Immediately after synthesis, Band III was found to be inserted into microsomal membranes in its mature configuration, with the N-terminal portion exposed to the cytoplasm and its hydrophobic C-terminal portion spanning the lipid bilayer. The newly-synthesized Band III was also provided with a high-mannose asparagine-linked oligosaccharide. Band III was found to be inserted into dog pancreatic microsomes in a co-translational manner; in synchronized translation studies microsomes could be added as late as the time when the hydrophilic N-terminal half of the protein had been synthesized and still allow normal trans-membrane insertion and glycosylation. There is no cleavage of any N-terminal peptide during membrane insertion. In many respects, therefore, the biosynthesis of Band III resembles that of co-translationally-inserted proteins whose N-terminal portions are exposed on the exterior of the cell, like vesicular stomatitis virus glycoprotein, HLA-A antigens, and glycophorin. However, our results suggest that Band III contains a sequence near the middle of the protein which directs its insertion into endoplasmic reticulum membranes.
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PMID:Synthesis and maturation of the erythrocyte anion transport protein--an internal sequence for membrane insertion. 682 84

In eukaryotic cells, secretory proteins and glycoproteins migrate from the rough endoplasmic reticulum, their site of synthesis, through Golgi vesicles before being released from the cell. Cellular and viral integral plasma membrane glycoproteins are co-translationally inserted into the rough endoplasmic reticulum membrane and follow a similar pathway to the cell surface. Previous studies using endoglycosidase H (Endo H) suggested that in rat hepatoma cells the vesicular stomatitis virus (VSV) G protein, albumin and transferrin migrate from the rough endoplasmic reticulum to the Golgi apparatus at different rates. Here we show directly that in human hepatoma HepG2 cells, five secreted proteins mature from the rough endoplasmic reticulum to Golgi vesicles at characteristic rates which differ at least threefold. The results are incompatible with bulk-phase movement of the luminal contents of the endoplasmic reticulum, and suggest that there is a membrane-bound receptor that selectively mediates the transport of secretory proteins from the rough endoplasmic reticulum to the Golgi.
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PMID:Hepatoma secretory proteins migrate from rough endoplasmic reticulum to Golgi at characteristic rates. 686 94

HIV-1 Vpu is a small transmembrane phosphoprotein of 16 kDa which performs critical roles in CD4 proteolysis and virus release. Previous studies have demonstrated that Vpu-induced degradation of CD4 occurs in the endoplasmic reticulum (ER), and that the proteolytic process is sequence specific requiring both the transmembrane and cytoplasmic domains of CD4. In the present study, we investigated the effects of Vpu expression on the intracellular membrane trafficking pathway of mammalian cells. In singly transfected cells, the HIV envelope glycoproteins and vesicular stomatitis virus glycoprotein (VSV G) were properly transported to the cell surface undergoing oligosaccharide modifications characteristic of their movement through the Golgi complex. In contrast, the cell surface delivery of glycoproteins was severely impeded in cells expressing Vpu. Biochemical analyses revealed that Vpu expression blocked the transfer of proteins from the ER-Golgi complex to the plasma membrane in a dose- and protein-dependent manner. Soluble gp120 exhibited extreme transport defects in the presence of Vpu, whereas transmembrane proteins (e.g., gp160, VSV) responded only moderately to wild-type Vpu. To gain insight into Vpu-mediated transport inhibition, we performed mutational analysis of the CK-2 phosphorylation sites (serines at 52 and 56) in the Vpu protein. CK-2 phosphorylation of Vpu has been shown to regulate the activity of the protein in reactions that involve the proteolysis of CD4 in the ER. We demonstrate here that the phosphorylation mutant is defective in both sequence-specific degradation of VRE-containing substrates and the transport inhibition of gp120 and VSV-G in the secretory pathway. Thus, these experiments have revealed that Vpu-mediated proteolysis and transport inhibition are mechanistically coupled requiring the same structural elements of the Vpu protein in both processes. We propose that the primary effect of Vpu expression is to impede the secretion process and then access glycoproteins bearing the VRE for Vpu-mediated proteolysis in the ER of mammalian cells.
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PMID:The human immunodeficiency virus type 1 Vpu protein: a potential regulator of proteolysis and protein transport in the mammalian secretory pathway. 749 87

