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)

Mouse mammary tumor virus (MMTV) is an endogenous murine retrovirus that is expressed in the epithelial cells of the mammary and salivary glands, lungs, kidneys, and seminal vesicles and in the lymphoid cells of the spleen and thymus. Several studies have shown that the long terminal repeat (LTR) of this virus can direct the expression of reporter genes to the same tissues in transgenic mice. To determine whether multiple regulatory elements within the LTR are involved in this tissue-specific expression, we have established lines of transgenic mice containing transgenes that have deletions in the MMTV LTR. Deletions of all LTR sequences upstream of -364 or of LTR sequences from -165 to -665 both result in the expression of linked reporter genes such as the simian virus 40 early region or the bacterial enzyme chloramphenicol acetyltransferase in novel sites, such as the heart, brain, and skeletal muscle; expression of endogenous MMTV and transgenes containing the full-length LTR is not detected in these organs. Negative regulation appears to involve more than one region, since deletion of sequences between either -201 and -471 or -201 and -344, as well as sequences upstream of -364, results in inappropriate expression in heart, brain, and skeletal muscle. Therefore, a negative regulatory element(s) in the MMTV LTR can suppress transcription from the viral promoter in several different organs. This represents the first example of generalized negative regulatory elements that act in many different tissues in transgenic mice to prevent inappropriate expression of a gene.
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PMID:Negative regulation in correct tissue-specific expression of mouse mammary tumor virus in transgenic mice. 170 Feb 74

Previous studies have demonstrated that the entire rat beta-casein (R beta C) gene and a -524/+490 R beta C fragment-chloramphenicol acetyltransferase (CAT) fusion gene are expressed preferentially in the mammary gland of transgenic mice in a developmentally regulated fashion. However, transgene expression was infrequent, less than 1% of that observed for the endogenous gene, and varied as much as 500-fold, presumably due to the site of chromosomal integration. To determine whether a heterologous hormone-responsive enhancer could be used to increase both the level and frequency of expression in the mammary gland, a fragment derived from the mouse mammary tumor virus long terminal repeat containing four hormone response elements (HREs) was inserted into the R beta C promoter at a site not known to contain transcriptional regulatory elements. Transgenic mice generated which carried HRE-enhanced R beta C-CAT fusion genes expressed CAT activity in the mammary glands of all founder lines examined at levels that were on average 13-fold greater than for lines generated with similar constructs not carrying HREs. In the highest expressing line, the level of HRE-enhanced transgene expression was found to be developmentally regulated, increasing 14-fold in the mammary gland from virgin to day 10 of lactation. In this line, expression was also observed in the thymus and spleen; however, the level of CAT activity was 4-fold lower than in the mammary gland and was not developmentally regulated. In adrenalectomized mice, the administration of dexamethasone stimulated CAT expression in the mammary gland but not in the thymus and spleen. These studies demonstrate that in the context of the R beta C promoter, the HRE functions in the mammary gland to increase both the frequency and level of transgene expression.
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PMID:A heterologous hormone response element enhances expression of rat beta-casein promoter-driven chloramphenicol acetyltransferase fusion genes in the mammary gland of transgenic mice. 177 34

TR3 receptor is a human homolog of mouse Nurr77 and N10 protein and the rat NGFI-B protein. A cDNA encoding a chimeric nuclear receptor composed of the N-terminal domain and C-terminal putative ligand-binding domain of the orphan receptor TR3 receptor and the DNA-binding domain of the androgen receptor was constructed. The chimeric receptor, called TR3/AR/TR3 receptor, when expressed in COS-1 monkey kidney cells or PC-3 human prostate tumor cells, cotransfected with an ARE-containing mouse mammary tumor virus long terminal repeat-linked reporter gene encoding chloramphenicol acetyltransferase (CAT), activated CAT expression in the absence of any added factor. The activation was dependent on the amount of expression vector transfected and appeared to be independent of the concentration of serum supplement. Intact TR3 receptor was not active in this system. A TR3/AR/TR3 receptor protein truncated in the putative ligand-binding domain also induced CAT activity. TR3 receptor appears to be a transcriptional factor that activates transcription independently of ligand or binds an endogenous ligand present constitutively in cultured cells.
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PMID:Transcriptional activation by TR3 receptor, a member of the steroid receptor superfamily. 184 11

We analyzed the long terminal repeat (LTR) of mouse mammary tumor virus for sequences that influence its promoter activity by using the chloramphenicol acetyltransferase assay. A series of LTR deletion mutants and recombinants between LTR and simian virus 40 regulatory sequences were used for these studies. Through transfection experiments in three different human cell lines (T47D, MCF-7, and HeLa), we identified a novel mammary cell line-specific enhancer element on a 98-bp BanII fragment (from position -1075 to -978 upstream of the start site of transcription) which interacts with the hormone-responsive element of LTR. We also identified nuclear factors that specifically interacted with this BanII fragment in the nuclear extract from the mammary tumor cell line, T47D, but not from the HeLa cell line.
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PMID:Identification of a novel mammary cell line-specific enhancer element in the long terminal repeat of mouse mammary tumor virus, which interacts with its hormone-responsive element. 184 7

