Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.30.1 (S1 nuclease)
 3,660 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Using a monoclonal antibody to the interleukin 3 (IL-3) receptor (anti-Aic2), we isolated a cDNA (AIC2B) from a mouse mast cell line which is homologous to the previously characterized gene for the IL-3 receptor (AIC2A). This cDNA encodes a polypeptide of 896 amino acid residues and has 91% amino acid sequence identity with the IL-3 receptor. A consensus sequence defining an additional cytokine receptor family is present in this clone. Compared to the AIC2A clone, the AIC2B cDNA encodes a protein with amino acid substitutions, insertions, and deletions dispersed throughout the entire protein. Oligonucleotide probes specific for each cDNA hybridized with different genomic fragments, indicating that the AIC2A and AIC2B proteins are encoded by two distinct genes. Fibroblasts transfected with the AIC2B cDNA expressed the protein at the cell surface as determined by binding with the anti-Aic2 antibody but did not bind IL-3 or other cytokines, including IL-2, IL-4, granulocyte-macrophage colony-stimulating factor, erythropoietin, and IL-9 (p40) at concentrations between 1 and 10 nM. An S1 nuclease protection assay was used to discriminate between the AIC2A and AIC2B transcripts. We found that the AIC2B gene was coexpressed with the AIC2A gene. These results suggest a potential involvement of AIC2B in cytokine signal transduction.
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PMID:Cloning and expression of a gene encoding an interleukin 3 receptor-like protein: identification of another member of the cytokine receptor gene family. 169 79

Anti-receptor antibodies have previously been used in two cytokine systems (IL-1 and TNF alpha) to identify the existence of different cytokine receptors on different cell types. In this study, we have similarly used two approaches to evaluate whether IL-4 receptors on different cell types are identical, or whether more than one species of IL-4 receptor exists. The first approach involved production of monoclonal antibodies specific for the IL-4 receptor expressed by the murine mast cell line, MC/9. Six anti-IL-4 receptor monoclonal antibodies were produced against the purified soluble extracellular domain of the recombinant IL-4 receptor derived from MC/9 cells. These antibodies were capable of binding to and specifically immunoprecipitating the soluble extracellular domain of the recombinant mast cell IL-4 receptor. Following biotinylation of the antibodies and addition of phycoerythrin-streptavidin, their binding to cell associated IL-4 receptors on MC/9 mast cells could be readily visualized by immunofluorescence. Using this approach, the anti-mast cell IL-4R antibodies were found to specifically bind IL-4 receptors expressed on a variety of other murine cell types, including T cells, B cells, macrophages, fibroblasts, and L cells. The antibodies did not bind to two human cell lines known to bind human but not murine IL-4. The intensity of staining was directly related to the number of IL-4 binding sites identified previously by receptor-ligand equilibrium binding analyses. As a second approach to evaluating potential receptor heterogeneity, we constructed S1 nuclease protection assay probes for two separate regions of the mast cell IL-4 receptor, one located in the extracellular domain and one in the intracellular domain. Subsequent S1 analyses showed that both regions are expressed by the following types of cells: T cells, B cells, macrophages, myeloid cells, L cells, and stromal cells. The two approaches used in this study therefore indicate that the same or highly similar IL-4 receptor species is expressed by a wide variety of hemopoietic and nonhemopoietic cells. Since the anti-IL-4 receptor antibodies produced in this study did not block binding of IL-4 to its receptor, we cannot exclude the possible existence of a second type of IL-4R coexpressed on the cells tested in this study, or expressed uniquely by other cell types that were not investigated.
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PMID:Evaluation of murine interleukin 4 (IL-4) receptor expression using anti-receptor monoclonal antibodies and S1 nuclease protection analyses. 206 18

