Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0043167 (pertussis)
 19,595 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have used monolayers of control 3T3 cells and 3T3 cells expressing transfected human neural cell adhesion molecule (NCAM) or chick N-cadherin as a culture substrate for PC12 cells. NCAM and N-cadherin in the monolayer directly promote neurite outgrowth from PC12 cells via a G-protein-dependent activation of neuronal calcium channels. In the present study we show that ganglioside GM1 does not directly activate this pathway in PC12 cells. However, the presence of GM1 (12.5-100 micrograms/ml) in the co-culture was associated with a potentiation of NCAM and N-cadherin-dependent neurite outgrowth. Treatment of PC12 cells with GM1 (100 micrograms/ml) for 90 min led to trypsin-stable increases in both beta-cholera toxin binding to PC12 cells and an enhanced neurite outgrowth response to N-cadherin. The ganglioside response could be fully inhibited by treatment with pertussis toxin. These data are consistent with exogenous gangliosides enhancing neuritic growth by promoting cell adhesion molecule-induced calcium influx into neurons.
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PMID:Ganglioside modulation of neural cell adhesion molecule and N-cadherin-dependent neurite outgrowth. 157 68

The interaction of pertussis toxin (PT) with cells and model membranes was investigated by examining PT-induced intoxication of Chinese hamster ovary cells and by studying the binding of PT and its subunits to phospholipid vesicles. Since certain bacterial toxins require an acidic environment for efficient interaction with membranes and subsequent entry into the cell, the requirement for an acidic environment for PT action was examined. PT, unlike bacterial toxins such as diphtheria toxin, did not require an acidic environment for efficient intoxication of Chinese hamster ovary cells. Potential modes by which PT might interact with biological membranes were studied by examining the binding of PT to a model membrane system. PT was found to be capable of interacting with phospholipid vesicles, however, efficient binding of the toxin to the vesicles occurred only in the presence of both ATP and reducing agent. The A subunit portion of the toxin bound preferentially to the vesicles while little binding of the B oligomer portion of PT to the model membranes was observed. Isolated A subunit, in the absence of the B oligomer, also bound to the vesicles with optimal binding occurring in the presence of reducing agent. After cleavage of the A subunit by trypsin, probably at Arg-181, Arg-182, and/or Arg-193, large fragments which lacked the C-terminal portion of the A subunit of PT no longer associated with the lipid vesicles. These results suggest that the A subunit of PT can interact directly with a lipid matrix and, if freed from the constraints imposed by the B oligomer, may be capable of interacting with cellular membranes.
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PMID:Interaction of pertussis toxin with cells and model membranes. 161 69

PC-3 human prostatic tumor sublines have been previously isolated which exhibit striking differences in their invasive and metastatic phenotypes. This work has been extended here to measure and compare the levels of kinesin, a microtubule-dependent translocator molecule, in the PC-3 sublines. Western blots, slot blots, radiolabeling, and immunoprecipitation analysis showed that kinesin was expressed in the highly invasive and metastatic sublines at levels which were elevated above the base-line levels observed in the parent PC-3 cells. In comparison, kinesin was not expressed in detectable amounts in the noninvasive cell lines. The conditioned medium of the metastatic PC-3 sublines contained a heat- and trypsin-sensitive protein which exhibited a dosage-dependent capacity to stimulate increased kinesin expression, type IV collagenase secretion, and invasion of Matrigel by the metastatic sublines. The noninvasive sublines failed to secrete a similar stimulatory factor(s) or respond to the conditioned medium of metastatic sublines. Various growth factors and cytokines tested (platelet-derived growth factor, epidermal growth factor, insulin-like growth factor, formylmethionineleucinephenylalanine) had no significant effect on either kinesin expression or protease secretion and invasion. Pertussis toxin blocked the stimulatory effects of the conditioned medium, but other agents known to interfere with adenylate cyclase pathways (i.e., cholera toxin, forskolin, 8-bromoadenosine) failed to block stimulation. The data show for the first time that kinesin, protease secretion, and the resulting invasion process may be regulated in a coordinated manner by an autocrine factor(s) which activates G-protein-dependent processes.
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PMID:Regulation of kinesin expression and type IV collagenase secretion in invasive human prostate PC-3 tumor sublines. 165 72

Autotaxin (ATX) is a potent human motility-stimulating protein that has been identified in the conditioned medium from A2058 melanoma cells. This protein has been purified to homogeneity utilizing a strategy involving five column steps. Homogeneity of ATX was verified by two-dimensional gel electrophoresis. The molecular size of ATX is 125 kDa, and it has an isoelectric point of 7.7 +/- 0.2. Purified ATX was digested with cyanogen bromide and trypsin, and the resulting ATX peptides were purified by reverse-phase high performance liquid chromatography. Eleven peptides were subjected to amino acid sequence analysis, and 114 residues were identified. The partial amino acid sequences and the amino acid composition obtained for ATX show that it does not exhibit any significant homology to known growth factors or previously described motility factors. At picomolar concentrations, ATX stimulates both random and directed migration of human A2058 melanoma cells. Pretreatment of the melanoma cells with pertussis toxin abolishes the response to purified ATX, indicating that ATX stimulates motility through a receptor acting via a pertussis toxin-sensitive G protein.
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PMID:Identification, purification, and partial sequence analysis of autotaxin, a novel motility-stimulating protein. 173 49

