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

The lateral mobility of the vesicular stomatitis virus spike glycoprotein (G protein) and various mutant G proteins produced by site-directed mutagenesis of the G cDNA has been measured. Fluorescence recovery after photobleaching results for the wild type G protein in transfected COS-1 cells yielded a mean diffusion coefficient (D) of 8.5 (+/- 1.3) X 10(-11) cm2/s and a mean mobile fraction of 75% (+/- 3%). Eight mutant proteins were also examined: dTM14, lacking six amino acids from the transmembrane domain; TA2, lacking an oligosaccharide in the extracellular domain; QN2, possessing an extra N-linked oligosaccharide in the extracellular domain; CS2, possessing a serine instead of a cysteine at residue 489 in the cytoplasmic domain, preventing palmitate addition to the glycoprotein; TMR-stop, lacking the entire cytoplasmic domain except an arginine at residue 483; and three chimeric proteins, G mu, G23, and GHA, containing in place of the 29 amino acid wild type cytoplasmic domain the cytoplasmic domains from the surface IgM from the spike protein of the infectious bronchitis virus or from the hemagglutinin protein of the influenza virus, respectively. The mean D for the mutant proteins varied over a relatively small range, with the slowest mutant, G23, exhibiting a value of 11.3 (+/- 1.4) X 10(-11) cm2/s and the fastest mutant, GHA, having a D of 28.6 (+/- 4.5) X 10(-11) cm2/s. The mean mobile fraction similarly varied over a small range, extending from 55 to 68%. None of the mutations resulted in the more rapid diffusion characteristic of membrane proteins embedded in artificial bilayers. Therefore, it appears that the cytoplasmic and transmembrane domains themselves contribute little to restraining the lateral mobility of this integral membrane protein when expressed in transfected cells.
 ...
PMID:Effects of mutations in three domains of the vesicular stomatitis viral glycoprotein on its lateral diffusion in the plasma membrane. 303 31

The recombinant polymerase protein L of vesicular stomatitis virus (VSV) expressed in COS cells is able to transcribe and replicate the viral genome, resulting in complementation of temperature-sensitive polymerase mutants of VSV at the restrictive temperature (M. Schubert, G. G. Harmison, C. D. Richardson, and E. Meier, Proc. Natl. Acad. Sci. USA 82:7984-7988, 1985). Here we report that the efficiency of complementation is dependent on the level of L protein expression. Unexpectedly, only cells expressing low levels of recombinant L protein efficiently complemented tsL gene mutants, whereas cells with high levels of L protein did not. In fact, in all cells with high levels of L protein expression, which at 40 h posttransfection represented almost the total number of transfected cells, viral replication not only of the temperature-sensitive mutant but also of wild-type VSV was excluded. The inhibition of VSV appeared to occur at an early stage of the infectious cycle, and wild-type virus of the same serotype (Indiana) as the recombinant L protein as well as wild-type virus of a different serotype (New Jersey) was affected. Measles virus, on the other hand, was not arrested in cells with high levels of recombinant L protein, demonstrating that these cells were still capable of supporting a viral infection. The expression of high levels of only the amino-terminal half of the L protein from a recombinant mutant L gene that contains a small out-of-frame deletion in the middle of the L gene did not inhibit a VSV infection. Since the level of amplification for both L- and truncated L-encoding vectors is similar, we conclude that the arrest of VSV was caused by high levels of functional full-length L protein itself and not by high levels of vector-encoded L mRNA or other vector products or by side effects of vector amplification. These data strongly support the idea that the highly conserved gene order of nonsegmented negative-strand viruses and the sequential and attenuated mode of transcription are important regulatory elements which balance the intracellular concentration of viral proteins. They both assure that the L gene is the last and the least frequently transcribed gene, giving rise to low levels of L protein necessary for efficient replication.
 ...
PMID:Homotypic and heterotypic exclusion of vesicular stomatitis virus replication by high levels of recombinant polymerase protein L. 304 Oct 35