The export of vesicular stomatitis virus glycoprotein (VSV-G) from the endoplasmic reticulum (ER) involves sorting and concentration, and has been proposed to require the function of heterotrimeric G proteins. To begin to identify the basic elements of a potential signaling pathway involved in vesicle assembly, we have examined whether protein kinase C (PKC) is required for ER to Golgi transport. Calphostin C, a specific inhibitor of the highly conserved cysteine-rich C6H2 motif present in the regulatory domain of PKC was found to be a potent inhibitor of export of VSV-G and vesicle budding from the ER in vivo and in vitro (IC50 approximately 60 nM). In contrast, the diacylglycerol analog phorbol 12-myristate 13-acetate, which activates PKC, enhanced the migration of VSV-G from the ER to pre-Golgi intermediates. Neither reagent had detectable effects on the oligomerization of VSV-G prior to export nor perturbed transport of protein between compartments of the Golgi stack. In contrast to the striking effects of calphostin C, reagents that inhibit the function of the catalytic domain of PKC (including the general kinase inhibitor staurosporine, as well as the more specific inhibitors H-7, H-8, pseudosubstrate inhibitor, or chelerythrine) did not inhibit export from the ER. Export was also insensitive to down-regulation of various PKC isoforms. These results suggest that a novel protein containing the conserved C6H2 motif may serve as a potential link in a signaling pathway regulating vesicle budding from the ER.
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PMID:Export of protein from the endoplasmic reticulum is regulated by a diacylglycerol/phorbol ester binding protein. 752 13

Newly synthesized membrane proteins are exported from the endoplasmic reticulum to the Golgi complex through an intermediate compartment. Incubation at low temperature (15 degrees C) arrests the proteins in the intermediate compartment and prevents the entry into the Golgi complex. We have studied, in living cells, the effect of dithiothreitol (DTT) and ATP depletion on the transport to the Golgi complex of proteins accumulated either in the endoplasmic reticulum or in the intermediate compartment after a temperature block. The morphological results obtained with vesicular stomatitis virus ts-O45 G glycoprotein and the biochemical analysis performed with human CD8 protein, an O-glycosylated protein, showed that: 1) ATP depletion blocks the export to the Golgi complex of proteins located either in the endoplasmic reticulum or in the intermediate compartment and ii) DTT interferes with the folding and export of proteins located in the endoplasmic reticulum, but it does not prevent the transfer from the intermediate compartment to the Golgi complex.
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PMID:Effect of ATP depletion and DTT on the transport of membrane proteins from the endoplasmic reticulum and the intermediate compartment to the Golgi complex. 758 83

Glycosylphosphatidylinositols (GPIs) are ubiquitous in eukaryotes and serve to anchor a variety of proteins to the exoplasmic leaflet of cellular membranes. GPIs are synthesized in the endoplasmic reticulum (ER), in excess of the amount needed for protein modification. The fate of the excess GPIs is unknown, but they may be retained in the ER, transported to other membranes, and/or metabolized. In relation to this problem, we were interested in determining whether GPIs were transported to the plasma membrane and whether, like GPI-anchored proteins, their presence was confined to the apical plasma membrane domain in polarized epithelial cells. Polarized Madin-Darby canine kidney epithelial cell monolayers were incubated with [3H]mannose or [3H]ethanolamine to label GPIs and then infected with enveloped viruses. We used influenza virus (flu) and vesicular stomatitis virus (VSV) for these experiments as these viruses are assembled at the cell surface and acquire their envelope lipids from the plasma membrane. Furthermore, flu and VSV bud specifically from the apical and basolateral plasma membrane domains, respectively. Flu and VSV were isolated from the apical and basolateral media, respectively, and subjected to lipid analysis. Radiolabeled GPIs were found in both viruses. Moreover, the membrane concentration of GPIs (i.e. GPI radioactivity normalized to membrane mass) in the two viruses was essentially the same. These observations suggest that (i) non-protein-linked GPIs are located at the plasma membrane; (ii) since GPIs are synthesized in the ER, they must be transported from the ER to the plasma membrane; and (iii) transport of nonprotein-linked GPIs is not influenced by the sorting processes that target GPI-anchored proteins exclusively to the apical plasma membrane.
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PMID:Nonpolarized distribution of glycosylphosphatidylinositols in the plasma membrane of polarized Madin-Darby canine kidney cells. 759 18


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