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)

Nanomolar concentrations of synthetic peptides corresponding to the calmodulin-binding domain of skeletal muscle myosin light chain kinase were found to inhibit calmodulin activation of seven well-characterized calmodulin-dependent enzymes: brain 61 kDa cyclic nucleotide phosphodiesterase, brain adenylate cyclase, Bordetella pertussis adenylate cyclase, red blood cell membrane Ca++-pump ATPase, brain calmodulin-dependent protein phosphatase (calcineurin), skeletal muscle phosphorylase b kinase, and brain multifunctional Ca++ (calmodulin)-dependent protein kinase. Inhibition could be entirely overcome by the addition of excess calmodulin. Thus, the myosin light chain kinase peptides used in this study may be useful antagonists for studying calmodulin-dependent enzymes and processes.
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PMID:Synthetic peptides based on the calmodulin-binding domain of myosin light chain kinase inhibit activation of other calmodulin-dependent enzymes. 290 35

This paper reports the solution conformation and calmodulin binding of a 43-residue peptide from the calmodulin-binding domain of Bordetella pertussis adenylate cyclase. The peptide (P225-267) was synthesized and 15N-labeled at specific amino acids. It binds calmodulin with an equilibrium dissociation constant of 25 nM. Assignment of the NMR spectrum of the free peptide and analysis of the NOE connectivities and secondary shifts of C alpha protons allowed us to identify a 10-amino acid fragment (Arg237 to Arg246) which is in rapid equilibrium between alpha-helical and irregular structures. Titration experiments showed that at substoichiometric molar ratios the two molecules are in intermediate exchange between free and bound conformations. Using 15N-edited methods we assigned a large part of resonances of the labeled residues in the bound peptide. Analysis of the chemical shift differences between free and bound states shows that the fragment Leu240-Ala257 is the most affected by the interaction. The proton spectra of the calmodulin, in the free and complexed states were extensively assigned using homonuclear experiments. Medium- and long-range NOE patterns are consistent with a largely conserved secondary and tertiary structure. The main changes in chemical shift of calmodulin resonances are grouped in six structural regions both in NH2- and COOH-terminal domains. Intermolecular NOE connectivities indicate that the NH2-terminal of the bound peptide fragment is engulfed in the COOH-terminal domain of calmodulin. The interaction geometry appears to be similar to those previously described for myosin light chain kinase or calmodulin kinase II fragments.
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PMID:Calmodulin binding of a peptide derived from the regulatory domain of Bordetella pertussis adenylate cyclase. 770 46

In these studies we show that stimulation of O2- generation by Tumor necrosis factor-alpha (TNF), or antibodies against the common beta 2 chain of leukocyte integrins (CD18), or LFA-1 (CD11a) displays a common and unique pattern of sensitivity or insensitivity to inhibitors of different signalling pathways. Both ways of stimulating neutrophil O2- generation were blocked by wortmannin, an inhibitor of myosin light chain kinase (Nakanishi, S., et al., 1992., J. Biol. Chem. 267, 2157-2163), and three different inhibitors of protein tyrosine kinases. Neither staurosporine, a protein kinase C inhibitor, nor Pertussis toxin, at concentrations which inhibited O2- generation in response to PMA, and FMLP, respectively, had any effect.
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PMID:Effect of inhibitors of distinct signalling pathways on neutrophil Q2- generation in response to tumor necrosis factor-alpha, and antibodies against CD18 and CD11a: evidence for a common and unique pattern of sensitivity to wortmannin and protein tyrosine kinase inhibitors. 809 58

The effects of zinc hydroxide on the respiratory burst and phagocytosis by rat neutrophils were examined. Zinc hydroxide induced an increase in oxygen consumption and O2- production. Electronmicroscopy showed that neutrophils engulfed zinc hydroxide particles by phagocytosis. Pertussis toxin (0.25, 0.5, 1.0 micrograms/ml) and EGTA (1, 2, 5 mM) inhibited zinc hydroxide-induced O2- production in a dose-dependent manner. The inhibitors of protein kinase C, 1-(5-isoquinolinesulfonyl)-2-methyl-piperazine and N-[2-(methylamino)ethyl]-5-isoquinolinesulfonamide inhibited zinc hydroxide-induced O2- production with IC50 values ranging between 10 microM and 25 microM. The inhibitory study using an inhibitor of myosin light chain kinase, 1-(5-iodo-naphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine, showed IC50 values ranging from 5 microM to 10 microM. These findings indicate that zinc hydroxide induces respiratory burst and phagocytosis by rat neutrophils.
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PMID:Zinc hydroxide induced respiratory burst in rat neutrophils. 815 83

