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Query: UMLS:C0038362 (stomatitis)
 8,852 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The vesicular stomatitis virus glycoprotein (G protein) is an integral membrane protein which assembles into noncovalently associated trimers before transport from the endoplasmic reticulum. In this study we have examined the folding and oligomeric assembly of twelve mutant G proteins with alterations in the cytoplasmic, transmembrane, or ectodomains. Through the use of conformation-specific antibodies, we found that newly synthesized G protein folded into a conformation similar to the mature form within 1-3 min of synthesis and before trimer formation. Mutant proteins not capable of undergoing correct initial folding did not trimerize, were not transported, and were found in large aggregates. They had, as a rule, mutations in the ectodomain, including several with altered glycosylation patterns. In contrast, mutations in the cytoplasmic domain generally had little effect on folding and trimerization. These mutant proteins, whose ectodomains were identical to the wild-type by several assays, were either transported to the cell surface slowly or not at all. We concluded that while correct ectodomain folding and trimer formation are prerequisites for transport, they alone are not sufficient. The results suggest that the cytoplasmic domain of the wild-type protein may facilitate rapid, efficient transport from the ER, which can be easily affected or eliminated by tail mutations that do not detectably affect the ectodomain.
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PMID:Differential effects of mutations in three domains on folding, quaternary structure, and intracellular transport of vesicular stomatitis virus G protein. 283 23

Wild-type vesicular stomatitis virus-infected cells contained multiple carboxy-terminal fragments of the envelope glycoprotein G. They migrated in 16% polyacrylamide gels with two dominant apparent molecular weights, 14,000 and 9,000. Both fragments were immunoprecipitated by two antibodies, anti-G(COOH) and anti-G(stem), made against the last 15 amino acids at the carboxy terminus and against the first 22 amino acids of the ectodomain adjacent to the transmembrane region of G, respectively. Pulse-chase experiments in the presence and absence of tunicamycin indicated that the higher-molecular-weight fragment, Gal, was generated first, presumably in the rough endoplasmic reticulum, and then apparently chased into the faster-migrating, stable fragment, Ga2. Exposure of infected cells to radioactive palmitic acid labeled Ga2. Ga2 was detected in purified virions. These results show that a polypeptide approximately 71 amino acids long is transported and incorporated into budding virions. What signals are operative and whether this C-terminal fragment of G protein is transported as a complex with other viral or host cell proteins are presently unknown.
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PMID:Membrane anchors of vesicular stomatitis virus: characterization and incorporation into virions. 283 85

Alterations of the cytoplasmic domain of the vesicular stomatitis virus glycoprotein (G protein) were shown previously to affect transport of the protein from the endoplasmic reticulum, and recent studies have shown that this occurs without detectable effects on G protein folding and trimerization (R. W. Doms et al., J. Cell Biol., in press). Deletions within this domain slowed exit of the mutant proteins from the endoplasmic reticulum, and replacement of this domain with a foreign 12-amino-acid sequence blocked all transport out of the endoplasmic reticulum. To extend these studies, we determined whether such effects of cytoplasmic domain changes were transferable to other proteins. Three different assays showed that the effects of the mutations on transport of two membrane-anchored secretory proteins were the same as those observed with vesicular stomatitis virus G protein. In addition, possible effects on oligomerization were examined for both transported and nontransported forms of membrane-anchored human chorionic gonadotropin-alpha. These membrane-anchored forms, like the nonanchored human chorionic gonadotropin-alpha, had sedimentation coefficients consistent with a monomeric structure. Taken together, our results provide strong evidence that these cytoplasmic mutations affect transport by affecting interactions at or near the cytoplasmic side of the membrane.
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PMID:Effects of altered cytoplasmic domains on transport of the vesicular stomatitis virus glycoprotein are transferable to other proteins. 284 89

The insertion of proteins into the endoplasmic reticulum is mediated by short hydrophobic domains called signal sequences, which are usually cleaved during insertion. We previously constructed DNAs encoding vesicular stomatitis virus glycoproteins with N-terminal extensions preceding the signal sequence and showed that these extensions allowed normal signal-sequence function and cleavage in vivo. To analyze signal sequence topology during membrane insertion, we generated a point mutation that blocks cleavage of these signal sequences. After expressing these proteins in HeLa cells, we used proteolysis of microsomal membranes to determine that the N terminus of the signal sequence and the C terminus of each protein remain on the cytoplasmic side of the endoplasmic reticulum after insertion. This result indicates that the proteins were inserted in a looped configuration. Extending this finding, we were able to reverse the orientation of such a mutant protein by deleting its normal C-terminal transmembrane and cytoplasmic domains. In addition to demonstrating that a signal sequence can function as a membrane anchor, these findings show that except for the presence of a cleavage site, the cleaved signal sequence of a type I transmembrane protein is structurally and functionally equivalent to the noncleaved signal sequences of type II transmembrane proteins.
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PMID:Evidence for the loop model of signal-sequence insertion into the endoplasmic reticulum. 284 15

