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

To define the effects of pp60v-src activity at different intracellular sites, we have constructed chimeric molecules which target the pp60v-src kinase to specific intracellular locations. pp60v-src was targeted to the nucleus by insertion of the SV40 large T antigen nuclear localization signal. Nuclear pp60v-src was active as a tyrosine kinase and phosphorylated nuclear proteins at tyrosine. However, cells expressing the nuclear pp60v-src were phenotypically normal by a number of criteria, and nuclear src kinase did not induce the expression of an mRNA (CEF-4) whose induction is characteristic of transformation by wild-type v-src. pp60v-src was targeted to perinuclear membranes by fusion to rat growth hormone and vesicular stomatitis G protein sequences. Cells expressing this chimeric molecule were phenotypically normal by most criteria. However the perinuclear src protein did induce elevated levels of CEF-4 mRNA, indicating that the v-src kinase expressed at this site induces partial transformation. The v-src and activated c-src kinases were targeted to adhesion plaques by fusion to the talin-binding sequence of vinculin. Cells expressing these fusion proteins were transformed by morphological, physiological and biochemical criteria, although the foci induced by these viruses were distinct from those induced by wild-type v-src. A chimeric protein which targeted c-src to adhesion plaques was not transforming. Thus targeting pp60src to adhesion plaques, although not sufficient to activate the transforming capacity of c-src, is sufficient to allow transformation by v-src.
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PMID:Intracellular targeting of pp60src expression: localization of v-src to adhesion plaques is sufficient to transform chicken embryo fibroblasts. 133 49

Membrane traffic has been shown to be regulated during cell division. In particular, with the use of viral membrane proteins as markers, endoplasmic reticulum (ER)-to-Golgi transport in mitotic cells has been shown to be essentially blocked. However, the effect of mitosis on other steps in the secretory pathway is less clear, because an early block makes examination of following steps difficult. Here, we report studies on the functional characteristics of secretory pathways in mitotic mammalian tissue culture cells by the use of a variety of markers. Chinese hamster ovary cells were transfected with cDNAs encoding secretory proteins. Consistent with earlier results following viral membrane proteins, we found that the overall secretory pathway is nonfunctional in mitotic cells, and a major block to secretion is at the step between ER and Golgi: the overall rate of secretion of human growth hormone is reduced at least 10-fold in mitotic cells, and export of truncated vesicular stomatitis virus G protein from the ER is inhibited to about the same extent, as judged by acquisition of endoglycosidase H resistance. To ascertain the integrity of transport from the trans-Golgi to plasma membrane, we followed the secretion of sulfated glycosaminoglycan (GAG) chains, which are synthesized in the Golgi and thus are not subject to the earlier ER-to-Golgi block. GAG chains are valid markers for the pathway taken by constitutive secretory proteins; both protein secretion and GAG chain secretion are sensitive to treatment with n-ethyl-maleimide and monensin and are blocked at 19 degrees C. We found that the extent of GAG-chain secretion is not altered during mitosis, although the initial rate of secretion is reduced about twofold in mitotic compared with interphase cells. Thus, during mitosis, transport from the trans-Golgi to plasma membrane is much less hindered than ER-to-Golgi traffic. We conclude that transport steps are not affected to the same extent during mitosis.
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PMID:Membrane traffic between secretory compartments is differentially affected during mitosis. 209 91

Caco-2 cells, derived from human colon, have the morphological, functional, and biochemical properties of small intestinal epithelial cells. After infection with enveloped viruses, influenza virions assembled at the apical plasma membrane while vesicular stomatitis virus (VSV) particles appeared exclusively at the basolateral membrane, similar to the pattern observed in virus-infected Madin-Darby canine kidney (MDCK). When grown in Millicell filter chamber devices and labeled with [35S]methionine, Caco-2 monolayers released all of their radiolabeled secretory products preferentially into the basal chamber. Among the proteins identified were apolipoproteins AI and E, transferrin, and alpha-fetoprotein. No proteins were observed to be secreted preferentially from the apical cell surface. The lysosomal enzyme beta-hexosaminidase was also secreted primarily from the basolateral surface of the cells in the presence or absence of lysosomotropic drugs or tunicamycin, which inhibit the targetting of lysosomal enzymes to lysosomes. Neither of these drug treatments significantly affected the polarized secretion of other nonlysosomal proteins. In addition, growth hormone (GH), which is released in a nonpolar fashion from MDCK cells, was secreted exclusively from the basolateral membrane after transfection of Caco-2 cells with GH cDNA in a pSV2-based expression vector. Similar results were obtained in transient expression experiments and after selection of permanently transformed Caco-2 cells expressing GH. Since both beta-hexosaminidase and GH would be expected to lack sorting signals for polarized exocytosis in epithelial cells, these results indicate that in intestinal cells, proteins transported via the basolateral secretory pathway need not have specific sorting signals.
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PMID:A specific sorting signal is not required for the polarized secretion of newly synthesized proteins from cultured intestinal epithelial cells. 245 57