We constructed a molecular clone encoding the N-terminal 379 amino acids of the polyomavirus middle-size tumor antigen, followed by the C-terminal 60 amino acids of the vesicular stomatitis virus glycoprotein G. This hybrid gene contained the coding region for the C-terminal hydrophobic membrane-spanning domain of the G protein in place of the C-terminal hydrophobic domain of the middle-size tumor antigen. The hybrid gene was expressed in COS-1 cells under the control of the simian virus 40 late promoter. The hybrid protein was located in cell membranes and was associated with a tyrosine-specific protein kinase activity, as was the middle-size tumor antigen. Plasmids encoding the hybrid protein failed to transform mouse NIH 3T3 or rat F2408 cells.
 ...
PMID:Construction and expression of a recombinant DNA gene encoding a polyomavirus middle-size tumor antigen with the carboxyl terminus of the vesicular stomatitis virus glycoprotein G. 632 57

The human papillomavirus-16 (HPV16) E5 gene is able to induce stable growth transformation and transient mitogenic stimulation in a variety of cultured cell systems. To characterize the biochemical properties of the hydrophobic HPV16 E5 transforming protein, we have constructed vectors expressing the wild-type HPV16 E5 gene and have generated antipeptide antisera. The 10-kDa E5 protein was readily detectable in transfected COS monkey cells by using these antisera either for immunoprecipitation of metabolically labeled cells or for immunoblotting. Coimmunoprecipitation analysis of cells coexpressing the viral protein and various growth factor receptors demonstrated stable complex formation between the E5 protein and the epidermal growth factor receptor, platelet-derived growth factor beta receptor, colony stimulating factor-1 receptor, and p185neu. The E5 protein also formed a stable complex with the vesicular stomatitis virus glycoprotein. These experiments indicated that the HPV16 E5 protein was able to participate in complex formation with a variety of transmembrane proteins, a property which may contribute to the biological activities of the viral protein. In addition, the expression vectors and antibodies described here will be useful reagents in examining various aspects of HPV16 E5 expression and function.
 ...
PMID:The HPV16 E5 protein: expression, detection, and stable complex formation with transmembrane proteins in COS cells. 764 15

Cellular ezrin-radixin-moesin (ERM) family proteins, members of the actin-binding proteins of the band 4.1 superfamily, were detected in the virions of enveloped viruses, such as rabies, vesicular stomatitis, Newcastle disease, and influenza viruses. To elucidate the mechanism of ERM protein incorporation, we investigated possible association of ERM proteins with viral components in rabies virus-infected BHK-21 cells. Double immunofluorescence studies demonstrated that the ERM proteins are concentrated in the microvilli, where the colocalized viral G protein was also seen. Viral G protein expressed in the G cDNA-transfected COS-7 cells also displayed similar distributions to those seen in the virus-infected cells. Both the ERM and viral envelope proteins were coprecipitated by anti-viral G antibody from lysates of the virus-infected cells, while the anti-ERM antibody coprecipitated viral G and ERM proteins. These observations suggest that the ERM proteins are closely associated with viral envelope proteins in the cell, which would be involved in the selective incorporation of cellular actin into the virion.
 ...
PMID:Cellular actin-binding ezrin-radixin-moesin (ERM) family proteins are incorporated into the rabies virion and closely associated with viral envelope proteins in the cell. 783 4

