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Query: UNIPROT:P04141 (granulocyte-macrophage colony-stimulating factor)
 6,790 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Selective mRNA degradation is an important control point in the transient expression of a variety of mRNAs coding for growth regulators. A variety of labile mRNAs coding for lymphokines, cytokines, and oncogenes contain within their 3'-untranslated region an AU-rich region shown to destabilize these messages. We recently identified a cytosolic protein, adenosine-uridine binding factor (AUBF), which complexes with four tandem AUUUA reiterations of a synthetic RNA transcript. We now show that AUBF forms RNase T1-resistant band-shifted complexes with a variety of in vitro transcribed mRNAs including granulocyte-macrophage colony-stimulating factor, interferon-gamma, interleukin-3, c-fos, and v-myc. Formation of complexes was specifically inhibited by AUUUA containing RNA, but not by irrelevant RNA. After brief ultraviolet light-induced cross-linking, AUBF.RNA complexes with the exception of c-fos comigrated on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Mutations within the AUUUA motifs demonstrate that both nucleotide sequence and secondary structure are important in AUBF.AUUUA RNA complex formation. Based upon these data, we suggest AUBF may interact with a variety of labile mRNAs with multiple AUUUA reiterations or single reiterations within an AU-rich 3'-untranslated region.
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PMID:The adenosine-uridine binding factor recognizes the AU-rich elements of cytokine, lymphokine, and oncogene mRNAs. 199 89

Post-transcriptional gene regulation plays an important role in the expression of granulocyte-macrophage colony-stimulating factor (GM-CSF). Cytokine secretion by activated lymphocytes or mast cells is preceded by dramatic stabilization of the normally labile GM-CSF mRNA. The 3'-untranslated region of GM-CSF and other labile mRNAs contain the destabilizing motif adenosine-uridine-uridine-uridine-adenosine (AUUUA). We recently identified a cytoplasmic protein denoted the adenosine-uridine binding factor (AUBF) which binds with high affinity and specificity to AUUUA elements in synthetic RNA transcripts. We now demonstrate that AUBF binds specifically to GM-CSF mRNA through the destabilizing AUUUA elements. The formation of AUBF-GM-CSF RNA complexes required calcium or magnesium which were sensitive to EDTA or EGTA. A variety of other divalent metals blocked magnesium-dependent AUBF activity. These observations suggest that AUBF may protect GM-CSF mRNA from rapid degradation and play a crucial role in the expression of cytokine genes.
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PMID:Adenosine-uridine binding factor requires metals for binding to granulocyte-macrophage colony-stimulating factor mRNA. 213 52

The granulocyte-macrophage CSF (GM-CSF) gene is known to be controlled at a variety of levels in different cell types. We showed previously that GM-CSF production by lectin or phorbol ester (12-O-tetradecanoyl-phorbol-13-acetate (TPA]-treated T cells was unaffected by cyclosporin A whereas IL-2 and IL-3 expression were. Cyclosporin A is thought to inhibit transcription that suggests that IL-2 and IL-3 are regulated primarily at the transcriptional level while GM-CSF is not. The lack of coordinate gene expression is of particular interest because all three mRNA share the presence of adenosine uridine-rich sequences in the 3' untranslated region and these sequences are believed to act by modulating mRNA stability. We measured the level of GM-CSF mRNA in untreated cells and found it to be extremely low. GM-CSF mRNA levels increased approximately 60-fold within 6 h of TPA-treatment. Nuclear run-on transcription analysis of the same cells showed readily detectable GM-CSF transcription in unstimulated cells that increased less than twofold after TPA treatment. However, IL-2 transcription was insignificant before TPA addition. Actinomycin D chase experiments showed that GM-CSF transcripts in untreated cells have a very short half-life (approximately 45 min) although transcripts in TPA-treated cells have a half-life exceeding 3 h. These findings indicate that GM-CSF production in EL-4 cells treated with TPA is regulated predominantly by modulation of cytoplasmic mRNA half-life.
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PMID:Post-transcriptional regulation of granulocyte-macrophage colony-stimulating factor synthesis in murine T cells. 219 29