Thrombin is believed to activate platelets via cell surface receptors coupled to G proteins. In order to better understand this process, we have examined the interaction of thrombin with HEL cells, a leukemic cell line that has served as a useful model for studies of platelet structure and function. In HEL cells, as in platelets, thrombin stimulated inositol trisphosphate (IP3) formation and suppressed cAMP synthesis. Both events were inhibited by pertussis toxin with 50% inhibition occurring at a toxin concentration that ADP-ribosylated 50% of the Gi alpha subunits present in HEL cells. IP3 formation was also stimulated by a second serine protease, trypsin. The trypsin response was identical to the thrombin response in time course, magnitude, and pertussis toxin sensitivity, suggesting that a similar mechanism is involved. Agonist-induced changes in the cytosolic-free Ca2+ concentration were used to test this hypothesis. Both proteases caused a transient increase in intracellular calcium [Ca2+]i that could be inhibited with D-phenylalanyl-L-prolyl-L-arginine chloromethyl ketone thrombin. Exposure to either protease desensitized HEL cells against subsequent increases in [Ca2+]i and IP3 caused by the other, although responses to other agonists were retained. This loss of responsiveness persisted despite repeated washing of the cells and the addition of hirudin. Complete recovery occurred after 20 h and could be prevented with cycloheximide. These observations suggest that 1) HEL cell thrombin receptors, like those on platelets, are coupled to phospholipase C and adenylylcyclase by pertussis toxin-sensitive G proteins, 2) the G proteins involved are equally accessible to pertussis toxin in situ, 3) when access is limited to the outside of the cell the response mechanisms for thrombin and trypsin are similar, if not identical, despite the broader substrate specificity of trypsin, 4) both proteases cause persistent changes that may involve proteolysis of their receptors or associated proteins, and 5) desensitization of the thrombin response occurs at a step no later than the activation of phospholipase C and requires protein synthesis for recovery.
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PMID:Receptor and G protein-mediated responses to thrombin in HEL cells. 184 99

Trypsin digestion of pertussis toxin (PT) preferentially cleaved the S1 subunit at Arg-218 without detectable degradation of the B oligomer. The fragment produced, termed the tryptic S1 fragment, appears to remain associated with the B oligomer. Chymotrypsin digestion of PT also preferentially cleaved the S1 subunit without detectable degradation of the B oligomer. The chymotryptic S1 fragment possessed a slightly lower apparent molecular weight than the tryptic S1 fragment and was more accessible to the respective protease. Trypsin- and chymotrypsin-treated PT and PT required the presence of dithiothreitol and ATP for optimal enzymatic activity. Trypsin-treated PT showed approximately a 2-4-fold higher level of expression of ADP-ribosyltransferase and NAD-glycohydrolase activities than PT. Chymotrypsin-treated PT also exhibited approximately a 2-fold greater level of ADP-ribosyltransferase activity than PT. The observed increase in activity of protease-treated PT was due primarily to a shorter time for activation in PT mediated ADP-ribosylation of transducin. In addition, trypsin-digested PT possessed the same cytotoxic potential for Chinese hamster ovary cell clustering as PT. One possible role for the generation of a proteolytic fragment of the S1 subunit of PT would be to produce a catalytic fragment with increased efficiency for ADP-ribosylation of G proteins in vivo.
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PMID:Protease treatment of pertussis toxin identifies the preferential cleavage of the S1 subunit. 185 Jul 38

Mast cells appear to promote fibroblast proliferation, presumably through secretion of growth factors, although the molecular mechanisms underlying this mitogenic potential have not been explained fully by known mast cell-derived mediators. We report here that tryptase, a trypsin-like serine proteinase of mast cell secretory granules, is a potent mitogen for fibroblasts in vitro. Nanomolar concentrations of dog tryptase strongly stimulate thymidine incorporation in Chinese hamster lung and Rat-1 fibroblasts and increase cell density in both subconfluent and confluent cultures of these cell lines. Tryptase-induced cell proliferation appears proteinase-specific, as this response is not mimicked by pancreatic trypsin or mast cell chymase. In addition, low levels of tryptase markedly potentiate DNA synthesis stimulated by epidermal growth factor, basic fibroblast growth factor, or insulin. Inhibitors of catalytic activity decrease the mitogenic capacity of tryptase, suggesting, though not proving, the participation of the catalytic site in cell activation by tryptase. Differences in Ca++ mobilization and sensitivity to pertussis toxin suggest that tryptase and thrombin activate distinct signal transduction pathways in fibroblasts. These data implicate mast cell tryptase as a potent, previously unrecognized fibroblast growth factor, and may provide a molecular link between mast cell activation and fibrosis.
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PMID:Mast cell tryptase is a mitogen for cultured fibroblasts. 186 60

