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Query: UMLS:C0043167 (pertussis)
 19,595 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Escherichia coli containing a cloned gene encoding the Bordetella pertussis serotype 2 fimbrial subunit failed to produce detectable levels of the gene product in whole-cell extracts. To engineer plasmids capable of directing the expression in E. coli of high levels of this product, both as a pre-protein and as a methionylated mature form the upstream signals of the fimbrial subunit gene were replaced by the lambda P(L) and P(R) promoters and the E. coli atpE translational initiation region. These constructs did not result in the expression of fimbrial subunit at detectable levels in several E. coli strains including DH5. However, they did in E. coli CAG629, which is lon protease and heat shock protein deficient. Both pre-protein and methionylated mature protein had molecular weights of 25.0 kD, which indicated that correct processing of the leader sequence had occurred and thus that it was transposed across the inner membrane. Electron microscopic investigation of the cell surface of E. coli cells expressing either form of the fimbrial gene failed to detect the presence of filamentous structures. The methionylated mature form of the recombinant fimbrial subunit was purified to apparent homogeneity. After dialysis in appropriate conditions it was seen to autoassemble into protein polymers. Antibodies raised against polymerized recombinant subunit reacted weakly with whole B. pertussis serotype 2 fimbriae in immunodot blot assays. However, such antibodies reacted in Western blots equally well with the recombinant and wild-type form of the fimbrial subunit.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Engineering upstream transcriptional and translational signals of Bordetella pertussis serotype 2 fimbrial subunit protein for efficient expression in Escherichia coli: in vitro autoassembly of the expressed product into filamentous structures. 196 7

GTP-binding proteins (G proteins), predominantly located at the inner surface of the plasma membranes of mammalian cells, dissociate into their constituent alpha and beta gamma subunits upon stimulation of G protein-coupled receptors by agonists. In the present studies, cytoplasmic proteins which might have an affinity for the dissociated beta gamma subunits were investigated by means of beta gamma subunit-immobilized affinity-column (beta gamma-immobilized column) chromatography. When soluble fractions obtained from various materials including rat liver, bovine brain, and HL-60 cells were applied to a beta gamma-immobilized column, some proteins were specifically eluted from the column with high-salt and detergent-containing solutions. One of the beta gamma subunit-binding proteins, of which the molecular weight was approximately 93,000 on SDS-PAGE, appeared to be commonly present in all tissues tested. The 93-kDa beta gamma-binding protein was identified as 90-kDa heat shock protein, hsp90, based on the findings of its partial amino acid sequences and its immunoreactivity to a monoclonal anti-hsp90 antibody. The brain hsp90 inhibited beta gamma-supported pertussis toxin-catalyzed ADP-ribosylation of alpha subunits. The hsp90 was also capable of binding to beta gamma subunits which had been reconstituted into phospholipid vesicles. The binding of hsp90 to beta gamma subunits was inhibited by the addition of GDP-bound alpha subunits, but not by GTP gamma S-bound ones. These results suggested that hsp90 could associate functionally with free beta gamma subunits dissociated from trimeric G proteins in vitro.
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PMID:Association of the beta gamma subunits of trimeric GTP-binding proteins with 90-kDa heat shock protein, hsp90. 805 61

Parathyroid hormone (PTH) activates both adenylate cyclase and phospholipase C in target cells, and cloned PTH/PTH-related protein (PTHrP) receptor can mediate both responses when expressed in host cells such as LLC-PK1 renal epithelial cells. Because calcitonin (CT) is known to augment 70-kDa heat shock protein (HSP70) mRNA by an adenosine 3',5'-cyclic monophosphate (cAMP)-independent mechanism in LLC-PK1 cells, we examined regulation of HSP70 transcription by PTH in these cells. Like CT, human PTH-(1-34) [hPTH-(1-34); 10(-10) to 10(-7) M)] increased porcine HSP70 mRNA and human HSP70 promoter-chloramphenicol acetyltransferase (CAT) expression within 4 h in LLC-PK1 cells that stably express > or = 100,000 PTH/PTHrP receptors per cell. The effect of PTH on HSP70 mRNA was not mimicked by cAMP analogues, forskolin, phorbol esters, Ca2+ ionophores, or alpha-thrombin; was insensitive to pertussis toxin; and was not due to increased mRNA stability. The upregulation of HSP70 gene transcription by hPTH (and CT) was clearly observed even after deletion of the functional heat shock consensus element in the promoter region of the human HSP70/CAT reporter. Upregulation of HSP70 transcription via endogenous PTH receptors also was observed in the osteoblastic cell lines SaOS-2 and ROS 17/2.8. Regulation of HSP70 gene transcription by PTH may be a common cellular response to the hormone, which, in some cells, may not be mediated by activation of adenylate cyclase or protein kinase C.
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PMID:Regulation of HSP70 by PTH: a model of gene regulation not mediated by changes in cAMP levels. 876 37

