Cell regulation最新文献

{"title":"Selective expression of high molecular weight basic fibroblast growth factor confers a unique phenotype to NIH 3T3 cells.","authors":"N Quarto,&nbsp;D Talarico,&nbsp;R Florkiewicz,&nbsp;D B Rifkin","doi":"10.1091/mbc.2.9.699","DOIUrl":"https://doi.org/10.1091/mbc.2.9.699","url":null,"abstract":"<p><p>The phenotypes of NIH 3T3 cells transfected with basic fibroblast growth factor (bFGF) cDNAs that express only the high molecular weight (HMW) forms of bFGF, the 18-kDa form, or all forms were examined. Cells producing the 18 kDa or all forms of bFGF were transformed at high levels of growth factor expression but were nontransformed at low levels. Cell producing low levels of HMW forms of bFGF were growth impaired when compared with the parental cells. These cells tended to form multinucleated giant cells, did not grow in soft agar, were nontumorigenic, had a normal bFGF receptor number, and had a nontransformed morphology. Cells expressing high levels of HMW bFGFs had a transformed morphology and were tumorigenic. These data suggest a specific functional role for HMWbFGF.</p>","PeriodicalId":9671,"journal":{"name":"Cell regulation","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1991-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1091/mbc.2.9.699","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"12825999","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Nerve growth factor induces the association of a 130-Kd phosphoprotein with its receptor in PC-12 pheochromocytoma cells.","authors":"M Ohmichi,&nbsp;S J Decker,&nbsp;A R Saltiel","doi":"10.1091/mbc.2.9.691","DOIUrl":"https://doi.org/10.1091/mbc.2.9.691","url":null,"abstract":"<p><p>To explore the molecular mechanisms of nerve growth factor (NGF) action, we have attempted to identify proteins that immunoprecipitate with the NGF receptor. An anti-NGF receptor antibody was developed that immunoprecipitated the 75-Kd receptor in PC-12 cells. In [35S]methionine-labeled cells lysed with nonionic detergent, immunoprecipitation with this antireceptor antisera specifically brought down several associated proteins, although prior treatment of cells with NGF produced no apparent change in the distribution of these proteins. However, in vitro phosphorylation assays of the immunoprecipitated complex revealed the presence of a serine kinase that phosphorylated two predominant substrates with Mrs of 60 and 130 Kd. Prior treatment of cells produced no change in the appearance of the 60-Kd phosphoprotein, but NGF did stimulate the appearance of the 130-Kd protein. This effect was observed with as little as 0.1 nM NGF and was maximal at 5 min, but declined thereafter. Prior treatment of cells with NGF did not increase the phosphorylation of enolase added exogenously to the immunoprecipitates, suggesting that this action of NGF may have reflected the hormone-dependent association of the 130-Kd protein with the receptor, rather than activation of a receptor-associated kinase. Thus the association of the NGF 75-Kd receptor with a 130-Kd protein may be involved in signal transduction for the growth factor, although the role of this receptor in the NGF-dependent tyrosine phosphorylation remains unclear.</p>","PeriodicalId":9671,"journal":{"name":"Cell regulation","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1991-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1091/mbc.2.9.691","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"12825998","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Regulation of Cl/HCO3 exchange in gastric parietal cells.","authors":"H A Thomas,&nbsp;T E Machen","doi":"10.1091/mbc.2.9.727","DOIUrl":"https://doi.org/10.1091/mbc.2.9.727","url":null,"abstract":"<p><p>Microspectrofluorimetry of the fluorescent indicators 2',7'-bis-(2-carboxyethyl)-5(and-6)carboxyfluorescein and 6-methoxy-N-(3-sulfopropyl)-quinolinium was used to measure intracellular pH (pHi), intracellular Cl (Cli), and transmembrane fluxes of HCO3 and Cl in single parietal cells (PC) in isolated rabbit gastric glands incubated in HCO3/CO2-buffered solutions. Steady-state pHi was 7.2 in both resting (50 microM cimetidine) and stimulated (100 microM histamine) PCs. Transmembrane anion (HCO3 or Cl) flux rates during Cl removal from or readdition to the perfusate were the same in resting and stimulated PCs. These rates increased at alkaline pHi, though this pHi dependence was small in the physiological range. Maximum velocity (Vmax) for Cl influx or HCO3 efflux was 80-110 mM/min at pHi 7.6-7.8, and the Km for extracellular concentrations of Cl (Clo) was 25 mM; in the physiological range (pHi 7.1-7.3), Vmax for anion fluxes was approximately 50 mM/min. Steady-state Cli in the unstimulated PC was 62 +/- 5 mM, but on histamine stimulation, Cli decreased rapidly to 25 mM and then increased back to a steady-state level of 44 mM. HCO3 fluxes due to Cl removal or readdition were completely blocked by 0.5 mM 4,4'-diisothiocyanatodihydrostilbene-2,2'-disulfonic acid (H2DIDS), but Cl fluxes were only inhibited by 80%. H2DIDS did not