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)

To study the mechanism of a novel herpes simplex virus (HSV) activity that stimulates expression of reporter genes containing beta interferon (IFN-beta)-coding sequences, we have established permanent DNA-transfected cell lines that each contain two distinct hybrid genes encoding mRNA species with different half-lives. These reporter genes comprised either the human IFN-beta- or bacterial chloramphenicol acetyltransferase (CAT)-coding and 3' untranslated regions placed under the transcriptional control of the powerful major immediate-early promoter-enhancer region (IE94) from simian cytomegalovirus. Most of the dual-transfected cell lines yielded significant levels of steady-state IE94-CAT mRNA and abundant constitutive synthesis of CAT enzyme activity, whereas no accumulation of IE94-IFN mRNA could be detected. However, infection with HSV type 1 resulted in a 300-fold increase in IE94-IFN-specific mRNA transcripts, compared with no more than 3- to 5-fold stimulation of IE94-CAT-specific mRNA. In contrast, cycloheximide treatment increased stable mRNA levels and transcription initiation rates from both the IE94-IFN and IE94-CAT hybrid genes. Run-on transcription assays in isolated nuclei suggested that induction of IE94-IFN gene expression by HSV type 1 occurred predominantly at the posttranscriptional level. Enhancement of the unstable IFN mRNA species after HSV infection was also observed in cell lines containing a simian virus 40 enhancer-driven IFN gene (SV2-IFN). Similarly, in transient-transfection assays, both SV2-IFN and IE94-IFN gave only low basal mRNA synthesis, but superinfection with HSV again led to high-level accumulation of IFN mRNA. Finally, substitution of the SV2-IFN gene 3' region with poly(A) and splicing signals from the SV2-CAT gene cassette led to stabilization of the IFN mRNA even in the absence of HSV. Therefore, we conclude that HSV infection leads to selective accumulation of IFN-beta mRNA by a posttranscriptional mechanism that is reporter gene specific and promoter independent.
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PMID:Herpes simplex virus infection selectively stimulates accumulation of beta interferon reporter gene mRNA by a posttranscriptional mechanism. 131 84

The expression of the human beta interferon (IFN-beta) gene was activated by the adenovirus E1B-19K protein. The sequence within the IFN-beta promoter which is related to the activation was analyzed by chloramphenicol acetyltransferase (CAT) assay. The repeated hexamer units, present within the region between -109 and -65 relative to the cap site, were required for the activation of the IFN-beta gene by the E1B-19K protein. The (AAGTGA)8 region, as a typical hexamer of the consensus sequences, was tested for function in the activation by the E1B-19K protein. When the hexamer (AAGTGA)4-8 was inserted upstream of several reporter genes (such as p55cat, pdlE1A-CAT, and pE1B-CAT) which were inefficiently stimulated, the CAT activities of these fusion genes were efficiently stimulated by the E1B-19K protein. These results show that the tandemly repeated hexamer sequences within the IFN-beta promoter can function as an inducible regulatory element in the activation by the adenovirus E1B 19K protein.
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PMID:Tandemly repeated hexamer sequences within the beta interferon promoter can function as an inducible regulatory element in activation by the adenovirus E1B 19-kilodalton protein. 213 95

Treatment of Daudi or HeLa cells with human interferon (IFN) alpha 8 before induction with either poly(I)-poly(C) or Sendai virus resulted in an 8- to 100-fold increase in IFN production. The extent of priming in Daudi cells paralleled the increase in the intracellular content of IFN-beta mRNA. IFN-alpha mRNA remained undetectable in poly(I)-poly(C)-treated Daudi cells either before or after priming. An IFN-resistant clone of Daudi cells was found to produce 4- to 20-fold more IFN after priming, indicating that priming was unrelated to the phenotype of IFN sensitivity. IFN treatment of either Daudi or HeLa cells transfected with the human IFN-beta promoter (-282 to -37) linked to the chloramphenicol acetyltransferase (CAT) gene resulted in an increase in CAT activity after induction with poly(I)-poly(C) or Sendai virus. A synthetic double-stranded oligonucleotide corresponding to an authentic 30-base-pair (bp) region of the human IFN-beta promoter between positions -91 and -62 was found to confer virus inducibility upon the reporter CAT gene in HeLa cells. IFN treatment of HeLa cells transfected with this 30-bp region of the IFN-beta promoter in either the correct or reversed orientation also increased CAT activity upon subsequent induction. IFN treatment alone had no detectable effect on the activity of either the 30-bp region or the complete human IFN promoter.
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PMID:Priming affects the activity of a specific region of the promoter of the human beta interferon gene. 215 28