Thrombin treatment causes a dose-dependent rounding of 1321N1 astrocytoma cells. This cytoskeletal response is rapid, peaking 2 h after thrombin stimulation, and reverses by 50% after 24 h. The thrombin receptor peptide SFLLRNP also induces cell rounding, whereas other G protein-linked receptor agonists such as carbachol, lysophosphatidic acid, or bradykinin fail to do so. Results of studies using pharmacological inhibitors do not support a requirement for phosphatidylinositol 3-kinase, mitogen-activated protein kinase, or Ca2+ mobilization in this response. Inhibition of protein kinase C or tyrosine kinase produces minimal blockade. Pertussis toxin treatment is also without effect. However, thrombin-induced rounding is fully blocked by the C3 toxin from Clostridium botulinum, which specifically ADP-ribosylates and inactivates the small G protein Rho. Thrombin also leads to a rapid, 2.4-fold increase in 32P incorporation into myosin light chain while carbachol does not. Myosin phosphorylation, like cell rounding is inhibited by inactivation of Rho with C3 exoenzyme, suggesting that myosin phosphorylation is necessary for this cytoskeletal response. This is supported by the observation that thrombin-induced rounding is also blocked by the myosin light chain kinase inhibitor KT5926. However, treatment with KT5926 fails to inhibit mitogenesis. Thus, cell rounding is not prerequisite to thrombin-induced DNA synthesis. We conclude that stimulation of the heterotrimeric G protein-coupled thrombin receptor in 1321N1 cells activates Rho-dependent pathways for both DNA synthesis and cell rounding, the cytoskeletal response being mediated in part through increases in myosin phosphorylation.
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PMID:Requirement for Rho-mediated myosin light chain phosphorylation in thrombin-stimulated cell rounding and its dissociation from mitogenesis. 955 56

To investigate the molecular mechanisms underlying transendothelial migration of tumor cells, an essential process for their hematogenous dissemination, we developed an in vitro model system that allows the separate monitoring of cell adhesion and transmigration processes. This system uses a human pre-B lymphoma cell line, Nalm-6, and a cultured mouse endothelial cell line, KOP2.16. Nalm-6 cells rapidly adhered to KOP2.16 and subsequently transmigrated underneath them. Using this model, we examined the effects on transendothelial migration, of various reagents which specifically interfere with the function of intracellular signal transduction molecules. Treatment of Nalm-6 cells with wortmannin (WMN), herbimycin A, pertussis toxin, or C3 exoenzyme of Clostridium botulinum, which specifically inhibit P13 kinase and/or myosin light chain kinase, herbimycin-sensitive tyrosine kinases, heterotrimeric G proteins, and the small G proteins, and the small G proteins rho/rac, respectively, reduced transmigration in a dose-dependent manner, Pretreatment of KOP2.16 endothelial cells with WMN also reduced transmigration in a dose-dependent manner. Binding of Nalm-6 binding to KOp2.16 was not affected, even when Nalm-6 or KOP2.16 cells were pretreated with these inhibitors, indicating that the reduction of transmigration was not due to a reduction of Nalm-6 to KOP2.16. These results also indicate that the signal transduction pathway(s) involved in transmigration can be dissociated from that of adhesion. Our results support the notion that endothelial cells are not a passive barrier in lymphoma extravasation, but that they assist lymphoma cell extravasation.
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PMID:Intracellular Signal-transducing elements involved in transendothelial migration of lymphoma cells. 968 62