inhibit the decrease in Cli that occurred with histamine treatment. Diphenylamine carboxylate (0.5 mM) inhibited Cl flux by only 50% and caused no additional inhibition of Cl flux when used in conjunction with H2DIDS. Transmembrane anion fluxes during solution Cl removal or readdition occurred 80% through the anion exchanger at the basal membrane and 20% through other pathway(s), presumably the Cl channel in the apical membrane. We conclude that the increase in transport activity via the Cl/HCO3 exchanger that occurs during histamine-induced increases in HCl secretion is due mostly to the decrease in Cli. In the resting cell with Cli = 62 mM, Clo = 120 mM, pHi = 7.2, and extracellular pH = 7.4, the anion exchanger is poised near its thermodynamic equilibrium. During histamine stimulation Cli drops from 62 mM to 44 mM, the thermodynamic equilibrium of the anion exchanger at the basolateral membrane is disturbed, and the anion exchanger then exchanges cellular HCO3 for extracellular Cl. Cli serves a crucial regulatory role in stimulus-secretion coupling in the PC.</p>","PeriodicalId":9671,"journal":{"name":"Cell regulation","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1991-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1091/mbc.2.9.727","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"12906297","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Potential oncogenic effects of basic fibroblast growth factor requires cooperation between CUG and AUG-initiated forms.","authors":"B Couderc,&nbsp;H Prats,&nbsp;F Bayard,&nbsp;F Amalric","doi":"10.1091/mbc.2.9.709","DOIUrl":"https://doi.org/10.1091/mbc.2.9.709","url":null,"abstract":"<p><p>Normal adult bovine aortic endothelial cells were infected with various recombinant retroviruses expressing one, two, or three human basic fibroblast growth factor (bFGF) proteins normally synthesized by an alternative use of translation initiation codons. We show here that the constitutive expression of the AUG-initiated from (18 kDa) leads the transfected cells to form colonies in soft agar. The expression of the high molar weight (HMW) forms (22.5 and 21 kDa) initiated at one of the two CUG initiation codons allows cell immortalization, whereas the tumorigenic potential is reached when the three forms are constitutively expressed. Furthermore, we provide evidence that constitutive expression of (HMW) bFGF forms has a down-regulation effect on bFGF synthesis from the gene naturally active in parental endothelial cells.</p>","PeriodicalId":9671,"journal":{"name":"Cell regulation","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1991-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1091/mbc.2.9.709","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"12826000","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Identification of tyrosine kinase Trk as a nerve growth factor receptor.","authors":"A H Ross","doi":"10.1091/mbc.2.9.685","DOIUrl":"https://doi.org/10.1091/mbc.2.9.685","url":null,"abstract":"","PeriodicalId":9671,"journal":{"name":"Cell regulation","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1991-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1091/mbc.2.9.685","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"12825997","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Degradation of T-cell receptor chains in the endoplasmic reticulum is inhibited by inhibitors of cysteine proteases.","authors":"T Wileman,&nbsp;L P Kane,&nbsp;C Terhorst","doi":"10.1091/mbc.2.9.753","DOIUrl":"https://doi.org/10.1091/mbc.2.9.753","url":null,"abstract":"<p><p>The endoplasmic reticulum, or an organelle closely associated with it, contains proteases that can be used to remove partially assembled or improperly folded proteins. Very little is known at present about the types of protease that degrade these proteins. The beta chain and cluster of differentiation (CD)3 delta subunit of the human T-cell antigen receptor (TCR) are degraded shortly after synthesis. In this study Chinese hamster ovary (CHO) cells transfected with either beta or delta were incubated with a panel of protease inhibitors, and the rates of degradation of the transfected proteins were followed using chain-specific enzyme-linked immunosorbent assays (ELISAs). Of the protease inhibitors tested, degradation of both chains was highly sensitive to sulfhydryl reagents and peptidyl inhibitors of cysteine proteases. Concentrations of inhibitors that produced near complete inhibition of degradation in the endoplasmic reticulum did not cause gross changes in cellular ATP levels nor did they significantly slow constitutive secretion from CHO cells. The inhibitors did not affect the ability of CHO cells to synthesize and assemble disulphide-linked TCR zeta dimers. We conclude that the protease inhibitors were not toxic to cells and did not affect the biosynthetic activity of the endoplasmic reticulum. Furthermore, they did not alter the ability of the endoplasmic reticulum to deliver its content to the Golgi apparatus. Taken together, these results suggest that the cysteine protease inhibitors slow degradation in the endoplasmic reticulum through an action on cysteine proteases. The results imply that the endoplasmic reticulum contains cysteine proteases that can be used to remove retained proteins.