Adenylate cyclase (AC) toxin from Bordetella pertussis interacts with and enters eukaryotic cells to catalyze the production of supraphysiologic levels of cyclic AMP. Although the calmodulin-activated enzymatic activity (ability to convert ATP to cyclic AMP in a cell-free assay) of this molecule is calcium independent, its toxin activity (ability to increase cyclic AMP levels in intact target cells) requires extracellular calcium. Toxin activity as a function of calcium concentration is biphasic, with no intoxication occurring in the absence of calcium, low level intoxication (200-300 pmol of cyclic AMP/mg of Jurkat cell protein) occurring with free calcium concentrations between 100 nM and 100 microM and a 10-fold increase in AC toxin activity at free calcium concentrations above 300 microM. The molecule exhibits a conformational change when free calcium concentrations exceed 100 microM as demonstrated by shift in intrinsic tryptophan fluorescence, an alteration in binding of one anti-AC monoclonal antibody, protection of a fragment from trypsin-mediated proteolysis, and a structural modification as illustrated by electron microscopy. Thus, it appears that an increase in the ambient calcium concentration to a critical point and the ensuing interaction of the toxin with calcium induces a conformational change which is necessary for its insertion into the target cell and for delivery of its catalytic domain to the cell interior.
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PMID:Adenylate cyclase toxin from Bordetella pertussis. Conformational change associated with toxin activity. 189 34

The guanine nucleotide binding proteins (G proteins) that couple hormone and other receptors to a variety of intracellular effector enzymes and ion channels are heterotrimers of alpha, beta, and gamma subunits. One way to study the interfaces between subunits is to analyze the consequences of chemically cross-linking them. We have used 1,6-bismaleimidohexane (BMH), a homobifunctional cross-linking reagent that reacts with sulfhydryl groups, to cross-link alpha to beta subunits of Go and Gi-1. Two cross-linked products are formed from each G protein with apparent molecular masses of 140 and 122 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Both bands formed from Go reacted with anti-alpha o and anti-beta antibody. The mobility on sodium dodecyl sulfate-polyacrylamide gel electrophoresis is anomalous since the undenatured, cross-linked proteins have the same Stokes radius as the native, uncross-linked alpha beta gamma heterotrimer. Therefore, each cross-linked product contains one alpha and one beta subunit. Activation of Go by guanosine 5'-3-O-(thio)triphosphate (GTP gamma S) does not prevent cross-linking of alpha to beta gamma, consistent with an equilibrium between associated and dissociated subunits even in the presence of GTP gamma S. The same cross-linked products of Go are formed in brain membranes reacted with BMH as are formed in solution, indicating that the residues cross-linked by BMH in the pure protein are accessible when Go is membrane bound. Analysis of tryptic peptides formed from the cross-linked products indicates that the alpha subunit is cross-linked to the 26-kDa carboxyl-terminal portion of the beta subunit. The cross-linked G protein is functional, and its alpha subunit can change conformation upon binding GTP gamma S. GTP gamma S stabilizes alpha o to digestion by trypsin (Winslow, J.W., Van Amsterdam, J.R., and Neer, E.J. (1986) J. Biol. Chem. 261, 7571-7579) and also stabilizes the alpha subunit in the cross-linked product. Cross-linked G o can be ADP-ribosylated by pertussis toxin. This ADP-ribosylation is inhibited by GTP gamma S with a concentration dependence that is indistinguishable from that of the control, uncross-linked G o. These two kinds of experiments indicate that alpha o is able to change its conformation even though it cannot separate completely from beta gamma. Thus, although dissociation of the subunits accompanies activation of G o in solution, it is not obligatory for a conformational change to occur in the alpha subunit.
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PMID:Structural and functional studies of cross-linked Go protein subunits. 189 68

This paper describes the development of a murine bank of monoclonal antibodies against Bordetella pertussis toxin, filamentous hemagglutinin (FHA), pili, lipopolysaccharide (LPS), or outer membrane proteins (OMPs). Subunits S1, S2, S3 of pertussis toxin (PT) bound immunoglobulins and glycoproteins such as fetuin and haptoglobin in an unspecific manner. The specificity of monoclonal antibodies towards subunits S1, S2, S3 or S4 of PT could be demonstrated by using purified immunoglobulins or their Fab2 fragments. A set of FHA-specific monoclonal antibodies could be differentiated on the basis of their binding to the various breakdown products present in FHA preparations. Pili-specific monoclonal antibodies reacted with either native pili or denatured pilin, and both demonstrated serotype specificity. Monoclonal antibodies to Bordetella pertussis OMPs were directed to either the virulent phase-regulated trypsin-sensitive, detergent-extractable OMPs 92 kDa, 32 kDa, and 30 kDa or the non-virulent phase-expressed, not-trypsin sensitive OMPs 38 kDa, 33kDa, and 18 kDa.
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PMID:Description of a hybridoma bank towards Bordetella pertussis toxin and surface antigens. 198


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