Liposome-mediated gene transfer is useful for DNA transfection into cells in culture. We wondered whether this method could be used to introduce new DNA into the intact lung. Fusion genes containing either the Rous sarcoma virus (RSV) promoter or the mouse mammary tumor virus (MMTV) promoter (which contains glucocorticoid response elements) were linked to the bacterial gene chloramphenicol acetyltransferase (CAT), an enzyme not present in mammalian cells. Plasmids containing the RSV-CAT fusion gene were mixed with cationic liposomes (Lipofectin; BRL, Inc., Grand Island, NY), and single doses were instilled into the cervical trachea of anesthetized rats. Control rats received either liposomes or plasmid. After 24, 48, and 72 h, lungs were perfused free of blood, homogenized, and analyzed for CAT enzyme activity. Liver and kidney tissue were also obtained. We found that rats given either intratracheal liposomes or plasmid had no detectable CAT activity. By contrast, 24 h after instillation of lipid:DNA complexes, lung CAT expression remained elevated for the next 48 h but was barely detectable in liver or kidney. In another group of rats, MMTV-CAT:liposome complexes were instilled intratracheally and then the rats were injected with either dexamethasone or saline. We found that the dexamethasone-treated rats had a 5- to 10-fold higher level of lung CAT expression at 24 and 48 h than the saline-treated controls had; liver and kidney CAT levels were negligible in both groups. Dexamethasone treatment did not increase RSV-CAT expression, indicating that the dexamethasone effect on MMTV-CAT expression was related to the presence of the MMTV promoter.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Localization and induced expression of fusion genes in the rat lung. 184 84

Transfection of HeLa cells with cDNA vectors expressing the wild-type human glucocorticoid receptor (GR) enabled dexamethasone to strongly repress cytokine- and second messenger-induced expression of cotransfected chimeric reporter genes containing transcription regulatory DNA elements from the human interleukin 6 (IL-6) promoter. Deletion of the DNA-binding domain or of the second Zn finger or a point mutation in the Zn catenation site in the second finger blocked the ability of GR to mediate repression of the IL-6 promoter. Unexpectedly, deletion of the first Zn finger, a point mutation in the Zn-catenation site in the first finger, or one in the steroid-specificity domain at the base of the first finger converted GR into a dexamethasone-responsive activator that enhanced basal and interleukin 1-induced IL-6 promoter function. These first-finger mutants of GR also mediated dexamethasone-responsive enhancement of expression of the herpesvirus thymidine kinase-chloramphenicol acetyltransferase (TK-105-CAT and TK-80-CAT) reporter genes but not of the murine mammary tumor virus long terminal repeat-CAT or the c-fos-CAT (pFC700) reporter genes. Wild-type GR was able to specifically bind to DNA fragments containing glucocorticoid response element sequences in both the murine mammary tumor virus and IL-6 promoters, albeit weakly to the latter, in a sequential DNA-binding immunoprecipitation assay. The first-finger mutants of GR, however, were inactive in this assay. Thus, mutations in the first Zn finger unmask unusual promoter-specific activation properties of GR that may not require direct high-affinity binding of the mutant GR to target DNA.
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PMID:Repressor to activator switch by mutations in the first Zn finger of the glucocorticoid receptor: is direct DNA binding necessary? 187 Nov 24

The androgen receptor (AR) is a signal-transducing protein required for sexual differentiation, development, and expression of the male phenotype. A series of human AR deletion mutants were created either by site-directed mutagenesis using restriction enzyme digestion, the polymerase chain reaction, or, for a series of unidirectional NH2-terminal deletions, exonuclease III digestion. Receptor mutants were expressed in monkey kidney COS cells as truncated AR proteins between 20 and 107 kDa as revealed on immunoblots, where wild type AR was a doublet of 114 and 108 kDa. Subcellular localization by immunocytochemical staining demonstrated androgen-dependent nuclear uptake of AR from a perinuclear region of the cytoplasm. A nuclear targeting signal similar in sequence and position to the glucocorticoid receptor and homologous to the SV40 large T antigen was required for androgen-induced nuclear uptake of wild type AR. AR mutants lacking the NH2-terminal and/or steroid binding domains were constitutively nuclear with reduced transcriptional activity. Transcriptional activation by wild type AR was androgen-dependent in cotransfection studies of CV1 cells using the chloramphenicol acetyltransferase reporter gene linked to the mouse mammary tumor virus promoter. Deletion mutagenesis revealed within the NH2-terminal region a domain required for full transcriptional activity and within the steroid binding domain, an inhibitory function, deletion of which yielded a constitutively active receptor. Inhibition of wild type AR by coexpression with an inactive NH2-terminal fragment suggested competition for nuclear factors required for transcriptional regulation. These studies demonstrate a concerted interplay among the domains of the AR protein in regulating gene transcription.
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PMID:Transcriptional activation and nuclear targeting signals of the human androgen receptor. 198 13

