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)

A rainbow trout major heat-shock-protein-like gene (hsp 70) and corresponding cDNA clones were isolated by hybridization to heterologous hsp70 probes. DNA sequencing revealed that this gene is structurally similar to a mammalian heat-shock-cognate hsc70 gene and consists of eight introns. Northern blot and primer extension analyses showed that the corresponding mRNA is constitutively abundant in different trout tissues and salmonid cell lines. Fragments of the isolated gene containing the -900 - +30 and -217 - +58 sequence were linked to a bacterial chloramphenicol acetyltransferase reporter gene and transiently transfected into salmonid cells. The expression pattern of these constructs supports our conclusion that the isolated genomic and cDNA clones correspond to a trout heat-shock-cognate hsc70 gene.
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PMID:Molecular cloning and characterization of a constitutively expressed heat-shock-cognate hsc71 gene from rainbow trout. 137 53

We established basic conditions for transient gene expression and selection of antibiotics in the cultured cell line of silkworm, Bombyx mori, by use of the promoter of the heat shock protein (hsp70) gene of Drosophila melanogaster. The control promoter (hsp70) promoted the expression of chloramphenicol acetyltransferase (CAT) gene ligated at the downstream, dependent on the orientation of the promoter in the silkworm cell. The cell line is able to be supplied for the promoter assay of the silkworm genes. The concentration for the drug selection was determined as 0.75 mg/ml on neomycin analog, G418 (geneticin).
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PMID:Basic conditions for the drug selection and transient gene expression in the cultured cell line of Bombyx mori. 148 68

Amber (UAG) and opal (UGA) nonsense suppressors were constructed by oligonucleotide site-directed mutagenesis of two Drosophila melanogaster leucine-tRNA genes and tested in yeast, Drosophila tissue culture cells and transformed flies. Suppression of a variety of amber and opal alleles occurs in yeast. In Drosophila tissue culture cells, the mutant tRNAs suppress hsp70:Adh (alcohol dehydrogenase) amber and opal alleles as well as an hsp70:beta-gal (beta-galactosidase) amber allele. The mutant tRNAs were also introduced into the Drosophila genome by P element-mediated transformation. No measurable suppression was seen in histochemical assays for Adhn4 (amber), AdhnB (opal), or an amber allele of beta-galactosidase. Low levels of suppression (approximately 0.1-0.5% of wild type) were detected using an hsp70:cat (chloramphenicol acetyltransferase) amber mutation. Dominant male sterility was consistently associated with the presence of the amber suppressors.
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PMID:Drosophila nonsense suppressors: functional analysis in Saccharomyces cerevisiae, Drosophila tissue culture cells and Drosophila melanogaster. 217 93

We have examined the expression of the heat shock protein (hsp70) gene in human cells. The transcription of the hsp70 gene and accumulation of cytoplasmic hsp70 mRNA is induced by serum stimulation. Populations of HeLa cells and human embryonic kidney cells (cell line 293) were serum starved. Upon serum stimulation, the level of hsp70 mRNA transiently increases between 12 and 18 hr to a 10-fold higher level. The increased levels of hsp70 mRNA can be accounted for by a 10- to 15-fold increase in the rate of transcription of the hsp70 gene. When cells were serum-stimulated in the presence of an inhibitor of DNA synthesis, 1-beta-D-arabinofuranosylcytosine (araC), the levels of hsp70 mRNA were induced to only 20% of the maximal level detected in the absence of the inhibitor. This suggests that the expression of the hsp70 gene is coupled to DNA synthesis. The cloned human hsp70 gene contains regulatory sequences that confer serum-stimulated transcriptional control. The endogenous hsp70 gene and the transfected chimeric gene containing sequences upstream of the hsp70 gene fused to bacterial chloramphenicol acetyltransferase are both temporally expressed in stable transfectants of cell line 293 cells. The endogenous hsp70 mRNA and the chimeric mRNA reach maximum levels 12-18 hr after serum stimulation.
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PMID:Transcription of the human hsp70 gene is induced by serum stimulation. 386 19

