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 examined the ability of recombinant human epidermal growth factor (EGF) and bradykinin (BK) to stimulate formation of inositol polyphosphates and sn-1,2-diacylglycerol (DAG), and mobilize intracellular Ca2+ ([Ca2+]i) in adult human keratinocytes (KC). Inositol polyphosphates were resolved by high performance liquid chromatography coupled with flow detector spectroscopy. Free intracellular calcium was quantitated using digital ratio imaging fluorescence microscopy of fura-2 loaded KC. The mass amount of DAG was quantitated using the DAG kinase reaction. When comparing maximal doses of BK (0.1 microM) and EGF (200 ng/ml), BK stimulated larger increases in all second messengers measured. The majority of cells responded rapidly to BK with global increases in [Ca2+]i. Cells responding to EGF were fewer in number and slower to respond with the Ca2+ signal being less pronounced. Treatment of cells with pertussis toxin (PTX) for 24 h significantly attenuated the BK-stimulated inositol polyphosphate formation and [Ca2+]i while the EGF response remained unaffected in both parameters. BK (10(-9) to 10(-6) M) did not stimulate DNA synthesis in KC as measured by [3H]-thymidine incorporation when cultures were treated for 5 days. These results demonstrate that the coupling and biochemical signals produced by stimulation of BK and EGF receptors in human KC are different and suggests that stimulation of second messenger formation from inositol lipid hydrolysis may not be an absolute requirement for the initiation of cell proliferation.
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PMID:Comparison of second messenger formation in human keratinocytes following stimulation with epidermal growth factor and bradykinin. 140 48

We have examined the possible involvement of pertussis toxin (PT)-sensitive GTP-binding protein and protein kinase C (PKC) in mitogen-induced tyrosine phosphorylation of the 41 kDa and 43 kDa cytosol proteins using PT-pretreated (inactivation of PT-sensitive GTP-binding protein) or phorbol 12-myristate 13-acetate (PMA)-pretreated (depletion of PKC) mouse fibroblasts. The effects of the inactivation of PT-sensitive GTP-binding protein and the depletion of PKC on mitogen-stimulated tyrosine phosphorylation of the proteins were similar and varied significantly and systematically in response to growth factors. The important finding was that such inhibitory effects of PT-sensitive GTP-binding protein inactivation and PKC depletion on protein tyrosine phosphorylation induced by each mitogen always correlated well with their inhibitory effects on each mitogen-stimulated DNA synthesis. Although the extent of platelet-derived-growth-factor-induced phosphorylation of the proteins was decreased to approx. 50% in PT- and PMA-pretreated cells compared with native cells, protein phosphorylation itself was not affected and occurred at identical sites on each protein in native, PT- and PMA-pretreated cells. These results suggest that: (1) 41 kDa and 43 kDa proteins are located downstream of PT-sensitive GTP-binding protein and PKC in the mitogenic signalling pathways of growth factors, (2) protein phosphorylation occurs via a cascade of events which includes the activation of the receptor tyrosine kinases, PKC and other unidentified kinase(s) which directly participate(s) in the phosphorylation of the 41 kDa and 43 kDa proteins, and (3) their phosphorylation may play an important role in integrating multiple mitogenic signalling pathways.
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PMID:Mitogen-induced tyrosine phosphorylation of 41 kDa and 43 kDa proteins. Potential role in integrating multiple mitogenic signalling pathways. 144 50

