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)

Antigen triggering of the T-cell receptor results in an accumulation of activated GTP-bound p21ras protein. To assess the role of ras protein in T-cell activation we have cotransfected the murine thymoma line EL4 with a construct capable of expressing a constitutively active, oncogenic form of Ha-ras and a reporter construct containing the human interleukin-2 promoter fused upstream of the bacterial gene for chloramphenicol acetyltransferase. We show that the ras oncoprotein contributes to interleukin-2 promoter activation. Its pattern of synergism with a calcium ionophore or the lymphokine interleukin-1 indicates that it replaces a signal mediated by protein kinase C. Interleukin-2 promoter activity in the presence of ras oncoprotein was inhibited by H7, a potent inhibitor of protein kinase C, but not by HA1004, an inhibitor of cyclic nucleotide-dependent kinase, suggesting that protein kinase C mediates the ras effect. In addition, we show that in these cells, expression of activated ras results in activation of a synthetic promoter containing several copies of an NF kappa B binding site.
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PMID:Interleukin-2 promoter activation in T-cells expressing activated Ha-ras. 153 20

The proteins encoded by cellular and viral src genes are believed to be involved in the transmission of mitogenic signals, the nuclear recipients of which are largely unknown. In this work, we report that four different v-src-transformed cell lines from three different species possess elevated levels of junB transcripts. Transient expression of junB promoter-chloramphenicol acetyltransferase constructs in NIH 3T3 cells was used to demonstrate that the increase in junB transcripts was specifically associated with v-src expression and could not be recapitulated with a c-src, v-H-ras, or v-raf expression vector. Deletion mutants were used to localize the v-src-responsive region in the junB promoter to a 121-nucleotide region encompassing the CCAAT and TATAA elements. This region is distinct from one in the 5' untranslated region of the junB gene which is required to maintain its high-level basal expression. Point mutagenesis of the junB TATAA box completely abolished v-src responsiveness, suggesting that proteins which bind to this element are modified by src transformation. Several v-src and c-src mutants were used to demonstrate that elevated tyrosine kinase activity of src proteins is required for the observed effects on junB expression. Finally, homology between the TATAA box regions of junB and the unrelated but src-responsive gene 9E3/CEF-4 suggests that modulation of gene activity through proteins which bind to this region may be a recurrent, although not exclusive, theme in src transforming action. Our results suggest that src proteins may modulate some nuclear effectors through pathways not involving cellular ras or raf gene products.
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PMID:Regulation of the junB gene by v-src. 163 Apr 51

Here the adeno-associated virus Rep78 gene product was found to inhibit the expression of the chloramphenicol acetyltransferase and the bladder cancer-derived EJ-H-ras coding sequences when they were under the control of the natural cellular H-ras regulatory sequences. However, Rep78 had little or no effect on the expression of these same coding sequences when they were under the control of the regulatory sequences of the murine osteosarcoma virus long terminal repeat. These data indicate that the inhibition of H-ras by Rep78 depends upon sequences present within the cellular H-ras upstream regulatory region. Furthermore, these and earlier data indicate that Rep78 functions as an "antioncogene" or transformation suppressor gene, inhibiting H-ras as well as several viral oncogenes.
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PMID:Inhibition of H-ras expression by the adeno-associated virus Rep78 transformation suppressor gene product. 164 67

Insulin induces a rapid activation of p21ras in NIH 3T3 and Chinese hamster ovary cells that overexpress the insulin receptor. Previously, we suggested that p21ras may mediate insulin-induced gene expression. To test such a function of p21ras more directly, we studied the effect of different dominant inhibitory mutants of p21ras on the induction of gene expression in response to insulin. We transfected a collagenase promoter-chloramphenicol acetyltransferase (CAT) gene or a fos promoter-luciferase gene into NIH 3T3 cells that overexpressed the insulin receptor. The activities of both promoters were strongly induced after treatment with insulin. This induction could be suppressed by cotransfection of two inhibitory mutant ras genes, H-ras(Asn-17) or H-ras(Leu-61,Ser-186). In particular, insulin-induced activation of the fos promoter was inhibited completely by H-ras(Asn-17). These results show that p21ras functions as an intermediate in the insulin signal transduction route leading to the induction of gene expression.
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PMID:Two dominant inhibitory mutants of p21ras interfere with insulin-induced gene expression. 165 21

