Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The dominant view has been that there is little or no activation of Type 2 cytokine production in human tuberculosis. A novel approach to quantitative nested reverse transcriptase-polymerase chain reaction has revealed that this conclusion was based on technical inadequacies of earlier studies, particularly the failure to discriminate between IL-4 and the IL-4 splice variant, IL4delta2. A new approach reveals that the largest cytokine change in tuberculosis is a 1-2 log increase in copy number for mRNAs encoding IL-4 and IL-13, accompanied by a small decrease in expression of mRNA encoding interferon-gamma. The increased IL-4 level correlates with disease severity and with serum levels of IgE and soluble CD30, and may be attributable to the recently observed increase in conversion of cortisone into cortisol in tuberculous lesions. The implications of these findings for pathogenesis, vaccine design and immunotherapy are discussed, as effective reagents will need to downregulate this inappropriate Th2 component.
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PMID:High levels of mRNA encoding IL-4 in unstimulated peripheral blood mononuclear cells from tuberculosis patients revealed by quantitative nested reverse transcriptase-polymerase chain reaction; correlations with serum IgE levels. 1123 43

The complementary DNAs encoding tropomyosins of the abalone Haliotis diversicolor, the scallop Chlamys nobilis, and the mussel Perna viridis were amplified by reverse transcriptase polymerase chain reaction and thereafter cloned in plasmid vectors for expression. Immunoblot analysis showed that recombinant proteins of abalone, scallop, and mussel tropomyosin were reactive to serum IgE antibodies from subjects allergic to shellfish but not to nonallergic controls. Nucleotide and amino acid sequences of abalone, scallop, and mussel tropomyosins are highly similar (mostly > 55%) to those of crustacean allergens identified as tropomyosins. Absorption experiments showed that recombinant tropomyosins from the 3 mollusks were able to remove serum IgE reactivity against the 38-kDa tropomyosin of the organisms. These results demonstrate that tropomyosin is the major allergen among various common edible mollusks. Yet a comparison between the amino acid sequences of putative epitopes in crustaceans and mollusks suggests that the epitopes in the two groups may be distinct. Parsimony analysis of the nucleotide sequences of tropomyosin from different mollusks suggests that the tropomyosin gene sequence is a useful tool for phylogenetic analysis of this group of animals.
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PMID:Tropomyosin Is the Major Mollusk Allergen: Reverse Transcriptase Polymerase Chain Reaction, Expression and IgE Reactivity. 1124 17

Because an airway-like inflammation has been reported in the gut of asthmatic patients, we sought to examine the expression of immunoregulatory cytokines like IL-4, IL-10, and IL-13 by gut mucosa. To establish this, we initiated this study to examine mRNA expressions of IL-4, IL-10, and IL-13 in duodenal mucosa from patients with asthma. Duodenal biopsy specimens were obtained from 20 asthmatic patients (10 allergic, 10 nonallergic) and 8 healthy controls. Cytokine mRNA was quantified with reverse transcriptase-competitive PCR, and results were expressed in proportion to the number of beta-actin mRNA in the same sample. IL-10 and IL-4 mRNA were detectable in all patients, whereas no IL-13 mRNA was detected. IL-10 mRNA concentrations were significantly higher in allergic subjects with asthma than in control subjects and nonallergic subjects with asthma. No significant difference was observed for IL-4. IL-10 mRNA expression was not related to asthma severity, FEV(1), blood eosinophilia, or IgE levels. Our results support the hypothesis that IL-10 overexpression may counterbalance the effects of proinflammatory cytokines and mitigate the inflammatory reaction found in gut mucosa of subjects with asthma.
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PMID:Overexpression of IL-10 mRNA in gut mucosa of patients with allergic asthma. 1129 68

Haematopoietic cell-specific transmembrane-4 (HTm4) is a four-transmembrane protein most closely related to CD20 and the beta subunit of the high affinity receptor for IgE (Fc(epsilon)RIbeta). To date, it has only been described in humans, where it is expressed in haematopoietic cells of both myeloid and lymphoid lineages. The function of HTm4 is unknown; however, as for CD20 and Fc(epsilon)RI-beta, it is likely to play a role in signal transduction as part of a multi-subunit cell surface receptor complex. In this study, we report the cDNA cloning and expression distribution of mouse HTm4. The deduced mouse HTm4 protein is of 213 amino acids, and contains four putative transmembrane domains. Mouse HTm4 shows 62% overall amino acid identity with human HTm4; the transmembrane regions are highly conserved between both species (75% identity), whereas the N- and C-terminal and inter-transmembrane loop regions are more divergent (52%). Interestingly, the N-terminal domain of mouse HTm4 is predicted to be 23 amino acids shorter, and the C-terminal domain 23 amino acids longer, than that of human HTm4. Northern blot and reverse transcriptase (RT)-PCR analysis suggest that mouse HTm4 mRNA is expressed at low levels only in spleen, bone marrow and peripheral blood leucocytes. This is the first report of the cloning of HTm4 from a species other than human, and provides important sequence information towards the understanding of the function of this poorly characterized four-transmembrane molecule.
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PMID:Cloning and characterization of a mouse homologue of the human haematopoietic cell-specific four-transmembrane gene HTm4. 1148 81