We have previously demonstrated that protein production and mRNA expression of granulocyte-macrophage colony-stimulating factor (GM-CSF), granulocyte colony-stimulating factor (G-CSF), and IL-3 are decreased in activated mononuclear cells (MNC) from human umbilical cord compared with adult peripheral blood. Reduced production of these colony-stimulating factors (CSF) during states of increased demand, as occurs during overwhelming bacterial infection, may play a role in the pathogenesis of neutropenia and thrombocytopenia in the newborn. To determine whether the reduced mRNA expression and CSF production from activated cord MNC is secondary to the decreased transcriptional activity of the corresponding genes, we determined the transcriptional rate of GM-CSF, G-CSF, IL-3, and M-CSF by nuclear run-on assays. Cord and adult MNC were isolated by Ficoll-Hypaque density centrifugation. A total of 10(8) MNC from cord and adult blood were stimulated as follows: GM-CSF and G-CSF [32 nmol/L phorbol-12-myristate-6-acetate (20 micrograms/L) + 2 mg/L phytohemagglutinin for 6 h]; IL-3 [32 nmol/L phorbol-12-myristate-6-acetate (20 micrograms/L) + 0.5 mumol/L A 23187 for 6 h]; and macrophage CSF (2 micrograms/L recombinant human GM-CSF for 24 h). The nuclei from unstimulated and stimulated cells were isolated and labeled with 32P-uridine triphosphate. Newly elongated 32P-labeled RNA transcripts were hybridized to slot blots of CSF DNA. To minimize cross hybridization artifacts, short fragments (0.5-1.0 kb) of cDNA were used.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Transcriptional rates of granulocyte-macrophage colony-stimulating factor, granulocyte colony-stimulating factor, interleukin-3, and macrophage colony-stimulating factor genes in activated cord versus adult mononuclear cells: alteration in cytokine expression may be secondary to posttranscriptional instability. 750 24

The adenosine-uridine (AU)-rich sequences within the 3' untranslated region (UTR) of many short-lived mRNAs are important in their rapid degradation. We present evidence that human embryonic lung fibroblasts (W138) contain five major proteins of 70, 45, 40, 38, 32.5 kd, which specifically bind the AU-rich region of human granulocyte-macrophage colony-stimulating factor (GM-CSF) 3'UTR containing 7 x AUUUA motifs. The 40 and 38 kd proteins also bound the 3x and 5 x AUUUA cassettes and even more strongly bound to the AUUUUUUUA motif. All five of these proteins showed more abundant localization in the nucleus than the cytoplasm. The 32.5 kd protein was the major cytoplasmic AU-binding protein. Incubation with actinomycin D resulted in a marked increase in binding activity of 45, 40, 38, and 32.5 kd proteins in the cytoplasm, accompanied by decreased binding activity of the 32.5 kd protein in the nucleus. Antibody against heterogeneous nuclear ribonucleoprotein C (hnRNP C) immunoprecipitated the 40 and 38 kd proteins, and antibody against the AU-rich element RNA-binding protein (AUF1) immunoprecipitated the 45, 40, and 38 kd proteins. The present results not only demonstrated that hnRNP C are AU-binding proteins which are present in the cytoplasm as well as the nucleus, but another group of AU-binding proteins (AUF1 [45, 40, 38 kd], and 32.5 kd), which are not hnRNP, have characteristics related to those of hnRNPs. Taken together with our previous results (Akashi et al., 1994, Blood, 83:3182-3187), AU-binding factors including hnRNP C and AUF1, which bind more than 3 x AUUUA motifs, may be involved in rapid degradation of these transcripts. No significant quantitative changes of these proteins in their binding activity to AU-rich sequences occurred in response to several stimuli that stabilize GM-CSF mRNA, indicating that binding of these proteins to their cognate RNA is not responsible for the stabilization of these transcripts.
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PMID:Characterization of adenosine-uridine-rich RNA binding factors. 759 27