A deletion mutant of the human melanoma-associated ME491 antigen gene starting at the first intron (lambda R31) differentially mediates the antigen expression depending on the cell type. Cryptic promoter activity residing in a 270-base-pair (bp) fragment of the first intron was examined by primer extension analysis and recombinant chloramphenicol acetyltransferase (CAT) assay. The cryptic promoter, further localized within a 153-bp fragment (fr153BN), exerted its effect in Ltk- and H-ras-transformed NIH3T3 (3T3-Hras) but not in parental NIH3T3 cells. The results suggested that the cryptic promoter was associated with a novel ras-responsive positive regulatory element, since fr153BN did not contain an AP-1-binding sequence motif, known as the ras-responsive enhancer element. The cryptic promoter activity of fr153BN was suppressed by an upstream 121-bp fragment (fr121SB) which contained a consensus sequence motif for binding of a repressor protein, GC factor, and regions showing sequence similarity with putative cis-acting repressor elements found in the vimentin gene. The degree of the suppression was greater in 3T3-Hras than in Ltk- cells. These positive and negative regulatory elements may be differentially involved in the regulation of ME491 antigen expression depending on the cell type.
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PMID:Activation and suppression of a cryptic promoter in the intron of the human melanoma-associated ME491 antigen gene. 175 82

Transcriptionally active domains have been identified and located within the 5'-region of the human normal and mutant T24 H-ras1 promoters, and have been characterised by linkage to the coding sequences of the bacterial chloramphenicol acetyltransferase (CAT) gene or by using DNaseI foot-printing analysis of the promoter sequence. It has been shown, using the latter method, that Sp-1 transcription factor binds to six GC sequences within the H-ras promoter. In the present study we have used unfractionated nuclear protein preparations from HeLa cells and a gel retardation assay to analyse specific binding of nuclear protein preparations from HeLa cells and a gel retardation assay to analyse specific binding of nuclear factors to several oligonucleotide sequences of the human H-ras1 promoter. Our data demonstrate the presence of three Spl specific binding sequences in the T24 promoter, one of them containing a Sp-1 consensus GGCGGC absent in the normal H-rasl promoter.
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PMID:Sp1 specific binding sites within the human H-ras promoter: potential role of the 6 bp deletion sequence in the T24 H-ras1 gene. 177 41

Differentiation of skeletal myoblasts is accompanied by induction of a series of tissue-specific genes whose products are required for the specialized functions of the mature muscle fiber. The program for myogenic differentiation is subject to negative control by several peptide growth factors and by the products of mutationally activated ras oncogenes, which persistently activate intracellular cascades normally triggered by specific growth factors. Previously, we reported that induction of the muscle creatine kinase (mck) gene during myogenesis was dependent on a distal upstream enhancer that cooperated with a proximal promoter to direct high levels of expression in developing muscle cells (E. A. Sternberg, G. Spizz, W. M. Perry, D. Vizard, T. Weil, and E. N. Olson, Mol. Cell. Biol. 8:2896-2909). To investigate the mechanisms whereby ras blocks the induction of muscle-specific genes, we have examined the ability of mck 5' regulatory elements to direct expression of the linked reporter gene for chloramphenicol acetyltransferase (cat) in C2 myoblasts bearing mutant N-ras and H-ras oncogenes. In this paper we report that expression of activated ras alleles abolishes activity of the mck upstream enhancer but does not affect the activity of the mck promoter. The ability of ras to repress the expression of mck-cat fusion genes that have been transfected either transiently or stably into myoblasts suggests that ras may exert its effects on muscle-specific genes through mechanisms independent of chromatin configurations or DNA methylation. These results also suggest that ras blocks establishment of the myogenic phenotype by preventing the accumulation of regulatory factors required for transcriptional induction of muscle-specific genes.
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PMID:A ras-dependent pathway abolishes activity of a muscle-specific enhancer upstream from the muscle creatine kinase gene. 265 1