In this report, we analyze the expression of the type II receptor for the Fc region of IgG (Fc gamma RII) in resting and lipopolysaccharide (LPS)-activated murine B lymphocytes. Fc gamma RII is encoded by two genes, alpha and beta. The beta gene encodes two mRNA, beta 1 and beta 2, which are generated by alternative splicing. Using an S1 nuclease protection assay, we found that resting and activated B lymphocytes express predominantly the beta 1 transcript. Very low levels of the beta 2 mRNA were detected in this assay, while no expression of the alpha transcript could be detected. Quantitative Northern blot analysis showed that the amount of Fc gamma RII beta mRNA was increased 9-fold in LPS-activated B lymphocytes. The expression of Fc gamma RII during the various phases of B cell activation was then studied by immunofluorescence using the monoclonal antibody 2.4G2. LPS stimulation induced an increase of the Fc gamma RII cellular pool as well as of its expression at the surface of B lymphocytes. The rise in Fc gamma RII surface expression occurred after the induction of class II antigens (Ia) and before transferrin receptor induction. Fc gamma RII expression was found to be enhanced during the G1 phase of the cell cycle since (a) only large cells (i.e. those that had entered the G1 phase) expressed an increased amount of Fc gamma RII and (b) blocking the entry of activated cells into the S phase (with the ion channel blocker quinine) did not affect the Fc gamma RII induction by LPS. Furthermore, only B cell activators that induced cells to enter into G1 [LPS and F(ab')2 anti-IgM antibodies, but not interleukin 4] caused an increase in the expression of Fc gamma RII. These results show that the increase in the membrane expression of Fc gamma RII occurs during the early G1 phase, establishing it as a marker for the entry of B lymphocytes into the cell cycle.
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PMID:Fc gamma RII expression in resting and activated B lymphocytes. 255 Feb 46

B cells can be activated by T-independent antigens or mitogens such as lipopolysaccharide (LPS) which will induce proliferation and differentiation of the B cells into Ig-secreting cells, without the intervention of T cells. The precise mechanism of T-independent proliferation and differentiation of B cells is still unclear. It is possible however that antigen-stimulated B cells may produce some factors which play a role in T-independent B-cell responses. In addition, since it has now been established that B cells can function as antigen-presenting cells, it is possible that they too secrete a molecule which is involved in the activation of T cells, analogous to IL-1 production by antigen-presenting macrophages. A number of human B-cell lines, as well as human normal B cells activated appropriately, have been shown to produce various cytokines, and similar studies are now being undertaken in the mouse. In the present study, six cloned murine B-cell lymphomas of different origin were analyzed for the presence of mRNA encoding a number of lymphokines by hybridization of specific cDNA probes to poly-A RNA, followed by the sensitive S1 nuclease digestion technique. The lymphokines included (IL-) 1, 2, 3, 4, 5, and neuroleukin. Whereas none of the lines expressed detectable levels of IL-2, IL-3, or IL-5 mRNA, all the lines expressed high levels of neuroleukin mRNA. Three of the lymphomas (CH12, CH31, and NBL) expressed low levels of IL-1 mRNA. The most striking finding was that one lymphoma, CH12, constitutively expressed IL-4 mRNA. This mRNA appeared to be functional, as IL-4 activity measured by the HT-2 T cell proliferation assay could be detected in supernatants collected from CH12 cells. The growth-inducing activity of CH12 supernatant on HT-2 cells could be completely blocked by an anti-IL-4 monoclonal (11B11), but not by an anti-IL-2 antibody (S4B6), consistent with our observations that CH12 cells produce IL-4 but not IL-2. CH12 cells were also found to express high affinity receptors for IL-4. Proliferation of CH12 cells was not affected by the addition of exogenous IL-4. Addition of anti-IL-4 antibodies to CH12 cells in culture caused a slight but reproducible increase in their proliferation at low cell numbers, which is probably not highly significant. These findings open the possibilities that murine B lymphocytes are capable of lymphokine production or alternatively that aberrant lymphokine production underlies B-lymphocyte transformation.
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PMID:Constitutive production of lymphokines by cloned murine B-cell lymphomas--CH12 B lymphoma produces interleukin-4. 278 29