Three gene libraries of Bordetella avium 197 DNA were prepared in Escherichia coli LE392 by using the cosmid vectors pCP13 and pYA2329, a derivative of pCP13 specifying spectinomycin resistance. The cosmid libraries were screened with convalescent-phase anti-B. avium turkey sera and polyclonal rabbit antisera against B. avium 197 outer membrane proteins. One E. coli recombinant clone produced a 56-kDa protein which reacted with convalescent-phase serum from a turkey infected with B. avium 197. In addition, five E. coli recombinant clones were identified which produced B. avium outer membrane proteins with molecular masses of 21, 38, 40, 43, and 48 kDa. At least one of these E. coli clones, which encoded the 21-kDa protein, reacted with both convalescent-phase turkey sera and antibody against B. avium 197 outer membrane proteins. The gene for the 21-kDa outer membrane protein was localized by Tn5seq1 mutagenesis, and the nucleotide sequence was determined by dideoxy sequencing. DNA sequence analysis of the 21-kDa protein revealed an open reading frame of 582 bases that resulted in a predicted protein of 194 amino acids. Comparison of the predicted amino acid sequence of the gene encoding the 21-kDa outer membrane protein with protein sequences in the National Biomedical Research Foundation protein sequence data base indicated significant homology to the OmpA proteins of Shigella dysenteriae, Enterobacter aerogenes, E. coli, and Salmonella typhimurium and to Neisseria gonorrhoeae outer membrane protein III, Haemophilus influenzae protein P6, and Pseudomonas aeruginosa porin protein F. The gene (ompA) encoding the B. avium 21-kDa protein hybridized with 4.1-kb DNA fragments from EcoRI-digested, chromosomal DNA of Bordetella pertussis and Bordetella bronchiseptica and with 6.0- and 3.2-kb DNA fragments from EcoRI-digested, chromosomal DNA of B. avium and B. avium-like DNA, respectively. A 6.75-kb DNA fragment encoding the B. avium 21-kDa protein was subcloned into the Asd+ vector pYA292, and the construct was introduced into the avirulent delta cya delta crp delta asd S. typhimurium chi 3987 for oral immunization of birds. The gene encoding the 21-kDa protein was expressed equivalently in B. avium 197, delta asd E. coli chi 6097, and S. typhimurium chi 3987 and was localized primarily in the cytoplasmic membrane and outer membrane. In preliminary studies on oral inoculation of turkey poults with S. typhimurium chi 3987 expressing the gene encoding the B. avium 21-kDa protein, it was determined that a single dose of the recombinant Salmonella vaccine failed to elicit serum antibodies against the 21-kDa protein and challenge with wild-type B. avium 197 resulted in colonization of the trachea and thymus with B. avium 197.
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PMID:Cloning and sequencing of a gene encoding a 21-kilodalton outer membrane protein from Bordetella avium and expression of the gene in Salmonella typhimurium. 144 40

Hydrolysis by phospholipase C (PLC) of phosphatidylinositol 4,5-bisphosphate is a key mechanism by which many extracellular signalling molecules regulate functions of their target cells. At least eight distinct isozymes of PLC are recognized in mammalian cells. Receptor-controlled PLC is often regulated by G proteins, which can be modified by pertussis toxin in some cells but not in others. In the latter cells, PLC-beta 1, but not PLC-gamma 1 or PLC-delta 1, may be activated by members of the alpha q-subfamily of the G protein alpha-subunits. An unidentified PLC in soluble fractions of cultured human HL-60 granulocytes is specifically stimulated by G protein beta gamma subunits purified from retina and brain. Identification of a second PLC-beta complementary DNA (PLC-beta 2) in an HL-60 cell cDNA library prompted us to investigate the effect of purified G protein beta gamma subunits on the activities of PLC-beta 1 and PLC-beta 2 transiently expressed in cultured mammalian cells. We report here that PLC-beta 1 and PLC-beta 2 were stimulated by free beta gamma subunits and that PLC-beta 2 was the most sensitive to beta gamma stimulation. Thus stimulation of PLC by beta gamma subunits is isozyme-selective and PLC-beta 2 is a prime target of beta gamma stimulation. Activation of PLC-beta 2 by beta gamma subunits may be an important mechanism by which pertussis toxin-sensitive G proteins stimulate PLC.
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PMID:Isozyme-selective stimulation of phospholipase C-beta 2 by G protein beta gamma-subunits. 146 33