Human herpesvirus 6 strain U1102 (HHV-6A) was shown to contain a 1,473-bp functional transformation suppressor gene (ts). ts exhibited 24% identity and 51% similarity to adeno-associated virus type 2 Rep68/78. Like adeno-associated virus type 2 Rep68/78, HHV-6A ts suppressed H-ras transformation of NIH 3T3 cells. Suppression of H-ras transformation was eliminated by translation termination linker mutation at amino acid 25, 125, or 245. These data indicated the importance of the C-terminal portion of the ts protein. H-ras transformation was suppressed by ts only when H-ras was expressed by its endogenous H-ras promoter and not when it was expressed by the heterologous murine osteosarcoma virus long terminal repeat (LTR). Furthermore, ts suppressed chloramphenicol acetyltransferase (CAT) activity when the CAT gene was expressed from the H-ras promoter but not the murine osteosarcoma virus LTR promoter. Taken together, the data showed that ts suppressed H-ras transformation at the level of the H-ras promoter. To further identify the interaction of ts with transcriptional regulatory elements, the human immunodeficiency virus type 1 (HIV-1) LTR was used. This promoter was selected because it has well-defined transcriptional regulatory elements for both basal and activated transcription, because its activity is inhibited by the Rep68/78 gene, and because both HHV-6 and HIV-1 naturally infect CD4+ T cells in vivo and have been shown to infect the same cell in vitro. ts suppressed expression from both wild-type and upstream mutant HIV-1 LTR-CAT constructs. However, downstream HIV-1 TAR mutations reversed ts suppression, indicating that TAR is one of the critical elements involved. The data presented demonstrated that HHV-6A ts functionally suppressed H-ras transformation and HIV-1 LTR expression and thus that it may be useful in future gene therapy.
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PMID:Human herpesvirus 6A ts suppresses both transformation by H-ras and transcription by the H-ras and human immunodeficiency virus type 1 promoters. 760 62

The involvement of p21ras in the induction of the early activation antigen CD69 was investigated in T cells. Expression of a v-Ha-ras coding for a constitutively active ras protein in Jurkat cells resulted in CD69 induction on the cell surface. Transfected ras was shown to be constitutively activated and functionally efficient, since it could be immunoprecipitated in the guanosine triphosphate (GTP)-bound form and it induced transactivation of an AP-1 consensus-chloramphenicol acetyltransferase reporter gene. The requirement for ras activation in T cell receptor (TcR) CD3-mediated CD69 induction was also investigated. The expression of a dominant negative c-Ha-ras-N17 mutant markedly reduced the amount of GTP that could be immunoprecipitated from ras proteins after TcR/CD3 triggering in Jurkat cells, and concomitantly decreased TcR/CD3-mediated CD69 induction. These results suggest a central role for ras in TcR/CD3-mediated CD69 expression in T cells.
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PMID:Involvement of p21ras activation in T cell CD69 expression. 790 94

The ras gene family encodes 21K proteins that reside on the inner face of the plasma membrane and bind GTP and GDP with an equally high affinity. Cotransfection of NIH 3T3 cells with a mammalian expression vector containing a viral Harvey-ras (v-Ha-ras) cDNA, together with a plasmid (pCMVCAT) carrying the immediate early (IE) enhancer of the murine cytomegalovirus (MCMV) linked to the chloramphenicol acetyltransferase (CAT) reporter gene strongly stimulated CAT activity. Basal levels of pCMVCAT expression as well as trans-activation by v-ras plasmid were both inhibited by cotransfection of an expression vector containing the dominant inhibitory mutant gene Ha-ras Asn-17. This indicates that the p21ras protein is responsible for these activities. High pCMVCAT activation was also observed in cell lines carrying stably transfected ras oncogenes, activated by point mutation or amplification. To define the cis-acting DNA elements in the MCMV IE enhancer responsible for this trans-activation by p21ras protein, we constructed several plasmids containing the CAT gene under control of MCMV IE enhancers that were deleted in different regions. The CAT assays demonstrated that several sequences were responsive to p21ras protein. These sequences are scattered throughout the IE enhancer, upstream of the transcription start site, and contain responsive elements that are homologous to the binding sites for cellular transcription factors such as NF kappa B, AP1, ATF and SP1. Activation of the p21ras protein may thus be one of the signals that regulate IE genes transcription during MCMV infection.
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PMID:Trans-activation of the mouse cytomegalovirus immediate early gene enhancer by ras oncogenes. 802 97


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