We have expressed the hybrid protein, GHG3, in baby hamster kidney cells to study protein turnover. GHG3 contains the cytoplasmic and transmembrane domains of vesicular stomatitis virus G protein linked to the C-terminus rat growth hormone. Turnover of GHG3 was prevented by lysosomal inhibitors (leupeptin, chloroquine, primaquine or monensin), while the accumulated GHG3 was localized to intracellular vesicles, results indicating that degradation occurred in lysosomes. The kinetics of degradation at 34 degrees C were determined in pulse-chase studies of metabolically labeled cells. After a lag period of 1 h, degradation was rapid (t1/2 = 1.25 h). The fate of GHG3 during the lag period was determined by immunofluorescence. We detected GHG3 on the cell surface when growth hormone antiserum was added to the growth medium 90 min prior to fixation and staining. No staining was observed if protein synthesis was inhibited with cycloheximide 90 min prior to the addition of growth hormone antiserum, a result indicating that GHG3 was rapidly removed from the cell surface. Unless the cells were pretreated with cycloheximide, antiserum was also detected in intracellular vesicles, which showed that GHG3 was endocytosed. These data indicate that a pool of GHG3 is transported rapidly to the cell surface, endocytosed and with little or no recycling directed to lysosomes for degradation.
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PMID:A growth hormone-vesicular stomatitis virus G hybrid protein is rapidly degraded in lysosomes following transport to the cell surface. 254 15

Various aspects of the biogenetic mechanisms that are involved in the insertion of nascent plasma membrane proteins into the endoplasmic reticulum (ER) membrane and their subsequent distribution through the cell have been investigated. For these studies chimeric genes that encode hybrid proteins containing carboxy-terminal portions of the influenza virus hemagglutinin (154 amino acids) or the vesicular stomatitis virus envelope glycoprotein (G) (60 amino acids) linked to the carboxy terminus of a nearly complete secretory polypeptide, growth hormone (GH), were used. In in vitro transcription-translation experiments, it was found that the insertion signal in the GH portion of the chimeras led to incorporation of the membrane protein segments into the ER membrane. Effectively, GH became part of the luminal segment of membrane proteins of which only very small segments, corresponding to the cytoplasmic portions of the G or HA proteins, remained exposed on the surface of the microsomes. When the chimeric genes were expressed in transfected cells, the products, as expected, failed to be secreted and remained cell-associated. These results support the assignment of a halt transfer role to segments of the membrane polypeptides that include their transmembrane portions. The hybrid polypeptide containing the carboxy-terminal portion of HA linked to GH accumulated in a juxtanuclear region of the cytoplasm within modified ER cisternae, closely apposed to the Golgi apparatus. The location and appearance of these cisternae suggested that they represent overdeveloped transitional ER elements and thus may correspond to a natural way station between the ER and the Golgi apparatus, in which further transfer of the artificial molecules is halted. The GH-G hybrid could only be detected in transfected cells treated with chloroquine, a drug that led to its accumulation in the membranes of endosome or lysosome-like cytoplasmic vesicles. Although the possibility that the chimeric protein entered such vesicles directly from the Golgi apparatus cannot be ruled out, it appears more likely that it was first transferred to the cell surface and was then internalized by endocytosis.
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PMID:Biosynthesis and intracellular sorting of growth hormone-viral envelope glycoprotein hybrids. 299 6

The mouse pituitary cell line, AtT-20, packages the adrenocorticotropic hormone (ACTH) in secretory vesicles and releases it when the cell is stimulated with secretagogues. These cells have the capacity, after transfection with the appropriate DNA, to package heterologous peptide hormones into the regulated secretory vesicles (Moore, H. P. H., M. D. Walker, F. Lee, and R. B. Kelly, 1983, Cell, 35:531-538). To test if other secreted proteins prefer a different route to the surface, we have transfected AtT-20 cells with DNAs coding for a fragment of a membrane protein, the vesicular stomatitis virus G protein from which the membrane spanning domain has been deleted (Rose, J. K., and J. E. Bergmann, 1982, Cell, 17:813-819). We found that the secreted vesicular stomatitis virus G proteins were not transported to the regulated secretory vesicles. Instead they preferentially exited the cell by the constitutive pathway previously found in these cells (Gumbiner, B., and R. B. Kelly, 1982, Cell, 28:51-59). In contrast, human growth hormone transfected into the cells by the same procedure was transported to the regulated pathway with a similar efficiency as the endogenous hormone ACTH. Transport of the secreted G protein to the regulated pathway, if it occurs at all, is at least 30-fold less efficient than peptide hormones. We conclude that the transport machinery in AtT-20 cells must selectively recognize different secreted proteins and sort them into distinct secretory pathways.
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PMID:Secretory protein targeting in a pituitary cell line: differential transport of foreign secretory proteins to distinct secretory pathways. 299 34