We produced the T cell hybridoma D9C1.12.17 by fusing an IL-4-producing T cell clone D9.1Hi with the AKR thymoma BW5147. The resulting hybridoma produced IL-2 as well as IL-4 even though none of the parental cells produced IL-2 after stimulation with Con A. The production of IL-2 was confirmed at the mRNA level by using an S1 nuclease protection assay. Further analysis indicated that Con A-induced IL-2 production was a common phenomenon among T cell hybridomas derived from this fusion. Although BW5147 does not produce detectable lymphokines after Con A stimulation, this line was able to produce IL-2, granulocyte-macrophage colony stimulating factor, and small amounts of IL-3 and IFN-gamma when stimulated with calcium ionophore and phorbol ester. The latter agents are thought to mimic the activating signal(s) delivered through the Ag:MHC TCR. This observation indicates that BW5147 has the ability to produce lymphokines but may lack component(s) which couple the extracellular signal to lymphokine production, and suggests that in T cell hybridomas, part of the spectrum of lymphokines produced may be contributed by BW5147.
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PMID:The AKR thymoma BW5147 is able to produce lymphokines when stimulated with calcium ionophore and phorbol ester. 312 29

Keratinocyte derived T-cell growth factor was initially described as a product of cultured neonatal keratinocytes and keratinocyte cell lines that induced the proliferation of HT-2 cells, a murine T-cell line that responds to IL-2 and IL-4 by incorporating 3H-Thymidine. Subsequently, KTGF has been purified to high specific activity and found to be distinct from IL-2 and IL-4 by a variety of biochemical, immunologic, and immunochemical criteria. Because it was found that certain HT-2 cell lines also proliferated in response to GM-CSF, the present study asked whether KTGF was related to GM-CSF. In this study, we demonstrate that antibodies to recombinant murine GM-CSF completely neutralize the capacity of KTGF to induce HT-2 proliferation without interfering with IL-2 or IL-4 induced HT-2 proliferation. Furthermore, poly-A+ RNA homologous to murine GM-CSF cDNA as judged by S1 nuclease analysis was detected in Pam 212 cells, and protein serologically homologous to GM-CSF was found in Pam 212 conditioned medium. We conclude that KTGF is identical to GM-CSF. The T-cell activating properties of GM-CSF require further exploration.
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PMID:Keratinocyte derived T-cell growth factor (KTGF) is identical to granulocyte macrophage colony stimulating factor (GM-CSF). 329 4

The expression of cytokine genes in cultures of human peripheral blood mononuclear cells (PBMC) stimulated with mannoprotein constituents (MP) of Candida albicans has been studied by means of S1 nuclease mapping analysis, polymerase chain reaction, and enzyme-linked immunosorbent assay. MP induced early, consistent, and long-lasting production of interleukin-1 beta (IL-1 beta), tumor necrosis factor alpha, and IL-6 mRNAs. Similar results were obtained when the same PBMC cultures were stimulated with the purified protein derivative (PPD) from Mycobacterium tuberculosis or with IL-2, although lower levels of IL-6 mRNA were detected in IL-2-stimulated cells than in MP- or PPD-stimulated cells. MP, PPD, and IL-2 induced appreciable levels of granulocyte-macrophage colony-stimulating factor and gamma interferon, but only MP and PPD were able to induce IL-2 mRNA. MP were unable to stimulate a consistent expression of the genes encoding for IL-4, IL-5, and IL-10, while low, sometimes barely detectable levels of these cytokine mRNAs were observed in PPD- or IL-2-stimulated PBMC cultures. When protein synthesis of MP-stimulated PBMC was inhibited by cycloheximide, a superinduction of mRNAs for IL-4 and IL-10 and, more markedly, gamma interferon was observed. Overall, these results highlight the powerful, selective induction of cytokine gene expression by MP constituents of C. albicans in human PBMC cultures, thus providing some functional clues to explain the efficient state of the anticandidal response in normal human subjects.
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PMID:Cytokine gene expression in human peripheral blood mononuclear cells stimulated by mannoprotein constituents from Candida albicans. 840 99