</p>","PeriodicalId":9671,"journal":{"name":"Cell regulation","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1991-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1091/mbc.2.9.753","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"12997061","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Wound-factor-induced and cell cycle phase-dependent expression of 9E3/CEF4, the avian gro gene.","authors":"M Martins-Green,&nbsp;C Tilley,&nbsp;R Schwarz,&nbsp;C Hatier,&nbsp;M J Bissell","doi":"10.1091/mbc.2.9.739","DOIUrl":"https://doi.org/10.1091/mbc.2.9.739","url":null,"abstract":"<p><p>The gro genes encode for secreted proteins with sequence homologies to inflammatory mediators. Little is known about the function of these proteins or their regulation. The chicken gro (9E3/CEF4) is expressed abundantly in the cells of proliferating cultures but at very low levels in confluent cultures. In vivo, this gene is expressed in connective tissue and overexpressed at sites of injury, especially in areas of neovascularization. Here we provide a bridge between these observations by examining in culture the effect on 9E3 expression and DNA synthesis induced by cell damage and by addition of factors known to be released on wounding. We mimicked wounding by scraping swaths across confluent cultures of embryonic fibroblasts and determined the time dependence of expression of 9E3 mRNA and incorporation of 3H-thymidine. We find that 9E3 is (1) transiently expressed after \"wounding\" or serum-stimulation; (2) expressed in a cell cycle phase-dependent manner; it is triggered during the G0-G1 transition or early in G1 and subsides during S-phase; and (3) stimulated to high levels by a-fibroblast growth factor (aFGF), bFGF, transforming growth factor alpha (TGF alpha), and TGF beta, to intermediate levels by platelet-derived growth factor and not stimulated by epidermal growth factor. We also find that cells that are constantly cycling do not express 9E3, indicating that they skip either the portion of the cell cycle where 9E3 is induced or that they constitutively express a repressor of transcription or an RNA-degrading enzyme. Taken together, these observations suggest that the product of this gene could play more than one role in vivo. For example, in normal tissues the 9E3 protein could be involved in the exit of cells from the resting stage, whereas during wound healing the secreted protein or its cleavage products also could play a role in angiogenesis.</p>","PeriodicalId":9671,"journal":{"name":"Cell regulation","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1991-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1091/mbc.2.9.739","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"12906298","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Temperature-dependent tyrosine phosphorylation of microtubule-associated protein kinase in epidermal growth factor-stimulated human fibroblasts.","authors":"R Campos-González,&nbsp;J R Glenney","doi":"10.1091/mbc.2.8.663","DOIUrl":"https://doi.org/10.1091/mbc.2.8.663","url":null,"abstract":"<p><p>Treatment of normal human fibroblasts with epidermal growth factor (EGF) results in the rapid (0.5 min) and simultaneous tyrosine phosphorylation of the EGF receptor (EGFr) and several other proteins. An exception to this tyrosine phosphorylation wave was a protein (42 kDa) that became phosphorylated on tyrosine only after a short lag time (5 min). We identified this p42 kDa substrate as the microtubule-associated protein (MAP) kinase using a monoclonal antibody to a peptide corresponding to the C-terminus of the predicted protein (Science 249, 64-67, 1990). EGF treatment of human fibroblasts at 37 degrees C for 5 min resulted in the tyrosine phosphorylation of 60-70% of MAP kinase as determined by the percent that was immunoprecipitated with antiphosphotyrosine antibodies. Like other tyrosine kinase growth factor receptors, the EGFr is activated and phosphorylated at 4 degrees C but is not internalized. Whereas most other substrates were readily tyrosine phosphorylated at 4 degrees C, MAP kinase was not. When cells were first stimulated with EGF at 4 degrees C and then warmed to 37 degrees C without EGF, tyrosine phosphorylation of MAP kinase was again observed. Treatment of cells with the protein kinase C activator phorbol myristate acetate (PMA) also resulted in the tyrosine phosphorylation of MAP kinase, and again only at 37 degrees C. Tryptic phosphopeptide maps demonstrated that EGF and PMA both induced the phosphorylation of the same peptide on tyrosine and threonine. This temperature and PMA sensitivity distinguishes MAP kinase from most other tyrosine kinase substrates in activated human fibroblasts.