The synthesis of fibrinogen in the liver is drastically enhanced during the inflammatory process. Two factors are involved: glucocorticoids and the hepatocyte stimulating factor which is identical with interleukin 6 (Il6), also called interferon beta 2. The function of the 5'-flanking region of the human beta-fibrinogen (beta-Fg) gene has been studied by deletion analysis with the chloramphenicol acetyltransferase (CAT) gene as a transient expression vector. In this analysis, a fragment containing 150 base pairs (bp) upstream from the cap site is sufficient to drive expression of the CAT gene in the hepatoma cells HepG2, but not in HeLa cells. The beta-Fg gene is induced by dexamethasone and Il6 in HepG2. We identify a domain located between -2900 and -1500 bp upstream from the transcription start point involved in dexamethasone sensibility. This distal regulatory region can confer hormone inductibility to a heterologous promoter and exert its effect in either orientation. The sequence located between -150 and -82 bp upstream from the transcription start point is responsive for the Il6-stimulated expression. This 68-bp sequence contains probably all the cis-acting Il6-responsive element of the human beta-Fg gene.
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PMID:Human beta-fibrinogen gene expression. Upstream sequences involved in its tissue specific expression and its dexamethasone and interleukin 6 stimulation. 231 33

The relationship between transcription of alpha and beta interferon (IFN-alpha and IFN-beta) genes and the interaction of IFN promoter-binding transcription factors has been examined in monoblastoid U937 cells following priming with recombinant IFN-alpha 2 (rIFN-alpha 2) and Sendai virus induction. Pretreatment of U937 cells with rIFN-alpha 2 prior to Sendai virus infection increased the mRNA levels of IFN-alpha 1, IFN-alpha 2, and IFN-beta as well as the final yield of biologically active IFN. Analysis of nuclear protein-IFN promoter DNA interactions by electrophoretic mobility-shift assays demonstrated increased factor binding to IFN-alpha 1 and IFN-beta regulatory domains, although no new induction-specific complexes were identified. On the basis of competition electrophoretic mobility-shift assay results, factors interacting with the IFN-alpha 1 and IFN-beta promoters appear to be distinct DNA-binding proteins. U937 factor binding was localized to the P2 domain (-64 to -55) of the IFN-beta regulatory element, a sequence motif with 80% homology to the recognition site of transcription factor NF-kappa B. Protein-DNA interactions within the IFN-beta P2 domain were, in fact, specifically competed by either excess homologous P2 fragment or the human immunodeficiency virus enhancer element which contains two duplicated NF-kappa B recognition sites. Hybrid promoter-chloramphenicol acetyltransferase fusion plasmids, containing either the IFN-beta regulatory element or the human immunodeficiency virus enhancer element linked to the simian virus 40 promoter, were analyzed for virus and phorbol ester inducibility in epithelial and lymphoid cells, respectively. In the 293 cell line, both plasmids were constitutively expressed but not virus inducible, while in Jurkat cells, chloramphenicol acetyltransferase activity from these plasmids was induced by tumor-promoting agent treatment. These experiments suggest that induction of IFN gene expression may be controlled in part by transcription regulatory proteins binding to an NF-kappa B-like site within the IFN-beta promoter.
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PMID:Induction of human interferon gene expression is associated with a nuclear factor that interacts with the NF-kappa B site of the human immunodeficiency virus enhancer. 254 71