In order to identify the amino acid sequences responsible for the internalization of the cloned rat brain neurotensin receptor, we carried out site-directed mutagenesis of the cDNA encoding the receptor followed by expression of the receptor into mammalian COS 7 cells. In cells transfected with the full-length neurotensin receptor, 56% of iodinated neurotensin specifically bound to the cells after 60 min of incubation at 37 degrees C was internalized. Deletions made in the third intracellular loop did not affect receptor internalization. By contrast, internalization was reduced to 5% of total in cells in which almost all the carboxyl-terminal tail of the receptor had been deleted (R392stop). In order to determine which part of the tail was responsible for this effect, several Ser and Thr residues were deleted in the carboxyl cytoplasmic sequence of the receptor. Almost all of these receptors were internalized as efficiently as the wild type. Only the form of the neurotensin receptor truncated at Glu-421 (deletion of the last three residues, TLY) produced a significant decrease in the amount of ligand internalized. Finally, point mutations of Thr-422 and Tyr-424 residues to Gly led to an almost complete loss of ligand internalization demonstrating the involvement of these 2 residues in the internalization process. Replacement of the last three amino acids by the cytoplasmic endocytosis signal of the vesicular stomatitis virus did not restore the efficiency of neurotensin receptor internalization. These biochemical results were confirmed by confocal microscopic analysis. Cell transfected with the wild type receptor showed a temperature-dependent intracellular accumulation of a fluorescent analog of neurotensin, whereas cells transfected with a receptor truncated at the carboxyl terminus showed a clustering of the fluorescent peptide at the cell surface.
 ...
PMID:Thr-422 and Tyr-424 residues in the carboxyl terminus are critical for the internalization of the rat neurotensin receptor. 785 3

Although basic fibroblast growth factor (bFGF/FGF-2) is found outside cells, it lacks a conventional signal peptide sequence; the mechanism underlying its export from cells is therefore unknown. Using a transient COS-1 cell expression system, we have identified a novel membrane-associated transport pathway that mediates export of FGF-2. This export pathway is specific for the 18-kD isoform of FGF-2, is resistant to the anti-Golgi effects of Brefeldin A, and is energy-dependent. In FGF-2-transfected COS-1 cells, this ER/Golgi-independent pathway appears to be constitutively active and functions to quantitatively export metabolically-labeled 18-kD FGF-2. Co-transfection and co-immunoprecipitation experiments, using a vector encoding the cytoplasmic protein neomycin phosphotransferase, further demonstrated the selectivity of this export pathway for FGF-2. When neomycin phosphotransferase was appended to the COOH-terminus of 18-kD FGF-2, the chimera was exported. However, the transmembrane anchor sequence of the integral membrane glycoprotein (G protein) of vesicular stomatitis virus (VSV) blocked export. The chimeric protein localized to the plasma membrane with its FGF-2 domain extracellular and remained cell-associated following alkaline carbonate extraction. Taken together, the data suggest that FGF-2 is "exported" from cells via a unique cellular pathway, which is clearly distinct from classical signal peptide-mediated secretion. This model system provides a basis for the development and testing of therapeutic agents which may block FGF-2 export. Such an intervention may be of considerable use for the treatment of angiogenesis-dependent diseases involving FGF-2.
 ...
PMID:Quantitative export of FGF-2 occurs through an alternative, energy-dependent, non-ER/Golgi pathway. 786 Jun 46

Many soluble resident proteins of the endoplasmic reticulum share a COOH-terminal Lys-Asp-Glu-Leu (KDEL) sequence. Current opinion favours a model in which these proteins can escape from the endoplasmic reticulum (ER) by bulk flow and are recognized and sorted in the Golgi apparatus by binding to a specific KDEL-receptor, which returns them to the ER. Through biochemical, morphological and mutational analysis we have studied the mechanisms that determine the localization of calreticulin, a soluble 60 kDa KDEL-protein of the ER. Immunogold labelling established the ER localization of calreticulin in transfected and nontransfected COS cells. Although the ER cisternae in transfected cells were enormously dilated and heavily labelled by gold particles we found no significant label in any other compartment. In vivo pulse chase experiments with [35S]methionine followed by biochemical fractionation of calreticulin overexpressing COS cells (50- to 100-fold) revealed that only a minor part of labelled calreticulin leaves the ER. Retrieval from the Golgi was confirmed by a partial redistribution of the endogenous KDEL-receptor as shown by double immunofluorescence. These data suggest a KDEL-independent retention of calreticulin in the ER. Further supporting evidence has come from morphological in vivo studies using calreticulin-transfected and vesicular stomatitis virus (ts045)-infected COS cells. Stimulation of vesicular transport from the ER by releasing the temperature-dependent transport block for the viral G-protein resulted in a small but significant appearance of calreticulin in a post-ER compartment. In contrast a calreticulin mutant, which lacked the Ca(2+)-binding domain but included the KDEL sequence, could escape from the ER to a much higher extent. Secretion of the nonmutated calreticulin was very low (1-2% of total calreticulin in 3 hours) compared to the mutated form (18% in 3 hours). Deletion of the KDEL sequence led to an increase in secretion to 29% over a 3 hour period, which is much less than expected for a secretory protein. Taken together these results strongly support the hypothesis of two independently operating retention/retrieval mechanisms for calreticulin: one providing for direct retention in the ER with a very high capacity and having Ca(2+)-dependent properties; the other a KDEL-based retrieval system for escaped calreticulin present in the Golgi apparatus.
 ...
PMID:Retention and retrieval: both mechanisms cooperate to maintain calreticulin in the endoplasmic reticulum. 787 39