Endothelial barrier function is regulated at the cellular level by cytoskeletal-dependent anchoring and retracting forces. In the present study we have examined the signal transduction pathways underlying agonist-stimulated reorganization of the actin cytoskeleton in human umbilical vein endothelial cells. Receptor activation by thrombin, or the thrombin receptor (proteinase-activated receptor 1) agonist peptide, leads to an early increase in stress fiber formation followed by cortical actin accumulation and cell rounding. Selective inhibition of thrombin-stimulated signaling systems, including Gi/o (pertussis toxin sensitive), p42/p44, and p38 MAP kinase cascades, Src family kinases, PI-3 kinase, or S6 kinase pathways had no effect on the thrombin response. In contrast, staurosporine and KT5926, an inhibitor of myosin light chain kinase, effectively blocked thrombin-induced cell rounding and retraction. The contribution of Rho to these effects was analyzed by using bacterial toxins that either activate or inhibit the GTPase. Escherichia coli cytotoxic necrotizing factor 1, an activator of Rho, induced the appearance of dense actin cables across cells without perturbing monolayer integrity. Accordingly, lysophosphatidic acid, an activator of Rho-dependent stress fiber formation in fibroblasts, led to reorganization of polymerized actin into stress fibers but failed to induce cell rounding. Inhibition of Rho with Clostridium botulinum exoenzyme C3 fused to the B fragment of diphtheria toxin caused loss of stress fibers with only partial attenuation of thrombin-induced cell rounding. The implication of Rac and Cdc42 was analyzed in transient transfection experiments using either constitutively active (V12) or dominant-interfering (N17) mutants. Expression of RacV12 mimicked the effect of thrombin on cell rounding, and RacN17 blocked the response to thrombin, whereas Cdc42 mutants were without effect. These observations suggest that Rho is involved in the maintenance of endothelial barrier function and Rac participates in cytoskeletal remodeling by thrombin in human umbilical vein endothelial cells.
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PMID:Regulation of the actin cytoskeleton by thrombin in human endothelial cells: role of Rho proteins in endothelial barrier function. 972 17

Contraction of normal esophageal circular muscle (ESO) in response to acetylcholine (ACh) is linked to M2 muscarinic receptors activating at least three intracellular phospholipases, i.e., phosphatidylcholine-specific phospholipase C (PC-PLC), phospholipase D (PLD), and the high molecular weight (85 kDa) cytosolic phospholipase A2 (cPLA2) to induce phosphatidylcholine (PC) metabolism, production of diacylglycerol (DAG) and arachidonic acid (AA), resulting in activation of a protein kinase C (PKC)-dependent pathway. In contrast, lower esophageal sphincter (LES) contraction induced by maximally effective doses of ACh is mediated by muscarinic M3 receptors, linked to pertussis toxin-insensitive GTP-binding proteins of the G(q/11) type. They activate phospholipase C, which hydrolyzes phosphatidylinositol bisphosphate (PIP2), producing inositol 1,4,5-trisphosphate (IP3) and DAG. IP3 causes release of intracellular Ca++ and formation of a Ca++-calmodulin complex, resulting in activation of myosin light chain kinase and contraction through a calmodulin-dependent pathway. Signal transduction pathways responsible for maintenance of LES tone are quite distinct from those activated during contraction in response to maximally effective doses of agonists (e.g., ACh). Resting LES tone is associated with activity of a low molecular weight (approximately 14 kDa) pancreatic-like (group 1) secreted phospholipase A2 (sPLA2) and production of arachidonic acid (AA), which is metabolized to prostaglandins and thromboxanes. These AA metabolites act on receptors linked to G-proteins to induce activation of PI- and PC-specific phospholipases, and production of second messengers. Resting LES tone is associated with submaximal PI hydrolysis resulting in submaximal levels of inositol trisphosphate (IP3-induced Ca++ release, and interaction with DAG to activate PKC. In an animal model of acute esophagitis, acid-induced inflammation alters the contractile pathway of ESO and LES. In LES circular muscle, after induction of experimental esophagitis, basal levels of PI hydrolysis are substantially reduced and intracellular Ca++ stores are functionally damaged, resulting in a reduction of resting tone. The reduction in intracellular Ca++ release causes a switch in the signal transduction pathway mediating contraction in response to ACh. In the normal LES, ACh causes release of Ca++ from intracellular stores and activation of a calmodulin-dependent pathway. After esophagitis, ACh-induced contraction depends on influx of extracellular Ca++, which is insufficient to activate calmodulin, and contraction is mediated by a PKC-dependent pathway. These changes are reproduced in normal LES cells by thapsigargin-induced depletion of Ca++ stores, suggesting that the amount of Ca++ available for release from intracellular stores defines the signal transduction pathway activated by a maximally effective dose of ACh.
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PMID:Signal transduction in esophageal and LES circular muscle contraction. 1078 May 77