The rat hst70 gene belongs to a heat shock hsp70 multigene family and its expression has been detected so far solely in spermatocytes. To investigate the cis-elements responsible for testis-specific expression of the hst70 gene we produced several lines of transgenic mice carrying fragments of the 5'-flanking regions of the hst70 gene fused to the chloramphenicol acetyltransferase (CAT) reporter gene. Hybrid genes of series B were constructed such that, besides the 780 bp, 343 bp and 163 bp 5'-flanking region these plasmids contained no other sequences of the hst70 gene. In hybrid genes of series D the CAT gene was ligated to 343 bp and 252 bp 5'-flanking regions together with the 57 bp of the 5'-end nontranslated (leader) sequences of the hst70 gene. We found that in 780/B, 343/B, 343/D and 252/D adult mice the transgene was specifically and highly expressed in testes. In developing testes the high CAT activity appeared in transgenic mice aged 3 weeks and older. None of the three 163/B transgenic lines exhibited CAT activity in any tissue analyzed. In all CAT expressing lines a weak but significant CAT activity (up to 5% of that in testis) was detected also in the brain. RNase protection assay confirmed that the endogenous hst70 gene transcripts are present in testis as well as in brain of nontransgenic rats and mice. Our data show that the cis-regulatory sequences responsible for testis-specific and developmentally regulated expression of the hst70 gene are localized within the 252 bp region 5' to the gene and neither the 5'-end nor 3'-end nontranslated sequences of the gene are important for this specificity.
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PMID:A 252 bp upstream region of the rat spermatocyte-specific hst70 gene is sufficient to promote expression of the hst70-CAT hybrid gene in testis and brain of transgenic mice. 749 63

Heat shock factors (HSF) are the transcriptional activators of the heat shock response. The conversion of constitutively expressed HSF to a form that can bind DNA requires the trimerization of the protein, involving leucine zipper interactions as shown for yeast, Drosophila, chicken and human HSFs. Like other metazoan HSFs, the endogenous Arabidopsis HSF displays heat shock-inducible DNA-binding activity in gel retardation assays. The heat shock-inducible binding of a recombinant Arabidopsis HSF (ATHSF1) expressed in Arabidopsis plants suggests that ATHSF1 is the major heat shock factor regulating the heat stress response. However, on transient expression in Drosophila and human cells, ATHSF1 fails to exhibit proper regulation, as demonstrated by constitutive binding to DNA, and by constitutive expression of a chloramphenicol acetyltransferase (CAT) reporter gene under the control of the Drosophila hsp70 promoter. These results suggest that the regulation of ATHSF1 is normally dependent on a specific factor that inhibits the DNA-binding and transcriptional activities under non-heat shock conditions.
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PMID:Arabidopsis heat shock factor is constitutively active in Drosophila and human cells. 765 36

The Larval serum protein-2 gene (Lsp-2) of Drosophila melanogaster is uniquely expressed in the fat body tissue from the beginning of the third instar to the end of adult life. Accumulation of the larval Lsp-2 transcript is enhanced by 20-hydroxyecdysone. To study the molecular basis for ecdysone regulated Lsp-2 activity, deletion mutants of the Lsp-2 5'-flanking region were constructed by fusion to either the Escherichia coli chloramphenicol acetyltransferase (CAT) gene or to an hsp70-lacZ hybrid gene encoding beta-galactosidase. Constructs transfected into Drosophila S2/M3 cells were shown to confer transient ecdysone inducibility on the reporter genes. A single functional ecdysone response element (EcRE) was localized at position -75 relative to the Lsp-2 transcription initiation site. In gel mobility shift assays using fat body nuclear extracts or nuclear receptors synthesized in vitro, a 27-bp sequence harboring the EcRE bound both the Drosophila ecdysone receptor and the Drosophila retinoid-X homologue, Ultraspiracle, in a cooperative manner. Competition experiments indicate that the affinity of the Lsp-2 EcRE for the ecdysone receptor complex is comparable to that of the canonical EcRE of the hsp27 gene and is at least 4-fold greater than that of Fbp1, another fat body-specific Drosophila gene. Our results suggest that structural features of this EcRE determine its ability to induce ecdysone responsiveness at a lower ligand concentration and may form the basis for differential hormone responsiveness within the fat body.
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PMID:Characterization of an EcR/USP heterodimer target site that mediates ecdysone responsiveness of the Drosophila Lsp-2 gene. 854 20