Thrombin at concentrations as low as 20 pM (0.002 U ml-1) was found to stimulate inositol phosphate levels in cultured human non-pigmented ciliary epithelial cells. Several other proteases, including trypsin and plasmin, had little or no effect, of several protease inhibitors tested, only those with specificity for thrombin blocked the effect. Studies with active site-blocked thrombin suggested that the esterolytic active site of thrombin is required for inositol phosphate stimulation, while gamma-thrombin, which has reduced binding affinity to fibrinogen also showed reduced effectiveness in stimulating inositol phosphates. In the presence of 10 mM LiCl, thrombin stimulated inositol monophosphate, inositol bisphosphate and inositol trisphosphate formation, with a prolonged rise of the first and transient early rises in the latter two species. Thrombin also elevated intracellular Ca2+ levels as measured with the fluorescent calcium probe, indo-1-AM. This elevation could be blocked by prior addition to cells of the thrombin inhibitor, hirudin, and was dependent upon extracellular Ca2+ for the maintenance of an elevated level in the presence of thrombin. Incorporation of thymidine into DNA in confluent cultures was also stimulated by thrombin, with a four-fold increase in incorporation at 35 hr in thrombin-treated cells compared to controls. The half-maximal concentration for this process was 0.25 U ml-1. Pretreatment with 100 ng ml-1 pertussis toxin greatly reduced the thrombin effect, which is consistent with a role for a G-protein in stimulation of DNA synthesis by thrombin.
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PMID:Thrombin stimulates inositol phosphate formation, intracellular calcium fluxes and DNA synthesis in cultured fetal human non-pigmented ciliary epithelial cells. 148 37

The expression of the pertussis toxin ptx operon is positively regulated in cis by a promoter region of about 170 base pairs and in trans by the bvg locus, which codes for the transcriptional activator protein BvgA. The promoter contains two direct repeats which are essential for its activity. When the position of these direct repeats relative to the transcription start point was changed, the activity of the promoter was strongly impaired. The repeated sequences therefore do not represent enhancer-like elements similar to those which have been identified in other positively regulated promoters; instead, the integrity of the whole promoter region seems to be an important feature of ptx regulation. A transcription interference assay was carried out to analyze in vivo binding of regulatory proteins to the ptx promoter. The results suggest that the direct repeats are the recognition sequence of a protein, which binds to them only under conditions in which the promoter is activated. In vitro DNA binding experiments with BvgA protein purified from an overproducing Escherichia coli strain were performed. However, no binding of BvgA to the ptx promoter was observed under conditions where binding of BvgA to the fha and bvg promoters occurred. This suggests that factors in addition to the bvg system are involved in the regulation of the Bordetella virulence regulon.
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PMID:Functional analysis of the pertussis toxin promoter. 148 51

The standard diagnostic methods for pertussis have several shortcomings. With the increased knowledge of the Bordetella pertussis genome a specific and conserved DNA sequence, present in about 70-80 copies in each genome, was selected for amplification with the polymerase chain reaction (PCR) technique in order to evaluate its diagnostic potential in children with suspected pertussis. The 400 basepair DNA sequence chosen was present and amplified in all 112 B. pertussis strains and in no other bacterial species examined. The specificity of the amplified material was documented by restriction enzyme cleavage. In nasopharyngeal aspirates a B. pertussis specific PCR product was visualized in 19/25 culture positive and in 5/50 culture negative children. In conclusion the present PCR assay for B. pertussis can be clinically useful and permit a specific diagnosis within 1 day after sampling. Further studies are requested to document its sensitivity, specificity and predictive value for positive and negative results.
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PMID:Amplification of DNA by the polymerase chain reaction for the efficient diagnosis of pertussis. 150 38