We previously demonstrated that occupancy of the epidermal growth factor (EGF) receptor reduced the ability of vaccinia virus to infect L cells [Eppstein et al: Nature 318:663, 1985]. This result suggested that vaccinia virus was utilizing the EGF receptor as one pathway to infect cells. We have studied this system further, and now find that antibodies to the EGF receptor also reduce the ability of vaccinia virus to infect cells productively. Inclusion of both EGF and antibodies to the EGF receptor did not cause inhibition over that obtained by EGF alone, providing another line of evidence that the antiviral effects on vaccinia virus were at the level of the EGF receptor. The antiviral effects of EGF or synthetic peptides corresponding to the third disulfide loop of TGF-alpha or the vaccinia virus growth factor were specific to vaccinia virus and did not inhibit replication of herpes simplex virus type 2 or vesicular stomatitis virus. The inhibitory effects on replication of vaccinia virus were obtained when EGF (but not insulin or growth hormone) was present prior to, but not after, productive viral adsorption. These results provided further evidence that the antivaccinia viral effects of EGF were at the level of initial receptor occupancy. As interferon (IFN) treatment has been shown to interfere with the action of some growth factors, including EGF, we examined the effects of IFN treatment of cells on the antivaccinia viral activity of EGF. Our results show that the antivaccinia effect of IFN-beta either interfered with or partially coalesced with the inhibitory effects of EGF.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Vaccinia virus and the EGF receptor: a portal for infectivity? 349 35

We have previously described the construction and expression of a hybrid gene encoding a membrane-anchored form of rat growth hormone. This protein is anchored in cellular membranes by a carboxy-terminal extension composed of the transmembrane and cytoplasmic domains of the vesicular stomatitis virus glycoprotein. The protein is transported efficiently to the Golgi apparatus but not to the cell surface. To examine the possibility that N-linked glycosylation might be required for protein transport to the cell surface, we created two mutant proteins (using in vitro mutagenesis) in which single amino acids at two random sites in anchored growth hormone were changed to generate consensus sequences required for addition of N-linked carbohydrate. These mutant proteins, and a protein with both glycosylation sites, were glycosylated and were transported to the cell surface. These results suggest that N-linked glycosylation can serve as a signal for protein transport to the cell surface.
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PMID:Glycosylation allows cell-surface transport of an anchored secretory protein. 392 68

Somatostatin is a14-amino acid peptide hormone that inhibits the secretion of a variety of other polypeptide hormones, including growth hormone. Here we describe an experimental system used to determine whether somatostatin can discriminate in its inhibition between secretory and plasma membrane proteins. Growth hormone-secreting cells (GH3) were infected with vesicular stomatitis virus and pulse-chased with [35S]methionine to follow the simultaneous intracellular transit of growth hormone and the viral membrane glycoprotein, G protein. Secretion of growth hormone was monitored by immunoprecipitation of chase media, while appearance of G protein on the plasma membrane was detected by cell surface labeling and virus purification. In the presence of somatostatin (10 micrograms/ml), the secretion of growth hormone was inhibited by 80%. In contrast, G protein appeared on the plasma membrane with slightly enhanced kinetics. When cells were treated with the ionophore monensin (0.2 microM), there was a dramatic inhibition of both the secretion of growth hormone and the incorporation of G protein into plasma membranes. Our results on the differential effect of somatostatin provide evidence for sorting of secretory and membrane proteins into distinct compartments in the secretory pathway. The data further suggest that this sorting event occurs late in the Golgi complex or after proteins exit from that organelle.
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PMID:Somatostatin discriminates between the intracellular pathways of secretory and membrane proteins. 614 20

We have carried out experiments designed to ask if it is possible to convert a secretory protein into an integral membrane protein by appending the membrane spanning domain of an integral membrane protein to its carboxy terminus. We first obtained expression of a cDNA clone encoding rat growth hormone (rGH) in eucaryotic cells, and found that this protein was secreted. We then constructed and expressed a hybrid gene encoding rGH fused to the membrane spanning and cytoplasmic domains of the vesicular stomatitis virus (VSV) glycoprotein (G). This fusion protein was anchored in microsomal membranes in the expected transmembrane configuration. The fusion protein was transported to the Golgi apparatus, and was esterified to palmitic acid, but it was not transported to the cell surface. We suggest that the sorting signal which allows rapid secretion of soluble rGH does not function when the protein is bound to the membrane.
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PMID:Conversion of a secretory protein into a transmembrane protein results in its transport to the Golgi complex but not to the cell surface. 658 49


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