Virus inducible elements (IE) in promoters of mouse alpha-interferon and human beta 1-interferon genes contain multiple copies of the hexanucleotide sequence AGT-GAA or its variants which are also found in the interferon-stimulated response element of genes transcriptionally induced by interferon. We have examined the similarities between virus and interferon induction of gene expression and the role of AGTGAA and AAT-GAA hexamers in these responses. Hybrid plasmids were constructed by inserting the IE region, the alpha 4 promoter, or the multiple copies of AGTGAA or AAT-GAA 5' to the inactive-45 human immunodeficiency-chloramphenicol acetyltransferase hybrid gene, and their inducible expression was studied in a transient expression assay. In L-cells, multiple hexamers were efficiently induced both by infection with Newcastle disease virus and by interferon treatment; while the alpha 4 promoter and the IE inducible region were induced predominantly by virus rather than by interferon. In order to dissociate the effect of virus and endogenous interferon on the induction process, we examined the gene expression in Vero cells, which have undergone homozygous deletion of type 1 interferon genes, and in VNPT-159 cells, which were derived from Vero cells by insertion of an inducible human interferon beta 1 gene. The results show that while the alpha 4 promoter was efficiently induced only by virus in both cell types, the constructs containing shorter segments of the IE were induced by both virus and interferon in Vero cells. However, the inducibility by interferon was not detected in VNPT-159 cells, suggesting that the presence of endogenous interferon suppresses interferon-induced expression of hexanucleotide repeats and the short inducible region. In contrast, virus inducibility of endogenous interferon-stimulated genes, ISG-15 and ISG-54, was about 100-fold more efficient in VNPT-159 cells than in Vero cells, suggesting that this induction is largely mediated through synthesis of endogenous interferon. Hence, endogenous interferon may play a role in the autoregulation of both interferon genes and interferon-stimulated genes.
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PMID:Virus infection and interferon can activate gene expression through a single synthetic element, but endogenous genes show distinct regulation. 255 Apr 51

A human transient expression assay was used to examine the inducible transcriptional activation of beta interferon (IFN-beta) and IFN-alpha 1 promoters in a homologous cellular environment. Use of 293 cells, an adenovirus DNA-transformed human embryonic kidney cell line, permitted Sendai virus-inducible expression of IFN-beta-CAT hybrid gene. Introduction of the simian virus 40 (SV40) enhancer 5' or 3' to the IFN-CAT gene increased basal (uninduced) levels of chloramphenicol acetyltransferase (CAT) activity; in one construct the SV40 enhancer--IFN-beta regulatory region combination increased the induced CAT activity 50- to 100-fold, suggesting that this may be a generally useful inducible enhancer-promoter combination. No expression from the IFN-alpha-CAT hybrid gene was detected in 293 cells, indicating that human epithelioid cells lack a factor required for expression of the IFN-alpha promoter. However, when the IFN-alpha regulatory region was combined with the SV40 enhancer, a low level of inducible CAT activity was detected in the human transient system.
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PMID:Transient expression of the beta interferon promoter in human cells. 282 99

A human transient expression system was used to measure the influence of simian virus 40 T antigen and adenovirus E1a proteins on the activation of alpha interferon subtype 1 (IFN-alpha 1) and IFN-beta promoters linked to the reporter chloramphenicol acetyltransferase gene. Large T-antigen production, amplified by expression plasmid replication in transfected 293 cells, was able to trans activate the IFN-beta promoter 5- to 10-fold, increasing both the constitutive and Sendai virus-induced levels of expression. Surprisingly, the previously quiescent transfected IFN-alpha 1 promoter in T-antigen-expressing cells displayed a level of inducibility similar to IFN-beta. The endogenous IFN-alpha 1 gene was also inducible to a limited extent in cells expressing T antigen. A truncated IFN-beta promoter deleted to position -37 relative to the CAP site was neither inducible nor trans activated by T antigen, suggesting that sequences required for efficient induction were also needed for trans activation. Since 293 cells express adenoviral E1a proteins, experiments were also performed in HeLa cells to assess the relative contribution of T antigen and E1a proteins to IFN trans activation. In HeLa cells, T-antigen coexpression increased the constitutive level of IFN-beta and IFN-alpha 1 promoter activity without augmenting relative inducibility. Coexpression of T antigen and E1a proteins did not have a cooperative effect on type 1 IFN expression.
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PMID:trans activation of type 1 interferon promoters by simian virus 40 T antigen. 285 Apr 92