The edema factor exotoxin produced by Bacillus anthracis is an adenylyl cyclase that is activated by calmodulin (CaM) at resting state calcium concentrations in infected cells. A C-terminal 60-kDa fragment corresponding to the catalytic domain of edema factor (EF3) was cloned, overexpressed in Escherichia coli, and purified. The N-terminal 43-kDa domain (EF3-N) of EF3, the sole domain of edema factor homologous to adenylyl cyclases from Bordetella pertussis and Pseudomonas aeruginosa, is highly resistant to protease digestion. The C-terminal 160-amino acid domain (EF3-C) of EF3 is sensitive to proteolysis in the absence of CaM. The addition of CaM protects EF3-C from being digested by proteases. EF3-N and EF3-C were expressed separately, and both fragments were required to reconstitute full CaM-sensitive enzyme activity. Fluorescence resonance energy transfer experiments using a double-labeled CaM molecule were performed and indicated that CaM adopts an extended conformation upon binding to EF3. This contrasts sharply with the compact conformation adopted by CaM upon binding myosin light chain kinase and CaM-dependent protein kinase type II. Mutations in each of the four calcium binding sites of CaM were examined for their effect on EF3 activation. Sites 3 and 4 were found critical for the activation, and neither the N- nor the C-terminal domain of CaM alone was capable of activating EF3. A genetic screen probing loss-of-function mutations of EF3 and site-directed mutations based on the homology of the edema factor family revealed a conserved pair of aspartate residues and an arginine that are important for catalysis. Similar residues are essential for di-metal-mediated catalysis in mammalian adenylyl cyclases and a family of DNA polymerases and nucleotidyltransferases. This suggests that edema factor may utilize a similar catalytic mechanism.
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PMID:An extended conformation of calmodulin induces interactions between the structural domains of adenylyl cyclase from Bacillus anthracis to promote catalysis. 1092 33

We compared stimulus-coupling pathways involved in bovine pulmonary artery (PA) and lung microvascular endothelial cell migration evoked by sphingosine-1-phosphate (S1P), a potent bioactive lipid released from activated platelets, and by vascular endothelial growth factor (VEGF), a well-recognized angiogenic factor. S1P-induced endothelial cell migration was maximum at 1 microM (approximately 8-fold increase with PA endothelium) and surpassed the maximal response evoked by either VEGF (10 ng/ml) (approximately 2.5-fold increase) or hepatocyte growth factor (HGF) (approximately 2.5-fold increase). Migration induced by S1P, but not by VEGF, was significantly inhibited by treatment with antisense oligonucleotides directed to Edg-1 and Edg-3 (endothelial differentiation gene) S1P receptors and by G protein modification. These strategies included pretreatment with pertussis toxin, or transfection with mini-genes encoding a betagamma subunit inhibitory peptide of the beta-adrenergic receptor kinase, or an 11-amino-acid peptide that inhibits G(1alpha2) signaling. Various strategies to interrupt Rho family signaling, including C(3) exotoxin, dominant/negative Rho, or the addition of Y27632, a cell-permeable Rho kinase inhibitor, significantly attenuated S1P- but not VEGF-induced migration. Conversely, pharmacologic inhibition of either myosin light chain kinase, src family tyrosine kinases, or phosphatidylinositol-3' kinase reduced basal endothelial cell migration and abolished VEGF-induced endothelial cell migration but did not inhibit the increase in S1P-induced migration. Whereas VEGF and S1P increased both p42/p44 extracellular regulated kinase and p38 mitogen-activated protein (MAP) kinase activities, only p38 MAP kinase inhibition significantly reduced VEGF- and S1P-stimulated migration. These data confirm S1P as a potent endothelial cell chemoattractant through G(1alpha2)-coupled Edg receptors linked to Rho-associated kinase and p38 MAP kinase activation. The divergence in signaling pathways evoked by S1P and VEGF suggests complex and agonist-specific regulation of endothelial cell angiogenic responses.
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PMID:Differential regulation of sphingosine-1-phosphate- and VEGF-induced endothelial cell chemotaxis. Involvement of G(ialpha2)-linked Rho kinase activity. 1141 36


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