We have used a protocol for internalization of ricin, a ligand binding to plasma membrane glycoproteins and glycolipids with terminal galactosyl residues, and infection with the vesicular stomatitis virus ts 045 mutant in BHK-21 cells to determine whether internalized plasma membrane molecules tagged by ricin reach distinct compartments of the biosynthetic-exocytic pathway. At 39.5 degrees C newly synthesized G protein of ts 045 was largely prevented from leaving the endoplasmic reticulum. At the same temperature ricin was endocytosed and reached, in addition to endosomes and lysosomes, elements of the Golgi complex. When the temperature was lowered to 19.5 degrees C, no more ricin was delivered to the Golgi complex, but now G protein accumulated in the Golgi stacks and the trans-Golgi network (TGN). Double-labeling immunogold cytochemistry on ultracryosections was used to detect G protein and ricin simultaneously. These data, combined with stereological and biochemical methods, showed that approximately 5% of the total amount of ricin within the cells, corresponding to 6-8 X 10(4) molecules per cell, colocalized with G protein in the Golgi complex after 60 min at 39.5 degrees C. Of this amount approximately 70-80% was present in the TGN. Since most of the ricin molecules remain bound to their binding sites at the low pH prevailing in compartments of the endocytic pathway, the results indicate that a fraction of the internalized plasma membrane molecules with terminal galactose are not recycled directly from endosomes or delivered to lysosomes, but are routed to the Golgi complex. Also, the results presented here, in combination with other recent studies on ricin internalization, suggest that translocation of the toxic ricin A-chain to the cytosol occurs in the TGN.
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PMID:Estimation of the amount of internalized ricin that reaches the trans-Golgi network. 289 43

The intracellular route followed by viral envelope glycoproteins in polarized Madin-Darby canine kidney cells was studied by using temperature-sensitive mutants of vesicular stomatitis virus (VSV) and influenza, in which, at the nonpermissive temperature (39.5 degrees C), the newly synthesized glycoproteins (G proteins) and hemagglutinin (HA), respectively, are not transported out of the endoplasmic reticulum. After infection with VSV and incubation at 39.5 degrees C for 4-5 h, synchronous transfer of G protein to the plasma membrane was initiated by shifting to the permissive temperature (32.5 degrees C). Immunoelectron microscopy showed that under these conditions the protein moved to the Golgi apparatus and from there directly to a region of the lateral plasma membrane near this organelle. G protein then seemed to diffuse progressively to basal regions of the cell surface and, only after it had accumulated in the basolateral domain, it began to appear on the apical surface near the intercellular junctions. The results of these experiments indicate that the VSV G protein must be sorted before its arrival at the cell surface, and suggest that passage to the apical domain occurs only late in infection when tight junctions are no longer an effective barrier. In complementary experiments, using the temperature-sensitive mutant of influenza, cultures were first shifted from the nonpermissive temperature (39.5 degrees C) to 18.5 degrees C, to allow entrance of the glycoprotein into the Golgi apparatus (see Matlin, K.S., and K. Simons, 1983, Cell, 34:233-243). Under these conditions HA accumulated in Golgi stacks and vesicles but did not reach the plasma membrane. When the temperature was subsequently shifted to 32.5 degrees C, HA rapidly appeared in discrete regions of the apical surface near, and often directly above, the Golgi elements, and later diffused throughout this surface. To ensure that the anti-HA antibodies had access to lateral domains, monolayers were treated with a hypertonic medium to dilate the intercellular spaces. Some labeling was then observed in the lateral plasma membranes soon after the shift, but this never increased beyond 1.0 gold particle/micron, whereas characteristic densities of labeling in apical surfaces soon became much higher (approximately 10 particles/micron). Our results suggest that the bulk of HA follows a direct pathway leading from the Golgi to regions of the apical surface close to trans-Golgi cisternae.
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PMID:Polarized delivery of viral glycoproteins to the apical and basolateral plasma membranes of Madin-Darby canine kidney cells infected with temperature-sensitive viruses. 298 Dec 29

The outer nuclear membrane is morphologically similar to rough endoplasmic reticulum. The presence of ribosomes bound to its cytoplasmic surface suggests that it could be a site of synthesis of membrane glycoproteins. We have examined the biogenesis of the vesicular stomatitis virus G protein in the nuclear envelope as a model for the biogenesis of membrane glycoproteins. G protein was present in nuclear membranes of infected Friend erythroleukemia cells immediately following synthesis and was transported out of nuclear membranes to cytoplasmic membranes with a time course similar to transport from rough endoplasmic reticulum (t 1/2 = 5-7 min). Temperature-sensitive mutations in viral membrane proteins which block transport of G protein from endoplasmic reticulum also blocked transport of G protein from the nuclear envelope. Friend erythroleukemia cells and NIH 3T3 cells differed in the fraction of newly synthesized G protein found in nuclear membranes, apparently reflecting the relative amount of nuclear membrane compared to endoplasmic reticulum available for glycoprotein synthesis. Nuclear membranes from erythroleukemia cells appeared to have the enzymatic activities necessary for cleavage of the signal sequence and core glycosylation of newly synthesized G protein. Signal peptidase activity was detected by the ability of detergent-solubilized membranes of isolated nuclei to correctly remove the signal sequence of human preplacental lactogen. RNA isolated from the nuclear envelope was highly enriched for G protein mRNA, suggesting that G protein was synthesized on the outer nuclear membrane rather than redistributing to nuclear membranes from endoplasmic reticulum before or during cell fractionation. These results suggest a mechanism for incorporation of membrane glycoproteins into the nuclear envelope and suggest that in some cell types the nuclear envelope is a major source of newly synthesized membrane glycoproteins.
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PMID:Role of the nuclear envelope in synthesis, processing, and transport of membrane glycoproteins. 298 6