Depression is often characterized by increased cortisol secretion caused by hyperactivity of the hypothalamic-pituitary-adrenal axis and by nonsuppression of cortisol secretion following dexamethasone administration. This hyperactivity of the hypothalamic-pituitary-adrenal axis could result from a reduced glucocorticoid receptor (GR) activity in neurons involved in its control. To investigate the effect of reduced neuronal GR levels, we have blocked cellular GR mRNA processing and/or translation by introduction of a complementary GR antisense RNA strand. Two cell lines were transfected with a reporter plasmid carrying the chloramphenicol acetyltransferase (CAT) gene under control of the mouse mammary tumor virus long terminal repeat (a glucocorticoid-inducible promoter). This gene construction permitted assay of the sensitivity of the cells to glucocorticoid hormones. Cells were also cotransfected with a plasmid containing 1,815 bp of GR cDNA inserted in the reverse orientation downstream from either a neurofilament gene promoter element or the Rous sarcoma virus promoter element. Northern (RNA) blot analysis demonstrated formation of GR antisense RNA strands. Measurement of the sensitivity of CAT activity to exogeneous dexamethasone showed that although dexamethasone increased CAT activity by as much as 13-fold in control incubations, expression of GR antisense RNA caused a 2- to 4-fold decrease in the CAT response to dexamethasone. Stable transfectants bearing the GR antisense gene fragment construction demonstrated a 50 to 70% decrease of functional GR levels compared with normal cells, as evidenced by a ligand-binding assay with the type II glucocorticoid receptor-specific ligand [3H]RU 28362. These results validate the use of antisense RNA to GR to decrease cellular response to glucocorticoids.
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PMID:Decreased glucocorticoid receptor activity following glucocorticoid receptor antisense RNA gene fragment transfection. 199 14

The testicular feminized (Tfm) mouse lacks completely androgen responsiveness; and therefore, is unique for studying the role of androgenic steroids in different biological processes. In order to understand the molecular basis of this mutation, 2.8 kilobases of cDNA encoding the Tfm mouse androgen receptor (AR) were amplified with a polymerase chain reaction (PCR) technique. No large deletion in the coding region of the Tfm mouse AR was detected. However, sequence analysis revealed a single base deletion in the coding region of the Tfm AR mRNA. This mutation, which is located in the amino-terminus domain of the receptor, is predicted to cause a frame-shift in translation resulting in a premature termination of AR synthesis at amino acid 412. In vitro translation studies of the recombinant wild type and Tfm AR's demonstrated that the Tfm AR cDNA failed to produce a full-length receptor. Furthermore, the Tfm AR was demonstrated to lack transcriptional activation capability by cotransfection experiments using the Tfm AR with a reporter plasmid of mouse mammary tumor virus long terminal repeat linked to the chloramphenicol acetyltransferase gene. These studies provide evidence of the molecular defect which causes androgen insensitivity in the Tfm mouse.
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PMID:A frame-shift mutation in the androgen receptor gene causes complete androgen insensitivity in the testicular-feminized mouse. 204 77

A cell line was generated from U7 cells (a subline of PC12 rat pheochromocytoma cells) that contains a stably integrated transforming mouse N-ras (Lys-61) gene under the control of the long terminal repeat from mouse mammary tumor virus. Such cells, designated UR61, undergo neuronal differentiation upon exposure to nanomolar concentrations of dexamethasone, as a consequence of expression of the activated N-ras gene (I. Guerrero, A. Pellicer, and D.E. Burstein, Biochem, Biophys. Res. Commun. 150:1185-1192, 1988). Exposure of UR61 cells to either nerve growth factor (NGF) or basic fibroblast growth factor (bFGF) results in a marked induction of c-fos RNA, with kinetics paralleling those of NGF- or bFGF-induced expression of c-fos RNA in PC12 cells. Dexamethasone-induced expression of activated N-ras p21 results in blocking of c-fos RNA induction by NGF or bFGF in a time-dependent manner. Activated N-ras p21-mediated inhibition of c-fos RNA induction in UR61 cells is selective for NGF and bFGF and is not due to selective degradation of c-fos RNA. Normal and transforming N-ras can trans activate the chloramphenicol acetyltransferase gene linked to mouse c-fos regulatory sequences when transient expression assays are performed. Our observations suggest that N-ras p21 selectively interacts with pathways involved in induction of c-fos expression which initiate at the receptors for NGF and bFGF.
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PMID:Oncogene N-ras mediates selective inhibition of c-fos induction by nerve growth factor and basic fibroblast growth factor in a PC12 cell line. 210 19


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