FE-3 cells were established by Hanashiro et al. by hybridizing mouse myeloma cells (Sp2/0-Ag14/SF) with rat spleen cells that were freshly isolated from Brown-Norway rats sensitized with DNP-As. FE-3 cells can constitutively secrete IgE without stimulation by cytokines. Our preliminary experiments demonstrated that the IgE secretion was decreased at 3 days after start of culture and the addition of exogenous IgE into culture media depressed the secretion of IgE. Thus, we hypothesized that the IgE production in FE-3 cells may be regulated by a signal transduction through the binding of IgE to its high affinity receptor (Fc(epsilon)RI) or to an IgE binding protein on the cell surface. In this study, we aimed to identify the nucleotide sequence of IgE FE-3 and compared with those of mouse IgE and IgE IR162 to find a structural heterogeneity in the Fc region of IgE FE-3. We also tested if the mRNA of Fc(epsilon)RI was expressed in FE-3 cells using the reverse transcriptase-polymerase chain reaction (RT-PCR) method with the combination of sequencing analysis. Consequently, the cDNA sequence of IgE FE-3 was identical to that of the CH3 and CH4 domains in the epsilon-chain of rat IgE IR162, whereas the cDNA of Fc(epsilon)RI was identical to that of mouse, suggesting that the genes of IgE FE-3 and Fc(epsilon)RI was derived from that of rat spleen cells and mouse myeloma cells, respectively.
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PMID:The nucleotide sequence of dinitrophenyl-specific IgE and Fc(epsilon)RI alpha-subunit obtained from FE-3 hybridoma cells. 1183 54

The elevation of non-specific IgE (total IgE) in Ascaris infection can be seen one week after infection, and reaches a peak after approximately two weeks. It has been reported that ABA-1 protein is the main constituent in the pseudocoelomic fluid of Ascaris suum. To investigate the effect of the ABA-1-like protein from Ascaris lumbricoides (ALB), the cDNA was cloned by reverse transcriptase polymerase chain reaction, using original primers based on the consensus sequences of ABA-1 and TBA-1, that is an ABA-1-like protein from Toxocara canis. The clone was sequenced, we constructed the recombinant polyprotein of ALB (rALB14 and rALB7) based on the ALB sequence, and rALB was administrated to BALB/c mice. Fourteen days after inoculation with rALB14 which is the full length of ALB, the elevation of total IgE which we supposed to contain non-specific IgE was observed, and the results were as we expected. Furthermore, in an in-vitro experiment, we confirmed that the spleen cells proliferated when stimulated by rALB14 and concanavalin A. Therefore, the whole conformation of ALB is considered to be involved in the elevation of non-specific IgE, and is involved in the activation of T cells.
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PMID:The biological activity of ABA-1-like protein from Ascaris lumbricoides. 1216 Feb 20

Mast cells are immunoregulatory effector cells capable of releasing different mediators and cytokines implicated in inflammatory tissue processes. Previous studies suggested that IL-3 regulates growth and function of murine mast cells and human mast cell precursors, but does not affect mature human mast cells. In the present study, we found expression of IL-3 receptors (IL-3R) in freshly isolated human intestinal mast cells by reverse transcriptase (RT)-PCR and in mast cells cultured with stem cell factor (SCF) using RT-PCR and flow cytometry. IL-3R expression was enhanced when the culture medium was supplemented with IL-4 in addition to SCF. In the presence of SCF, IL-3 significantly enhanced mast cell growth in a dose-dependent fashion (179+/-51% of control, p</=0.004, n=9, ED(50) approximately 15 ng/ml) by decreasing mast cell apoptosis rather than inducing proliferation. Furthermore, IL-3 selectively enhanced histamine (from 39.6+/-12.4 to 51.2+/-15.7% specific release, p<0.02, n=8) and leukotriene C(4) (LTC(4), 5.1+/-3.4 to 10.8+/-5.5 ng/10(6) mast cells, p<0.03, n=6) release triggered by IgE receptor cross-linking without affecting prostaglandin D(2) production. In conclusion, our data show that human intestinal mast cells express functional IL-3R, indicating that IL-3 not only regulates growth and function of immature, but also that of mature human mast cells.
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PMID:Cultured human intestinal mast cells express functional IL-3 receptors and respond to IL-3 by enhancing growth and IgE receptor-dependent mediator release. 1220 44