Recently, the interleukin-2 receptor (IL-2R) was shown to be present on human neutrophils, and IL-2-neutrophil interactions are believed to be important in both tumor rejection and increased susceptibility to bacterial infections. Furthermore, neutrophils have been shown to synthesize host defense proteins, such as cytokines. In this study, we analyzed the effects of IL-2 on the induction of de novo RNA and protein synthesis in this cell type. When cells were stimulated with IL-2 alone, the level of incorporation of either [5-3H]-uridine or [35S]-methionine and [35S]-cysteine was similar to unstimulated cells. However, when cells were stimulated with the combination of a fixed concentration of granulocyte-macrophage colony-stimulating factor (GM-CSF), a dose-dependent effect of IL-2 was observed on the induction of both RNA and protein synthesis. In the presence of tumor necrosis factor-alpha or formyl-methionyl-leucyl-phenylalanine, however, IL-2 exerted no similar effect. Furthermore, the study of a large number of normal subjects (n = 55) showed reproducible categories of responders (low, intermediate, and high). The binding of IL-2 to the IL-2R complex on human neutrophils increased on GM-CSF-stimulated neutrophils compared with unstimulated cells. However, no increase in the level of expression of either the alpha or beta chains of this receptor complex was observed. This finding suggests that GM-CSF functionally activates the IL-2R, but does not regulate its level of expression. Finally, we found that human neutrophils constitutively express IL-2R gamma chain mRNA and thus have the potential to express the functional IL-2R complex. Our findings on IL-2-neutrophil interactions should lead to new avenues of research in understanding the responses of patients undergoing GM-CSF or IL-2 therapy.
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PMID:Effects of interleukin-2 on gene expression in human neutrophils. 762 Jan 70

In vitro decay of granulocyte-macrophage colony-stimulating factor (GM-CSF) mRNA was examined on polysomes prepared from normal human peripheral blood mononuclear cells stimulated with phorbol ester (TPA) and phytohemagglutinin for 14 h. GM-CSF mRNA decayed with a half-life of 90 min while 18 S rRNA was stable. RNA gel mobility assay performed on crude cytosolic lysate (S20) with radiolabeled AUUUA containing RNA identified a 42-kDa RNA-protein complex on SDS-polyacrylamide gel electrophoresis. The binding specificity was identical to that of the previously described adenosine-uridine binding factor (AUBF) (Malter, J. S. (1989) Science 246, 664-666). Further fractionation of the S20 cytosol through a sucrose gradient showed > 90% of AUBF activity associated with polysomes and < 10% with the S130 fraction. Solution phase RNAs containing AUUUA reiterations specifically competed for polysome-bound AUBF and accelerated the decay of GM-CSF mRNA (t1/2 = 17 min). We linked biotinylated AUUUA RNA to streptavidin magnetic beads and removed > 95% of polysome-associated AUBF. A decay system thus depleted of AUBF activity also showed accelerated decay of GM-CSF mRNA (t1/2 = 20 min). These data show that AUBF is preferentially located on polysomes and that its removal destabilizes GM-CSF mRNA. Therefore, AUBF likely prevents GM-CSF mRNA decay by binding to the AUUUA instability determinants in the 3'-untranslated region.
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PMID:Modulation of granulocyte-macrophage colony-stimulating factor mRNA stability in vitro by the adenosine-uridine binding factor. 792 35