</p>","PeriodicalId":9671,"journal":{"name":"Cell regulation","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1991-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1091/mbc.2.8.663","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"12828901","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Isoprenoid pathway activity is required for IgE receptor-mediated, tyrosine kinase-coupled transmembrane signaling in permeabilized RBL-2H3 rat basophilic leukemia cells.","authors":"G G Deanin,&nbsp;J R Pfeiffer,&nbsp;J L Cutts,&nbsp;M L Fore,&nbsp;J M Oliver","doi":"10.1091/mbc.2.8.627","DOIUrl":"https://doi.org/10.1091/mbc.2.8.627","url":null,"abstract":"<p><p>Previously, we reported that the isoprenoid pathway inhibitor, lovastatin, blocks the activation by IgE receptor cross-linking of 45Ca2+ influx, 1,4,5-inositol trisphosphate production, secretion, and membrane changes (ruffling, spreading) in intact RBL-2H3 rat basophilic leukemia cells. These results indicated that an isoprenoid pathway intermediate, very likely an isoprenylated protein, is importantly involved in the control of IgE receptor-mediated signal transduction. Here, we show that 20 h of pretreatment with lovastatin also inhibits antigen-induced secretion and membrane responses in streptolysin O-(SLO)-permeabilized cells. However, lovastatin does not inhibit secretion stimulated by the nonhydrolyzable GTP analog, GTP gamma S. Furthermore, the membrane responses to GTP gamma S persist, although in an attenuated form, in lovastatin-treated permeabilized cells. The relative insensitivity of GTP gamma S-induced responses to lovastatin was one of several indications that antigen and GTP gamma S may activate separate pathways leading to transmembrane responses in permeabilized cells. Further experiments showed that the beta-thio derivative of GDP, GDPBAS, inhibits the secretory and membrane responses to GTP gamma S, as expected for a GTP-binding protein-dependent signaling pathway, while having little effect on antigen-induced responses. Conversely, genistein blocks the secretory and membrane responses to antigen, as expected for a tyrosine kinase-dependent pathway, without altering the GTP gamma S-induced responses. From these results, and from additional data from cells treated with tyrphostins and sodium orthovanadate, we propose that IgE receptor-mediated secretion from permeabilized RBL-2H3 cells occurs by a tyrosine kinase-dependent pathway that requires isoprenoid pathway activity for function. We propose further that RBL-2H3 cells contain a separate GTP-binding protein-mediated signaling pathway whose direct activation by GTP gamma S is either independent of isoprenoid pathway activity or depends on the activity of an isoprenylated protein that is not significantly depleted after 20 h of lovastatin treatment.</p>","PeriodicalId":9671,"journal":{"name":"Cell regulation","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1991-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1091/mbc.2.8.627","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"12941205","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Down-regulation of cellular platelet-derived growth factor receptors induced by an activated neu receptor tyrosine kinase.","authors":"L Lehtola,&nbsp;M Nistér,&nbsp;E Hölttä,&nbsp;B Westermark,&nbsp;K Alitalo","doi":"10.1091/mbc.2.8.651","DOIUrl":"https://doi.org/10.1091/mbc.2.8.651","url":null,"abstract":"<p><p>The functional integration of growth factor signaling occurs at several levels in target cells. One of the most proximal mechanisms is receptor transmodulation, by which one activated receptor can regulate the expression of other receptors in the same cells. Well-established transregulatory loops involve platelet-derived growth factor (PDGF) down-regulation of epidermal growth factor (EGF) receptors and beta-type transforming growth factors modulation of PDGF receptors. We have studied the relationship between neu tyrosine kinase activation and the expression of the PDGF receptors in transfected NIH/3T3 cells. Expression of the neu oncogene, but not of the neu proto-oncogene, was associated with a decrease of PDGF alpha- and beta-receptors on the cell surface, as measured by [125-I]PDGF-AA and -BB binding. These results were corroborated by metabolic labeling and immunoprecipitation of the PDGF beta-receptors. PDGF alpha- and beta-receptor mRNAs were strongly decreased in the neu oncogene-transformed cells in comparison with control cells expressing the neu proto-oncogene. Down-regulation of the PDGF receptors and their mRNAs was also observed after EGF treatment of cells expressing a chimeric EGF receptor/neu receptor, where the neu tyrosine kinase is activated by EGF binding. These results show that the neu tyrosine kinase can down-modulate PDGF receptor expression, and the effect is mediated via decreased PDGF receptor mRNA levels.</p>","PeriodicalId":9671,"journal":{"name":"Cell regulation","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1991-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1091/mbc.2.8.651","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"12850282","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}