The hallmark of "beta 2-interferon (IFN-beta 2)/hepatocyte-stimulating factor/interleukin 6" gene expression is its inducibility in different types of human cells (fibroblasts, monocytes, epithelial cells, and endothelial cells) by different stimuli, which include cytokines such as tumor necrosis factor, interleukin 1 (IL-1) and platelet-derived growth factor, different viruses, and bacterial products such as endotoxin. The activation by cytokines, viruses, and second messenger agonists of the IFN-beta 2 promoter linked to the bacterial chloramphenicol acetyltransferase (CAT) gene was studied after transfection into HeLa cells. A chimeric gene containing IFN-beta 2 DNA from -1180 to +13 linked to the CAT gene was inducible approximately 10-fold by phorbol 12-myristate 13-acetate (PMA), followed, in decreasing order, by pseudorabies and Sendai viruses (7- to 11-fold each); serum (6- to 9-fold); the cytokines tumor necrosis factor, IL-1, and epidermal growth factor (3- to 5-fold each); the cAMP agonists BrcAMP and forskolin and the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (2- to 6-fold each); poly(I).poly(C) (2- to 4-fold); 1,2-diacylglycerol and the calcium ionophore A23187 (1.5- to 2-fold each). Bacterial endotoxin did not activate this IFN-beta 2/CAT fusion gene in HeLa cells. Deletion of the 5' boundary of the IFN-beta 2 DNA from -1180 to -596 in the fusion gene preserved its activation by IL-1, tumor necrosis factor, epidermal growth factor, serum, pseudorabies, and Sendai viruses and by PMA, Br-cAMP, and forskolin; deletion to -225 led to a small reduction (by a factor of 1.5-2) in the responsiveness to serum, PMA, and Sendai virus but not to the other inducers; a further deletion to -112 greatly reduced all responsiveness. Thus, the region between -225 and -113 in IFN-beta 2, which contains DNA motifs similar to the regulatory elements in the human c-fos gene, appears to contain the major cis-acting regulatory elements responsible for the activation of the IFN-beta 2 promoter by several different cytokines, viruses, and second messenger agonists.
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PMID:Activation of the human "beta 2-interferon/hepatocyte-stimulating factor/interleukin 6" promoter by cytokines, viruses, and second messenger agonists. 304 22

We previously isolated a cDNA clone encoding interferon consensus sequence-binding protein (ICSBP), a member of the interferon regulatory factor (IRF) family, that binds to the interferon (IFN)-stimulated response element (ISRE) of many IFN-regulated genes. In this investigation, we studied the functional role of ICSBP by transient cotransfection of ICSBP cDNA with IFN-responsive reporter genes into the human embryonal carcinoma cell line N-Tera2. These cells were shown not to express ICSBP or IRF-2, thus allowing functional analysis of transfected cDNAs. Cotransfection of ICSBP into cells treated with retinoic acid or any of the IFNs (alpha, beta, or gamma) repressed expression of a chloramphenicol acetyltransferase reporter driven by the major histocompatibility complex class I gene promoter. Similarly, ICSBP repressed expression of chloramphenicol acetyltransferase reporters driven by the ISREs of the 2'-5' oligoadenylate synthetase, guanylate-binding protein, and ISG-15 genes in IFN-treated cells. The repression was dependent on the presence of the ISRE in the reporter. Deletion analysis showed that the putative N-terminal DNA binding domain of ICSBP by itself is capable of mediating the repression. Using the same cotransfection conditions as for ICSBP, a similar repression of these reporters was observed with IRF-2. Finally, ICSBP repressed the IRF-1-mediated induction of major histocompatibility complex class I and IFN-beta reporters in the absence of IFN or retinoic acid. Taken together, these results suggest that ICSBP is a negative regulatory factor capable of repressing transcription of target genes induced by IFN, retinoic acid, or IRF-1.
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PMID:Interferon consensus sequence-binding protein, a member of the interferon regulatory factor family, suppresses interferon-induced gene transcription. 767 54


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