The envelope glycoprotein G of vesicular stomatitis virus induces membrane fusion at low pH. Site-directed mutagenesis of specific amino acids within a segment spanning amino acids 123 to 137 of G protein, which is highly conserved in vesiculoviruses and was previously shown by us to be involved in fusogenic activity (Y. Li, C. Drone, E. Sat, and H. P. Ghosh, J. Virol. 67:4070-4077, 1993), was used to determine the role of this region in low-pH-induced membrane fusion. The mutant glycoproteins expressed in COS cells were assayed for acid-pH-induced cell-cell fusion. Substitution of the variant Pro-123 with Leu had no effect on the fusogenic activity, while substitution of conserved Phe-125 and Asp-137 with Tyr and Asn, respectively, shifted the pH optimum of membrane fusion to a more acidic pH value and decreased the fusion efficiency. The deletion of amino acid residues 124 to 127, 131 to 137, or 124 to 137 produced mutants defective in transport. Mutation of the conserved residues Gly-124 and Pro-127 to Ala and to Gly or Leu, respectively, inhibited cell-cell fusion activity by about 90% without affecting transport of the mutant proteins to the cell surface, suggesting that these two residues may be present within the fusion peptide and thus may be directly involved in fusion. This highly conserved domain containing neutral amino acids of G protein may therefore represent the putative fusion domain of vesicular stomatitis virus G protein.
 ...
PMID:Characterization of the putative fusogenic domain in vesicular stomatitis virus glycoprotein G. 813 3

We have used the glycoprotein gB of herpes simplex virus type 1 (gB-1), which buds from the inner nuclear membrane, as a model protein to study localization of membrane proteins in the nuclear envelope. To determine whether specific domains of gB-1 glycoprotein are involved in localization in the nuclear envelope, we have used deletion mutants of gB-1 protein as well as chimeric proteins constructed by replacing the domains of the cell surface glycoprotein G of vesicular stomatitis virus with the corresponding domains of gB. Mutant and chimeric proteins expressed in COS cells were localized by immunoelectron microscopy. A chimeric protein (gB-G) containing the ectodomain of gB and the transmembrane and cytoplasmic domains of G did not localize in the nuclear envelope. When the ectodomain of G was fused to the transmembrane and cytoplasmic domains of gB, however, the resulting chimeric protein (G-gB) was localized in the nuclear envelope. Substitution of the transmembrane domain of G with the 69 hydrophobic amino acids containing the membrane anchoring domain of gB allowed the hybrid protein (G-tmgB) to be localized in the nuclear envelope, suggesting that residues 721 to 795 of gB can promote retention of proteins in the nuclear envelope. Deletion mutations in the hydrophobic region further showed that a transmembrane segment of 21 hydrophobic amino acids, residues 774 to 795 of gB, was sufficient for localization in the nuclear envelope. Since wild-type gB and the mutant and chimeric proteins that were localized in the nuclear envelope were also retained in the endoplasmic reticulum, the membrane spanning segment of gB could also influence retention in the endoplasmic reticulum.
 ...
PMID:Membrane anchoring domain of herpes simplex virus glycoprotein gB is sufficient for nuclear envelope localization. 813 12


<< Previous 1 2 3 4 5 Next >>