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:2.3.1.28 (chloramphenicol acetyltransferase)
5,100 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have analyzed in various human leukemic cell lines a previously unrecognized region within the human TNF gene promoter that contains the sequence motif 5'-CCGCCCCCGCG-3'. This GC-rich sequence maps to bps -170 and -160 of the TNF gene. Electrophoretic mobility shift assays (EMSA) combined with methylation interference analysis revealed the binding of two distinct proteins with overlapping recognition sites. Supershift assays identified the constitutive transcription factor Sp1 and the immediate-early growth-response transcription factor Egr-1/Krox-24. Interestingly, this Egr-1-related factor was induced by PMA but not by TNF. The TNF gene GC-rich sequence conferred PMA responsiveness when linked to a heterologous minimal c-fos promoter. To examine the involvement of Egr-1/Krox-24 in TNF gene regulation, a Krox-24 expression vector was used, pSCTKr24. In Jurkat T cells pSCTKr24 stimulated pTNF-286CAT that contains sequences -286 to +34 of the human TNF gene fused to the chloramphenicol acetyltransferase (CAT) gene. Moreover, pSCTKr24 also stimulated the TNF gene GC-rich sequence linked to the minimal c-fos promoter. However, deletion of this site did not result in markedly reduced TNF promoter activity, suggesting that the Egr-1/Krox-24 response element may play an auxiliary role in TNF gene regulation.
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PMID:Characterization of an Krox-24/Egr-1-responsive element in the human tumor necrosis factor promoter. 791 37

We have previously isolated a HeLa cell cDNA encoding a 21-kDa polypeptide that is 48% similar to transcription factor IIS. To explore the possibility that p21 plays a role in transcriptional regulation in vivo, we tested the effect of p21 expression on the synthesis of reporter chloramphenicol acetyltransferase (CAT) in transfected COS-1 cells. CAT formation under control of the Rous sarcoma virus long terminal repeat (RSV LTR) promoter was decreased nearly 20-fold in cells coexpressing p21. In contrast, CAT production under control of other sequence elements was only slightly reduced (human immunodeficiency virus type 1 LTR, simian virus 40 early promoter), unaffected (human heat shock protein of 70-kDa promoter, adenovirus major late promoter TATA box), or increased (terminal deoxynucleotidyltransferase initiator element, c-fos promoter) by p21 coexpression as compared to cells cotransfected with the parental vector. The abundance of steady-state CAT transcripts from RSV LTR was also decreased by p21 expression in a dose-dependent manner, suggesting that transcription of RSV LTR/CAT is under negative control by p21. Consistent with an effect on transcription, p21 was localized in nuclei of transfected cells. Deletion analysis of p21 indicated that the sequences essential for inhibition of RSV LTR function include the previously identified ARg/Ser-rich region and zinc finger-like motif. Proliferation of chicken embryo fibroblasts transfected with an infectious molecular clone of RSV was diminished by p21 expression, which also resulted in fewer transformed foci.
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PMID:Down-regulation of Rous sarcoma virus long terminal repeat promoter activity by a HeLa cell basic protein. 797 97