Expression of the alpha(v)beta(3) integrin by murine bone marrow macrophages is regulated by cytokines such as IL-4 and GM-CSF through transcriptional activation of the beta(3) subunit gene. To characterize the molecular mechanisms by which such regulation occurs, we isolated the murine beta(3) integrin promoter. To this end, we first cloned a full length beta(3) cDNA and used the 5'UTR and leader peptide coding sequence to identify genomic clones containing the beta(3) promoter region. The transcriptional start site, identified by primer extension and S1 nuclease assay, is 34 nt upstream of the translation initiation codon. A 1.1 kb fragment of the promoter region drives IL-4 responsive transcription in transiently transfected murine bone marrow macrophages. Deletion analysis of the beta(3) promoter indicates the IL-4 responsive element lies between -465 to -678 nt relative to the transcriptional start site. This promoter fragment contains two overlapping STAT consensus recognition sites and nuclear extracts from BMMs contain an IL-4-inducible DNA binding factor, identified by super shift analysis, as STAT-6. Furthermore, an oligonucleotide which includes the two STAT recognition sites residing in the IL-4 responsive region of the beta(3) promoter, competes for STAT-6 binding. Confirming IL-4 induction of the integrin subunit is specifically mediated by STAT-6, beta(3) mRNA is not enhanced in BMMs derived from STAT-6 deleted mice, which however, retain their capacity to respond to GM-CSF.
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PMID:Cloning and characterization of the murine beta(3) integrin gene promoter: identification of an interleukin-4 responsive element and regulation by STAT-6. 1124 72

Diesel exhaust particles (DEP) are known to modulate the production of cytokines associated with acute and chronic respiratory symptoms and allergic respiratory disease. Tolerance is an important mechanism through which the immune system can maintain nonresponsiveness to common environmental antigens. We examined the effect of DEP on IL-10 and TGF-beta, cytokines produced by macrophages and repressor (Tr-like) lymphocytes which influence tolerance. Human PBMCs (n = 22) were incubated with 1-100 ng/ml of DEP, and suboptimally primed with LPS. IL-10 gene expression was assessed by the S1 nuclease protection assay, and production of IL-10, TGF-beta, TNF-alpha, IL-1 beta and IL-4 stimulated CD23 was evaluated by ELISA after 24 and 48 h. The effect of the order of exposure to DEP and LPS was evaluated on IL-10 protein and mRNA in cells (1) preincubated with LPS followed by DEP, or (2) exposed first to DEP followed by LPS. IL-10 was further evaluated using benzo[a]pyrene and [alpha]naphthoflavone as a surrogate for the polyaromatic hydrocarbons (PAHs) adsorbed to DEP. Control cells were incubated with carbon black, without PAHs. In PBMCs exposed to DEP with LPS, or preincubated with LPS before DEP, IL-10 production and mRNA fall significantly. TGF-beta is similarly suppressed, IL-1 beta secretion is significantly stimulated, and IL-4 stimulated CD23 release rises in the atopic subjects. In contrast, when DEP is added prior to LPS, IL-10 production rises, and IL-1 beta falls to zero. These effects on IL-10 are reproduced with benzo[a]pyrene and reversed by the coaddition of [alpha]naphthoflavone, its known antagonist. The carbon black fraction has no effect on IL-10 production. The effect of DEP on IL-10 can be inhibitory or stimulatory, depending on the order of exposure to DEP and LPS. Pro-inflammatory cytokines and factors rise when IL-10 is inhibited, and are suppressed when IL-10 is stimulated. These results are duplicated with benzo[a]pyrene, suggesting that the PAH portion of the DEP is the active agent.
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PMID:The influence of diesel exhaust particles on mononuclear phagocytic cell-derived cytokines: IL-10, TGF-beta and IL-1 beta. 1173 50