We have developed a new selection procedure for mammalian cell mutants defective in protein export by the use of diphtheria toxin, and devised a new screening method for defective protein secretion using nitrocellulose membranes. By the combination of these procedures, we have isolated a temperature-sensitive mutant clone of Chinese hamster ovary cells which shows a pleiotropic defect in protein export. This mutant, designated DS28-6, is temperature-sensitive for growth. Secretion of a series of proteins is markedly inhibited at the non-permissive temperature. These proteins seem to be normally synthesized and accumulated within the cell at the non-permissive temperature and secreted upon shift down to the permissive temperature. When this mutant is infected with vesicular stomatitis virus, oligosaccharide processing of G-protein is arrested at an endoglycosidase-H-sensitive stage at the non-permissive temperature. The lesion of this mutant appears to be in the endoplasmic reticulum or the cis Golgi or both.
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PMID:A temperature-sensitive Chinese hamster ovary cell mutant pleiotropically defective in protein export. 298 43

The structures of the asparagine-linked oligosaccharides of several variant forms of the vesicular stomatitis virus glycoprotein transiently expressed from cloned cDNAs have been determined. Glycopeptides isolated from forms of the G protein that reach the cell surface or that are secreted into the medium are virtually identical; they contain complex-type oligosaccharides whose nonreducing ends terminate in galactose and sialic acid residues. In contrast, forms of the G protein that remain intracellular possess oligosaccharides at intermediate stages in the processing pathway. One deletion mutant, delta 1473, codes for a protein that remains in the rough endoplasmic reticulum (Rose, J. K., and J. E. Bergmann, 1982, Cell, 30:753-762) and contains only high mannose-type oligosaccharides. Another mutant, delta 1554, codes for a glycoprotein that contains oligosaccharides of primarily two classes. One class is of the high mannose type and is similar to those found on the protein coded for by delta 1473. However, the major class contains biantennary and more highly branched complex-type oligosaccharides that terminate in N-acetylglucosamine rather than galactose or sialic acid residues. These data suggest that the protein coded for by delta 1554 migrates to the Golgi apparatus, but does not enter the more distal compartment(s) of the organelle which contains galactosyl- and sialyltransferases.
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PMID:Processing of the asparagine-linked oligosaccharides of secreted and intracellular forms of the vesicular stomatitis virus G protein: in vivo evidence of Golgi apparatus compartmentalization. 299 Dec 99

The synthesis and oligosaccharide processing of the glycoproteins of SA11 rotavirus in infected Ma104 cells was examined. Rotavirus assembles in the rough endoplasmic reticulum (RER) and encodes two glycoproteins: VP7, a component of the outer viral capsid, and NCVP5, a nonstructural protein. A variety of evidence suggests the molecules are limited to the ER, a location consistent with the high mannose N-linked oligosaccharides modifying these proteins. VP7 and NCVP5 were shown to be integral membrane proteins. In an in vitro translation system supplemented with dog pancreas microsomes, they remained membrane associated after high salt treatment and sodium carbonate-mediated release of microsomal contents. In infected cells, the oligosaccharide processing of these molecules proceeded in a time-dependent manner. For VP7, Man8GlcNAc2 and Man6GlcNAc2 were the predominant intracellular species after a 5-min pulse with [3H]mannose and a 90 min chase, while in contrast, trimming of NCVP5 halted at Man8GlcNAc2. VP7 on mature virus was processed to Man5GlcNAc2. It is suggested that the alpha-mannosidase activities responsible for the formation of these structures reside in the ER. In the presence of the energy inhibitor carbonyl cyanide m-chlorophenylhydrazone (CCCP), processing of VP7 and the vesicular stomatitis virus G protein was blocked at Man8GlcNAc2. After a 20-min chase of [3H]mannose-labeled molecules followed by addition of CCCP, trimming of VP7 could continue while processing of G protein remained blocked. Thus, an energy-sensitive translocation step within the ER may mark the divergence of the processing pathways of these glycoproteins.
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PMID:Processing of the rough endoplasmic reticulum membrane glycoproteins of rotavirus SA11. 299 4


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