The prn gene encoding the 68 kDa protective outer-membrane protein of Bordetella bronchiseptica (P.68 pertactin) was cloned, sequenced and expressed in Escherichia coli. The gene was isolated by DNA:DNA hybridization experiments using a radioactively-labelled fragment of the homologous prn gene from Bordetella parapertussis. DNA sequence analysis reveals that the gene is capable of encoding a protein with a molecular mass of 93996 Da (P.94); this precursor molecule is processed to form the P.68 antigen on the surface of B. bronchiseptica. Heterologous expression of the full-length gene encoding P.94 in Escherichia coli results in similar processing, with the P.68 antigen targeted to the bacterial outer membrane. Comparison of P.94 with the P.93 and P.95 precursors, encoding homologous proteins from Bordetella pertussis and B. parapertussis, shows a high degree (greater than 90%) of homology. The major differences between all three proteins occur in the number of repeats of the two families (Gly-Gly-Xaa-Xaa-Pro)n and (Pro-Gln-Pro)n of reiterated sequence motifs.
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PMID:Cloning, nucleotide sequence and heterologous expression of the protective outer-membrane protein P.68 pertactin from Bordetella bronchiseptica. 152 10

A brief exposure of pancreatic islets to the cytokine interleukin-1 beta (IL-1 beta) induces an initial stimulatory phase, which is followed by inhibition of islet function and eventually beta-cell damage. In the present study we have investigated the effects of IRAP, a blocker of type I IL-1 receptor and actinomycin D, an inhibitor of DNA transcription, on both the stimulatory and inhibitory effects of IL-1 beta on rat pancreatic islets in vitro. The two test agents counteracted the initial stimulatory actions of IL-1 beta on both islet glucose-induced insulin release and glucose oxidation rates. Furthermore, cycloheximide, an inhibitor of protein synthesis, could also prevent the early IL-1 beta-induced stimulation of insulin release. When islets were exposed for 1 hr to IL-1 beta and studied after 12 hr, there was a 75% inhibition of glucose induced insulin release, a 50% decrease in glucose oxidation rates and a 30% decrease in (pro)insulin biosynthesis. These effects were completely counteracted by coincubation with IRAP or actinomycin D, but were not affected by coincubation with pertussis toxin. Islet exposure to IL-1 alpha also induced a 60-80% inhibition of glucose-induced insulin release after 12 hr. As observed with rIL-1 beta, IRAP was also able to block the suppressive effects of IL-1 alpha on islet function. Mouse islets exposed for 2 hr to IL-1 beta and studied after 12 hr presented a 50% decrease in the glucose-induced insulin release. This effect was completely blocked by coincubation with a rat monoclonal antibody generated against the type I mouse IL-1 receptor.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Role of receptor binding and gene transcription for both the stimulatory and inhibitory effects of interleukin-1 in pancreatic beta-cells. 153 17

Human breast cancer cells have been recently reported to produce endothelin (ET) 1. To investigate the potential regulation of ET production in breast cancer cells, we have measured the release of ET-like immunoreactivity from the T47D cell line in response to various paracrine/endocrine factors. Bombesin (0.1 microM) and cortisol (1 microM) stimulated maximal respective increases in IR-ET release to 580 and 369% of basal values after 6 h. The responses to cortisol and bombesin were additive. The response to bombesin was dose dependent with a median effective dose around 1 nM and was inhibited by the receptor antagonist [Leu13-psi-CH2NH-Leu14]bombesin. Pretreatment of T47D cells with pertussis toxin had no effect on bombesin-induced inositol lipid hydrolysis but inhibited ET-like immunoreactivity release in response to bombesin in the presence of glucocorticoid, by 56%. ET-1 (10 nM) and insulin-like growth factor (10 ng/ml) stimulated modest separate increases in DNA synthesis in human breast fibroblasts of 35 and 71%, respectively, but together exhibited a strong synergistic response to 905% of control values. This in vitro study demonstrates the potential for bombesin and glucocorticoid to regulate ET production in human breast cancer cells, which may in turn have a paracrine influence on neighboring stromal cell function.
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PMID:Bombesin and glucocorticoids stimulate human breast cancer cells to produce endothelin, a paracrine mitogen for breast stromal cells. 155 Nov 9


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