Functional antagonism between retinoic acid (RA) receptors and activator protein-1 (AP-1) transcription factors might regulate expression of genes involved in the response to injury in the kidney. We designed experiments to analyze the mechanisms by which RA inhibits AP-1-directed transcriptional responses in glomerular mesangial cells. RA inhibited serum-stimulated mesangial cell proliferation as assessed by measurements of [3H]thymidine uptake and cell number. In transient transfection assays with a chloramphenicol acetyltransferase reporter, RA completely blocked transcription directed by an AP-1 cis-element in cells stimulated by serum. AP-1 DNA binding was analyzed in electrophoretic gel mobility shift assays using nuclear extracts from control or RA-pretreated cells stimulated with serum. RA did not abolish AP-1 DNA binding activity under the conditions of this assay. The apparent equilibrium dissociation constant, maximal density of binding, and association rate for the AP-1-DNA interaction were similar in serum-stimulated cells or RA-pretreated cells stimulated with serum. RA repressed serum-stimulated induction of the immediate early genes c-fos and c-jun, whose protein products dimerize to form AP-1. Repression was relatively selective for c-fos/c-jun; induction of other immediate early transcription factors (junB, c-myc, and egr-1) was not downregulated by RA. That repression of c-fos by RA might contribute to anti-AP-1 activity was suggested by experiments with an antisense c-fos expression vector, which demonstrated that c-fos induction was required for serum-stimulated AP-1 activity. Together, these data demonstrate that RA antagonizes AP-1-directed transcription without inhibiting AP-1 DNA-binding in mesangial cells. Selective repression of c-fos and c-jun might contribute to the anti-AP-1 activity of RA.
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PMID:Anti-AP-1 activity of all-trans retinoic acid in glomerular mesangial cells. 797 84

Transforming growth factor beta (TGF-beta) is a potent modulator of cell growth in many systems. In normal rat kidney fibroblasts, TGF-beta 1 increases epidermal growth factor (EGF) receptor gene transcription and synergizes with EGF to stimulate growth in soft agar, a characteristic of the transformed phenotype. In order to identify the target of TGF-beta 1 action, we have used a series of 5' deletion mutants of the EGF receptor promoter linked to a chloramphenicol acetyltransferase reporter gene (ERCAT). The TGF-beta response element(s) was localized to a cis-regulatory region which resides between positions -919 and -860 relative to the ATG translation initiation codon of the EGF receptor promoter. This 60-base pair region contains a repressor of the EGF receptor promoter and a TGF-beta inhibitory element that mediates TGF-beta 1 suppression of transin/stromelysin gene transcription through binding of a Fos-containing protein complex. Cotransfection of c-fos, c-jun, or both expression vectors with the intact or 5'-deleted ERCAT constructs identified several Fos-responsive inhibitory regions within the EGF receptor promoter, but these did not localize to the -919 to -860 promoter region. Mobility shift assays showed binding of the 60-base pair DNA fragment to proteins in extracts from untreated normal rat kidney cells; the binding was specifically competed by oligonucleotides containing a CAGATG sequence but not by oligonucleotides containing the EGF receptor repressor or the TGF-beta inhibitory element. TGF-beta 1 treatment but not anti-Fos antibody caused a decrease in specific 60-base pair DNA-protein complex formation.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Induction of epidermal growth factor receptor gene transcription by transforming growth factor beta 1: association with loss of protein binding to a negative regulatory element. 798 46

Receptor-bound growth factors elicit intracellular signals that lead to the phosphorylation and activation of numerous intracellular kinases and transcription factors with consequent changes in patterns of gene expression. Several oncogene products are able to mimic these signals, resulting in cell transformation and proliferation. For example, the introduction of oncogenic forms of Raf-1 kinase into fibroblasts induces transformation and leads to the constitutive expression of, among others, the c-fos proto-oncogene. Here it is shown that the elevation of c-fos promoter activity brought about by v-raf is mediated by TCF/Elk-1, which forms a ternary complex with SRF at the serum response element and is a substrate for mitogen-activating protein kinases in vitro. In NIH 3T3 fibroblasts, v-raf activates Erk2, and overexpression of an interfering mutant of Erk2 both blocks the ability of v-raf to activate the c-fos promoter and suppresses transformation. Mutation of individual mitogen-activating protein kinase phosphoacceptor sites in TCF/Elk-1 also compromises v-raf-activated expression of a Gal-Elk/Gal-chloramphenicol acetyltransferase reporter system. However, in at least one instance the introduction of glutamate, but not aspartate, at a phosphoacceptor site is compatible with activation. These results provide compelling evidence that phosphorylation of TCF/Elk-1 by Erk2 is a major link in the Raf-1 kinase-dependent signal transduction pathway that activates c-fos expression.
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PMID:Inhibition of v-raf-dependent c-fos expression and transformation by a kinase-defective mutant of the mitogen-activated protein kinase Erk2. 800 80