Rhodotorula mucilaginosa (also known as R. rubra) is among the most commonly found yeast strains in our environment. However, allergens from R. mucilaginosa have not yet been characterized at the molecular level. The purpose of this study was to characterize the enolase allergen from R. mucilaginosa and examine the allergenic/antigenic cross-reactivity among fungal enolases. The full-length cDNA encoding the R. mucilaginosa enolase was isolated through the reverse transcriptase-polymerase chain reaction in conjunction with the 5'-end and 3'-end rapid amplification cDNA end reactions. The corresponding natural enolase from R. mucilaginosa was identified using two-dimensional gel electrophoresis and N-terminal amino acid sequence analysis. The results showed that the enolase from R. mucilaginosa is a protein of 439 residues and is encoded by a cDNA of 1497 bp. It shares high sequence identity with enolase allergens from Candida albicans (85%), Saccharomyces cerevisiae (76%), Penicillium citrinum (76%), Aspergillus fumigatus (76%), Cladosporium herbarum (76.5%), and Alternaria alternata (74%). A 47-kD component in the R. mucilaginosa extracts was found to react with IgE or rabbit anti-enolase antiserum and has an N-terminal amino acid sequence identical to that deduced from the isolated enolase cDNA. Sera from three (21%) of 14 allergic patients sensitized to R. mucilaginosa showed IgE binding to this 47-kD R. mucilaginosa component and the His-tagged recombinant enolase. A rabbit antiserum against the P. citrinum enolase and a monoclonal antibody (MoAb; Afueno 8) against the A. fumigatus enolase reacted with all 5 fungal enolases tested. However, an MoAb (E2a) generated by using the Saccharomyces enolase as antigen could only recognize the immunizing enolase. In addition, heterogeneity in immunoblot profiles of IgE antibodies in serum samples from 9 allergic patients against 5 different fungal enolases tested was also observed. The presence of IgE cross-reactivity among enolase allergens from R. mucilaginosa, C. albicans and P. citrinum was detected by immunoblot inhibition. In conclusion, a new and cross-reactive enolase allergen from R. mucilaginosa (Rho m 1) was identified. Although enolases are highly conserved allergens among different fungal species, most of the allergic patients examined in this study differed in their IgE reactivity to the 5 different fungal enolases tested. The results obtained will be of value in understanding the role of enolase allergen in clinical mould allergy.
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PMID:Characterization of enolase allergen from Rhodotorula mucilaginosa. 1243 31

Mast cells play a central role in IgE-dependent allergic responses. Although they have been reported to express only low-affinity IgG receptors and no high-affinity receptors (FcgammaRI), our recent study showed that canine mastocytoma CM-MC cells are activated by monomeric canine IgG, suggesting the presence of FcgammaRI on CM-MC cells. In the present study, we measured the affinity of canine IgG with CM-MC cells, determined the presence of the FcgammaRI protein and mRNA, and identified the cDNA sequence of it. The results showed that (7.5+/-3.1)x10(4) receptor molecules are expressed on a CM-MC cell with a Ka of (9.1+/-1.6)x10(7)M(-1) for binding to monomeric canine IgG. Canine IgG-conjugated beads precipitated an approximately 72-kDa surface protein, whose size is consistent with that of the FcgammaRI alpha subunit of humans and mouse. The expression of FcgammaRI mRNA was detected by reverse transcriptase polymerase chain reaction (RT-PCR), and the cDNA encoding the FcgammaRI alpha subunit was found to be 84% and 78% similar to that of humans and the mouse, respectively. The predicted amino acid sequence was 72% and 63% identical, respectively. Canine mastocytoma CM-MC cells are therefore very useful for studying FcgammaRI-mediated signal transduction in mast cells.
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PMID:Presence and primary sequence of a high-affinity IgG receptor on canine mastocytoma (CM-MC) cells. 1281 28

We investigated the expression and function of matrix metalloproteinase-12 (MMP-12) in a model of allergic airway inflammation. Mice were sensitized mucosally by exposure to aerosolized ovalbumin (OVA) daily over a period of 10 d in the context of adenovirus-mediated granulocyte macrophage colony-stimulating factor (GM-CSF) expression. The ensuing inflammatory response is characterized by a Th2 cytokine profile, OVA-specific IgE, and airway eosinophilia. Using real-time, quantitative reverse transcriptase-polymerase chain reaction we assessed MMP-12 mRNA expression in whole lung tissue. We observed a 12- and 70-fold increase in expression at Days 7 and 11, respectively, in OVA-exposed mice when compared with naive controls. Immunoblot analysis revealed an increase in MMP-12 protein in the bronchoalveolar lavage fluid of mice exposed to OVA in the context of GM-CSF. No such elevation was observed in mice exposed to saline only in the context of GM-CSF. To assess functional role of MMP-12, MMP-12 knockout (KO) mice were subjected to the aforementioned protocol. We observed an 80% reduction in eosinophils in the bronchoalveolar lavage fluid of KO mice compared with their wild-type littermates. Using interleukin-13 KO mice, we demonstrated that expression of MMP-12 is interleukin-13-dependent. Collectively, our data indicate a novel function for MMP-12 in the process of airway eosinophil accumulation.
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PMID:Interleukin-13-dependent expression of matrix metalloproteinase-12 is required for the development of airway eosinophilia in mice. 1284 50


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