The level of granulocyte-macrophage colony-stimulating factor (GM-CSF) mRNA is fourfold lower in phorbol myristate acetate (PMA) + phytohemagglutinin (PHA)-activated mononuclear cells (MNC) from newborns compared with adults. The GM-CSF transcription rate is similar in umbilical cord and adult MNC, but transcript half-life is threefold lower in cord activated MNC. Interaction of RNA binding proteins, such as the cloned adenosine + uridine-rich element, binding factor, AUF1, with eight AUUUA motifs in the human GM-CSF mRNA 3'-untranslated region (GM-3'-UTR) has been implicated in regulating transcript stability. Translational inhibition by cycloheximide (CHX) significantly increased GM-CSF mRNA accumulation and half-life by three-fold in activated cord MNC, but had a minimal effect in activated adult MNC as compared with PMA + PHA alone. Electrophoretic mobility-shift assays with a 32P-labeled, 305-nucleotide RNA comprising the GM-3'-UTR revealed two RNaseT1-resistant, bound complexes that were almost twice as abundant in cord than in adult MNC extracts. Mobility-shift competition assays and RNaseT1 mapping localized the binding site of both complexes to a 52-nucleotide region containing seven of eight AUUUA motifs. Inclusion of AUF1 antiserum produced a supershifted complex at 35-fold higher levels in cord than in adult MNC extracts. Extracts from the carcinoma cell line 5637, with extended GM-CSF mRNA half-life, also had very low levels of anti-AUF1 supershifted complex. Anti-AUF1 immunoblotting showed significantly higher levels of two AUF1 protein isoforms and lower levels of one in cord than in adult MNC or 5637 extracts. These results suggest that destabilization of GM-CSF mRNA in cord MNC is translation-dependent and that increased levels of specific AUF1 isoforms in cord MNC may target transcripts for increased degradation, which could account in part for dysregulation of neonatal phagocytic immunity.
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PMID:Increased granulocyte-macrophage colony-stimulating factor mRNA instability in cord versus adult mononuclear cells is translation-dependent and associated with increased levels of A + U-rich element binding factor. 887 85

The 3'-untranslated region of granulocyte/macrophage colony-stimulating factor (GM-CSF) mRNA contributes to the post-transcriptional regulation of gene expression. Degradation is partly mediated by adenosine- uridine-rich sequence elements (ARE), which serve as binding sites for specific proteins. Stabilization of RNA by phytohemagglutinin and concanavalin A treatment is dependent on regulatory sequence elements upstream of ARE. We have performed northwestern blot and filter binding assays using cell extracts and RNA sequences containing or lacking ARE. Murine and human T cell extracts (EL-4 and Jurkat) yielded two specific proteins of 93 and 94 kDa, respectively, that were binding to sequences upstream of ARE. Within this region, the human and murine RNA do not share any obvious sequence identity, yet both are target sites for the binding proteins. The smallest RNA fragments protected by the proteins from RNase A digestion, were 44 in the murine, and 38 ribonucleotides long in the human sequence. The binding activity of the 94 kDa protein derived from human Jurkat cells could be enhanced by phytohemagglutinin. The interaction with regulatory mRNA sequences and the responsiveness to phytohemagglutinin suggests that the proteins are involved in controlling GM-CSF mRNA turnover.
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PMID:Identification of proteins binding specifically to the 3'-untranslated region of granulocyte/macrophage-colony stimulating factor mRNA. 917 Oct 94

Human eosinophils activated by calcium ionophore produce granulocyte-macrophage colony-stimulating factor (GM-CSF). In T lymphocytes GM-CSF messenger RNA (mRNA) stability is regulated by 3' untranslated region (UTR) adenosine-uridine-rich elements (AREs). We show endogenous GM-CSF mRNA is rapidly induced in an eosinophil cell-line (AML14.3D10) after activation with ionomycin. To calculate the decay rate of GM-CSF mRNA in activated cells, eosinophils were transfected with wild-type, full-length GM-CSF mRNA or a mutant version lacking the AUUUA motifs. In unstimulated cells, wild-type GM-CSF mRNA decayed with a half-life time (t1/2) of 6+/-2 min while the mutant decayed with a t1/2 of 20+/-4 min, demonstrating the dominant, destabilizing effect of multiple AUUUA motifs. Within 1 hr of activation by ionomycin, the half-life of transfected wild-type mRNA increased by 2.5-fold, which increased up to 4-fold after 2 hr of activation. The half-life of the mutant GM-CSF was unaffected by ionomycin, demonstrating that ionophore-mediated stabilization requires intact AUUUA motifs. Actinomycin D (ActD) stabilized wild-type GM-CSF mRNA as well, causing poly(A) tail elongation and translation inhibition. These data show that in eosinophil-like cell lines, GM-CSF mRNA is exquisitely unstable but can be markedly stabilized by calcium ionophore. Both effects require intact 3' UTR AREs.
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PMID:Stabilization of granulocyte-macrophage colony-stimulating factor RNA in a human eosinophil-like cell line requires the AUUUA motifs. 982 39


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