Prolonged expression of activated ras mutants resulted in both neoplastic transformation and suppression of serum-induced c-fos expression in Rat1 fibroblasts. Expression of other serum-inducible genes, including c-jun and beta-actin, was not suppressed in ras-transformed Rat1 cells, indicating that these effects are specific for c-fos and that growth-factor signal transduction pathways remain essentially intact. Run-on transcription studies indicated that c-fos transcription was blocked at the level of initiation in these cells. Transient transfection studies using 360 bp from the wild-type c-fos promoter as well as a series of mutated c-fos promoter fragments linked to the chloramphenicol acetyltransferase gene indicated that repression of c-fos was mediated by approximately 49 bp immediately upstream of the dyad symmetry element (DSE). Deletion of this region, referred to as the upstream repressor region (URR), restored serum inducibility to the c-fos promoter in ras-transformed cells. In contrast, suppression of c-fos transcription was not affected by either deletion of 240 bp between the DSE and the TATA element or by base-substitution mutations that inactive the ternary complex factor and fos-AP-1-like binding sites. In addition, in vitro competition studies indicated that ras-transformed cells express one or more repressor factors that interact with as-yet-unidentified elements within the c-fos promoter (possibly the URR) and block serum induction of c-fos. These findings suggest that prolonged expression of activated ras results in the activation of one or more as-yet-unidentified proteins that suppress transcription of the c-fos gene by interacting with the URR.
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PMID:Mediation of suppression of c-fos transcription in rasT24-transformed rat cells by a cis-acting repressor element. 803 67

Matrix metalloproteinases are secreted enzymes important in inflammation and tumor invasion. Earlier, we demonstrated that in normal human FS-4 fibroblasts, collagenase and stromelysin mRNA levels are increased not only after treatment with known matrix metalloproteinase inducers such as tumor necrosis factor (TNF), interleukin-1, and 12-O-tetradecanoylphorbol-13-acetate, but also with interferon-beta (IFN-beta). In this study, we compared the regulation of these matrix metalloproteinase genes by TNF and IFN-beta. We show that both TNF and IFN-beta increase steady-state levels of collagenase and stromelysin mRNAs with similar slow kinetics. The glucocorticoid dexamethasone blocked matrix metalloproteinase induction by both cytokines. The protein synthesis inhibitor cycloheximide inhibited collagenase mRNA induction by TNF or IFN-beta, suggesting that induction by both agents is indirect. Consistent with these observations, both TNF and IFN-beta increased c-fos and c-jun mRNA levels. Furthermore, treatment with TNF or IFN-beta increased the transcriptional activity of activator protein-1-responsive chloramphenicol acetyltransferase reporter gene constructs, including a native collagenase promoter-driven chloramphenicol acetyltransferase construct. These findings show that regulation of matrix metalloproteinase gene expression by both TNF and IFN-beta involves the transcription factor activator protein-1 and demonstrate a novel indirect mechanism of type I IFN-induced gene expression.
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PMID:Interferon-beta induces metalloproteinase mRNA expression in human fibroblasts. Role of activator protein-1. 806 4

Photodynamic therapy (PDT) generates reactive oxygen species which initiate the cytotoxic events of this tumor treatment. We demonstrate that PDT mediated oxidative stress induced a transient increase in the early response genes c-fos, c-jun, c-myc, and egr-1 in murine radiation-induced fibrosarcoma cells. Incubation of exponentially growing cells with porphyrin based photosensitizers in the dark also induced an increase in mRNA levels of early response genes. However, the xanthine photosensitizer, rose bengal, produced increased c-fos mRNA levels only following light treatment. Nuclear runoff experiments confirmed that the induction of c-fos mRNA is controlled in part at the level of transcription. Likewise, a chloramphenicol acetyltransferase reporter construct containing the major c-fos transcriptional response elements was inducible by porphyrin and PDT. Signal transduction pathways associated with PDT mediated c-fos activation were examined by treating cells with protein kinase inhibitors. Staurosporine and 1-(5-isoquinolinesulfonyl)-2-methylpiperazine inhibited PDT mediated c-fos activation while N-(2-guanidinoethyl)-5-isoquinoline-sulfonamide had no effect. In addition, quinacrine, which can inhibit phospholipase activity, blocked PDT induced c-fos mRNA expression. These results suggest that photosensitizer mediated oxidative stress acts through protein kinase-mediated signal transduction pathway(s) to activate early response genes.
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PMID:Photodynamic therapy mediated induction of early response genes. 811 27

