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)

The p10 gene of Autographa californica nucleopolyhedrovirus has two putative AATAAA polyadenylation signals. The downstream signal is used predominantly, as was determined by analysing 3' cDNA ends. This downstream motif is followed by a GT-rich sequence, known to be important for efficient polyadenylation in mammalian systems. To analyse the importance of polyadenylation for p10 gene expression, recombinant viruses with altered 3' untranslated regions (UTRs) were tested using chloramphenicol acetyltransferase (CAT) as a reporter. Surprisingly, after inactivation of the downstream AATAAA motif, CAT expression remained at the same high level as observed with a wild-type 3' UTR. Polyadenylation occurred 24-28 nucleotides further downstream, probably due to an ATTAAA sequence motif. Replacing the p 10 3' UTR with the SV40 early terminator sequence as part of an hsp70-lacZ-SV40 gene cassette, which is commonly used in baculovirus expression vectors, resulted in a reduction in reporter gene expression. Polyadenylation occurred far more efficiently in the original p10 3' UTR than in the SV40 terminator sequence, as was shown by testing the SV40 terminator separately. These results indicate that in order to obtain high levels of foreign gene expression, vectors that provide a wild-type p10 3' UTR are to be preferred over those containing the hsp70-lacZ-SV40 gene cassette. Comparison of the p10 genes of various baculoviruses showed the presence of at least one AATAAA or ATTAAA motif in combination with a GT-rich sequence in the 3' UTR, suggesting an evolutionary conservation of these two elements, thereby maintaining the high level of p10 gene expression.
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PMID:Role of the 3' untranslated region of baculovirus p10 mRNA in high-level expression of foreign genes. 1046 25

Using mouse L929 cells stably transfected with a glucocorticoid receptor (GR)-responsive murine mammary tumor virus-chloramphenicol acetyltransferase (MMTV-CAT) reporter gene (LMCAT2 cells), we have shown that cellular stress (heat or chemical shock) can cause a dramatic increase in the levels of dexamethasone (Dex)-induced CAT gene expression. We refer to this response as the heat shock potentiation effect, or HSPE. As the cellular heat shock response also involves the activation of heat shock transcription factor (HSF), we have, in the present study, examined the role of HSF in the stress potentiation of GR by use of a flavonoid compound, quercetin, recently shown to selectively inhibit the stress response in a variety of human and murine cell lines. Analysis of the HSPE, as well as heat shock protein synthesis and activation of HSF during time-courses of recovery following heat shock, revealed a similar pattern for each response, with peak activities occurring about 16 h after stress. These data suggest a correlation between the activation of both GR and HSF in stressed cells. In L929 cells stably transfected with a CAT reporter plasmid under the control of the HSF-responsive hsp70 promoter (LHSECAT cells), pretreatment with quercetin was found to cause a dose- and time-dependent inactivation of HSF activity following heat shock, but only when added before the stress event. In LMCAT2 cells, quercetin similarly inhibited both heat and chemical shock potentiation of Dex-induced GR activity. This activity of quercetin was not the result of post-transcriptional or general cytotoxic properties, as quercetin (1) did not significantly affect GR or HSF activities when added after the stress event, (2) did not reduce CAT gene expression as controlled by the constitutive SV40 early promoter, and (3) did not alter normal (non-stress), Dex-induced MMTV-CAT expression. Thus, quercetin appears to be an effective and selective inhibitor of HSF stress-induced activation and its ability to prevent the stress potentiation of GR suggests either a direct or indirect involvement by stress-activated HSF in this process, or the existence of a regulatory step common to both the heat shock and HSPE responses.
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PMID:Inhibition of heat shock factor activity prevents heat shock potentiation of glucocorticoid receptor-mediated gene expression. 1059 Aug 36

We present structural and comparative analysis of the flanking regions of the rat hsp70.1 stress gene. Several repetitive sequences, microsatellites and short interspersed repetitive elements (SINEs) were found, as well as a significant gap in the 3' UTR, as compared to the orthologous mouse gene. We also show that the complex microsatellite region composed of partially overlapping inverted repeat and long homopurine-homopyrimidine sequence, which is localized 1.8 kbp upstream of the transcription start site, is capable to adopt non-B DNA structures (an H-DNA and a cruciform structure) in vitro. Functional analysis performed with the use of various fragments of the 5'end flanking regions ligated to the chloramphenicol acetyltransferase (CAT) reporter gene revealed a crucial role of cooperation between heat shock element (HSE) regulatory sequences, while none of the three HSEs alone is able to drive efficient heat induced transcription of the reporter gene. We also found that the microsatellite region does not influence transcription by itself, however, it abolishes the effect of the adjacent putative silencing element. To our knowledge, this is a first extensive structural and functional analysis of the promoter region of the mammalian heat inducible hsp70i gene localized distally to the hsp70-related spermatid-specific gene in the major histocompatibility complex III.
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PMID:Structure of gene flanking regions and functional analysis of sequences upstream of the rat hsp70.1 stress gene. 1252 28

The aim of this study was to assess the potential of a human cell line containing the hsp70 promoter linked to the chloramphenicol acetyltransferase reporter gene in evaluating the toxic potential of complex mixtures. Cells were exposed to eluates of industrial wastes and the cellular responses were compared with the metal contents of the samples and with standardized aquatic (microalgal growth inhibition, daphnia Immobilization, bacterial luminescence inhibition, Ceriodaphnia dubia reproduction inhibition) and terrestrial (earthworm lethality, plant growth inhibition) tests. The hsp70 promoter was significantly induced by 11 of 14 samples, with different dose-response patterns. Significant correlations of in vitro induction potency with aquatic ecotoxicity, especially with chronic tests, and with the metal contents of the samples were observed. Our study provides new information on the relevance of hsp70 gene induction as a criterion of toxicity and suggests its usefulness for the detection of toxicity associated with metallic pollution in complex mixtures.
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PMID:Evaluation of an in vitro hsp70 induction test for toxicity assessment of complex mixtures: comparison with chemical analyses and ecotoxicity tests. 1254 40


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