In an aortic smooth muscle cell line, A10 cells, we investigated the effect of sphingosine 1-phosphate on the induction of heat shock protein 27 (HSP27), a low-molecular-weight heat shock protein. Sphingosine 1-phosphate significantly induced the accumulation of HSP27 in a pertussis toxin-sensitive manner. The effect was dose-dependent in the range between 0.1 and 30 microM. Sphingosine 1-phosphate stimulated an increase in the levels of mRNA for HSP27. Sphingosine 1-phosphate stimulated both p42/p44 mitogen-activated protein (MAP) kinase and p38 MAP kinase activation. PD98059, an inhibitor of the upstream kinase that activates p42/p44 MAP kinase, did not affect sphingosine 1-phosphate-stimulated HSP27 induction. In contrast, SB203580, an inhibitor of p38 MAP kinase, reduced sphingosine 1-phosphate-induced HSP27 induction. SB203580 reduced the levels of mRNA for HSP27 induced by sphingosine 1-phosphate. These results indicate that sphingosine 1-phosphate stimulates the induction of HSP27 via p38 MAP kinase activation in aortic smooth muscle cells.
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PMID:Sphingosine 1-phosphate regulates heat shock protein 27 induction by a p38 MAP kinase-dependent mechanism in aortic smooth muscle cells. 1041 91

There is evidence that the myocytes produce dynorphin and dynorphin-like peptides, which are kappa opioid receptor (kappa-OR) agonists. Activation of kappa-OR, a dominant opioid receptor in the heart, alters the cardiac function in vivo and in vitro. The observations suggest that the endogenous kappa-opioid peptides may act as autocrines or paracrine in regulation of cardiac functions. Myocardial ischemia is a common cause of heart disorders, which is manifested in decreased myocardial performance, arrhythmia and infarct. When myocardial ischemia occurs, the sympathetic discharge increases, which in turn increases the work-load and oxygen consumption. This exacerbates the situation induced by ischemia. One of the mechanisms with which the body protects against ischemia-induced injury/arrhythmia is inhibition of stimulation of beta-adrenoceptor (beta-AR), the receptor mediating the actions of sympathetic stimulation. kappa-Opioids inhibit the beta-AR activation. The inhibition of the beta-AR activation is due to inhibition of Gs-protein and to a lesser extent the adenylyl cyclase of the signaling pathway mediating beta-AR stimulation by a pertussis sensitive G-protein that mediates kappa-OR activation. Another mechanism against ischemia-induced injury is preconditioning, which is defined as prior exposures to ischemia or other insults make the heart more tolerant to subsequent and more severe insults. Protection occurs immediately or 1-3 days after preconditioning. kappa-OR mediates protection of preconditioning with ischemia or metabolic inhibition, one of the consequences of ischemia, in the heart. Activation of kappa-OR by U50488H, a selective kappa-OR agonist (pharmacological preconditioning with U50488H, UP), activates protein kinase C (PKC), opens K(ATP) channels and increases the production of heat shock proteins. Blockade of PKC, or closing of the K(ATP) channels or inhibition of the synthesis of the heat shock protein abolishes the cardioprotection of UP. The findings indicate the important roles of PKC, the K(ATP) channels and the heat shock protein in cardioprotection of UP. In addition, UP also attenuates the Ca(2+) overload, a precipitating cause of cardiac injury, induced by ischemic insults, indicating that UP may confer cardioprotection via at least partly attenuating the Ca(2+) overload. Most interestingly, blockade of the K(ATP) channels with channel blockers, that abolishes the delayed cardioprotection of UP, also attenuates the inhibitory effect of UP on Ca(2+) overload, suggesting that the cardioprotective effect of opening of the K(ATP) channels may be due at least partly to the prevention/attenuation of Ca(2+) overload.
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PMID:Roles of kappa opioid receptors in cardioprotection against ischemia: the signaling mechanisms. 1271 97

The endocannabinoid 2-arachidonoylglycerol (2-AG) enhances cell migration through the CB2 cannabinoid receptor. In this study, using an immunoprecipitation and mass spectrometry-based proteomic approach, we first identified the 90-kDa heat shock protein (Hsp90), a chaperone protein with novel signaling functions, as a CB2-interacting protein. The CB2/Hsp90 interaction was confirmed in human embryonic kidney 293 cells expressing transfected CB2 and in differentiated HL-60 cells expressing endogenous CB2, by coimmunoprecipitation and Western blot experiments, as well as by treatment with geldanamycin (GA), a specific Hsp90 inhibitor. Disruption of the CB2/Hsp90 interaction by treatment with GA or reducing Hsp90 levels with specific short interfering RNAs markedly inhibited 2-AG-induced cell migration, demonstrating that Hsp90 is crucial for 2-AG-induced cell migration. 2-AG treatment resulted in a CB2-mediated stimulation of Rac1 activity, and treatment with GA blocked 2AG-induced activation of Rac1. It is noteworthy that expression of the dominant-negative form of Rac1 reduced 2-AG-induced cell migration. These data demonstrate that 2-AG-induced activation of Rac1 is essential for 2-AG-induced cell migration, and the CB2/Hsp90 interaction is needed for 2-AG-induced activation of Rac1. Furthermore, 2-AG-induced Rac1 activation was sensitive to pertussis toxin treatment, hence involving G(i) proteins. In addition, treatment with GA significantly inhibited the CB2/Galpha(i2) interaction. As a whole, our data indicate that Hsp90 may serve as scaffold to keep the CB2 receptor and its signaling components, including Galpha(i2), in proximity, thus facilitating CB2-mediated signaling to cell migration through the G(i)-Rac1 pathway. By demonstrating that Hsp90 is essential for CB2-mediated signaling to cell migration, this study reveals a novel role of Hsp90 in the signaling events mediated by a G protein-coupled receptor.
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PMID:Involvement of the 90-kDa heat shock protein (Hsp-90) in CB2 cannabinoid receptor-mediated cell migration: a new role of Hsp-90 in migration signaling of a G protein-coupled receptor. 1769 52