To study of structure of RNA polymerase (pol) II transcription units a nd the influence of temperature on the regulation of gene expression in Trypanosoma brucei, and hsp70 intergenic region promoter was characterized. In T. brucei, the hsp70 locus contains, from 5' to 3', a cognate hsp70-related gene (gene 1) which is separated by about 6 kb of DNA from a cluster of five identical hsp70 genes (genes 2 to 6). Transcription proceeds on the entire 23-kb locus, and polycistronic transcription occurs in hsp70 genes 2 to 6. Transcription of hsp70 genes 2 to 6 is only moderately sensitive to UV irradiation, indicating that it cannot be driven by a single far-upstream promoter, which suggests that promoters could be located in the region close to the hsp70 coding region. Transient transformations demonstrated that sequences located upstream of hsp70 gene 2 and in the intergenic region between hsp70 genes 2 and 3 are able to direct transcription of the reporter gene, the chloramphenicol acetyltransferase (CAT) gene. The plasmid DNA driven by the hsp70 intergenic region promoter gave CAT activity approximately 85-fold above to background level. This is equivalent to approximately 1% of that derived from a CAT plasmid driven by the procyclic acidic repetitive protein gene promoter, which is controlled by RNA pol I. The hsp70 intergenic region promoter can drive alpha-amanitin-sensitive transcription at an internal position of the chromosome as well as an episome, suggesting that it is controlled by RNA pol II. However, this hsp70 intergenic region promoter, along with the 3' splice site and the 5' untranslated region of the hsp70 genes that controls the transcription of the reporter gene, cannot up-regulate the expression of the reporter gene during heat shock. This result is consistent with the previous observation that expression of the hsp70 genes in T. brucei is mainly controlled at the posttranscriptional level.
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PMID:An RNA polymerase II promoter in the hsp70 locus of Trypanosoma brucei. 862 66

We have previously examined the efficiency of two Drosophila melanogaster promoters to enable reporter gene expression in embryos of the Australian sheep blowfly, Lucilia cuprina. Both the hsp70 heat-shock promoter and the actin5C promoter resulted in low levels of expression of a reporter gene in these embryos. In this study, the D. pseudoobscura hsp82 promoter (phsp82) was tested for its ability to direct the expression of the Escherichia coli chloramphenicol acetyltransferase-encoding gene (cat). We report that the level of CAT activity in L. cuprina embryos was comparable to that obtained with the same construct in D. melanogaster, indicating that phsp82 functions efficiently in this non-drosophilid insect. The results suggest that phsp82 may be utilised in other non-drosophilid insects in which poor expression levels are obtained from constructs containing the hsp70 or actin5C promoters.
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PMID:The 5' regulatory region from the Drosophila pseudoobscura hsp82 gene results in a high level of reporter gene expression in Lucilia cuprina embryos. 891 99

Transcriptional activation of heat shock genes is a reversible and multistep process involving conversion of inactive heat shock factor 1 (HSF1) monomers into heat shock element (HSE)-binding homotrimers, hyperphosphorylation, and further modifications that induce full transcriptional competence. HSF1 is controlled by multiple regulatory mechanisms, including suppression by additional cellular factors, physical interactions with HSP70, and integration into different cellular signaling cascades. However, the signaling mechanisms by which cells respond to stress and control the HSF1 activation-deactivation pathway are not known. Here we demonstrate that HSP90, a cellular chaperone known to regulate several signal transduction molecules and transcription factors, functions in the regulation of HSF1. The existence of HSF1-HSP90 heterocomplexes was shown by coimmunoprecipitation of HSP90 with HSF1 from unshocked and heat-shocked nuclear extracts, recognition of HSF1-HSE complexes in vitro by using HSP90 antibodies (Abs), and recognition of HSF1 in vivo by HSP90 Abs microinjected directly into oocyte nuclei. The functional impact of HSP90-HSF1 interactions was analyzed by using two strategies: direct nuclear injection of HSP90 Abs and treatment of cells with geldanamycin (GA), an agent that specifically blocks the chaperoning activity of HSP90. Both HSP90 Abs and GA delayed the disassembly of HSF1 trimers during recovery from heat shock and specifically inhibited heat-induced transcription from a chloramphenicol acetyltransferase reporter construct under control of the hsp70 promoter. HSP90 Abs activated HSE binding in the absence of heat shock, an effect that could be reversed by subsequent injection of purified HSP90. GA did not activate HSE binding under nonshock conditions but increased the quantity of HSE binding induced by heat shock. On the basis of these findings and the known properties of HSP90, we propose a new regulatory model in which HSP90 participates in modulating HSF1 at different points along the activation-deactivation pathway, influencing the interconversion between monomeric and trimeric conformations as well as transcriptional activation. We also put forth the hypothesis that HSP90 links HSF1 to cellular signaling molecules coordinating the stress response.
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PMID:HSP90 interacts with and regulates the activity of heat shock factor 1 in Xenopus oocytes. 971 May 78


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