The mouse epidermal cell line 308 contains an activated Ha-ras gene and forms benign papillomas when transplanted to the skin of athymic nude mice. A radiation-associated malignant variant of this cell line, 308-10Gy5, has been isolated and shown to form squamous cell carcinomas in nude mice. To further examine the molecular events involved in malignant conversion of 308-10Gy5, we assessed the activator protein-1 (AP-1) binding and transactivating ability of 308 and 308-10Gy5. In nuclear protein extracts of 308, AP-1 sequence-specific binding to an oligonucleotide containing a single high-affinity AP-1 binding site was induced by the tumor promoter 12-O-tetradecanoylphorbol-13-acetate, as determined by gel shift analysis. Nuclear extracts of 308-10Gy5 bound to the AP-1 oligonucleotide without treatment with tumor promoters. Not only was sequence-specific AP-1 DNA binding constitutively active in malignant versus benign tumor cells, but so was transactivation of a unique AP-1-responsive chloramphenicol acetyltransferase reporter construct, pTiCTaK. Constitutive transactivation of this AP-1-responsive reporter construct was observed in the malignant but not the benign tumor cells. Furthermore, steady-state transcript levels of the tumor-associated AP-1-responsive genes stromelysin, urokinase-type plasminogen activator, c-jun, and c-fos were higher in malignant 308-10Gy5 cells than in benign 308 cells. These results suggest that acquisition of constitutive AP-1 DNA binding and transactivation can result in sustained deregulation of gene expression. While malignant progression in keratinocytes is probably not due solely to the acquisition of constitutive cellular AP-1 activity, the effect of deregulated expression of AP-1-regulated genes, especially basement membrane-degrading enzymes, may be functionally related to malignant conversion.
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PMID:Constitutive AP-1 DNA binding and transactivating ability of malignant but not benign mouse epidermal cells. 814 9

Transcription of the cytoskeletal beta-actin gene is rapidly induced by phorbol 12-myristate 13-acetate (PMA) and the calcium ionophore, A23187, in cultured H4IIE hepatoma (H4) cells. PMA directly activates protein kinase C (PKC) and activation of PKC is necessary for the cellular actions of PMA, including induction of beta-actin gene transcription. In the present study, we determined the DNA sequence requirements for induction of the beta-actin gene by PMA and A23187. Constructs containing progressive deletions of normal and mutated human beta-actin 5' sequences fused to the reporter gene, bacterial chloramphenicol acetyltransferase, were analysed in transient transfections of H4 cells. We delineated the PMA response DNA element of the human beta-actin gene to the proximal CCArGG box (-62 to -53) in the 5' flanking region. In contrast, A23187 did not induce expression of transfected gene constructs containing this CCArGG box. Additionally, we demonstrated that CCArGG boxes from two other PMA-induced genes in H4 cells, c-fos and gamma-actin, could confer PMA inducibility to a heterologous promoter. This CCArGG box specifically interacts with one or more proteins present in nuclear extracts of H4 cells. These results indicate that in cultured cells, PMA-dependent induction of the beta-actin gene is mediated through the proximal CCArGG box. This suggests that the CCArGG box is a target for PKC action and may be involved in the control of other PKC regulated genes.
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PMID:Identification of beta-actin sequences necessary for induction by phorbol esters and calcium ionophores. 818 67


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