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 complementary DNA (cDNA) of hepatitis delta virus (HDV), constructed as tandem repeats under appropriate exogenous promoters, has been used to initiate viral replication in transfection experiments. Whether the structure of tandem repeats is essential has not yet been systematically examined. In this study, expression vectors containing only an HDV cDNA monomer, permutated at unique position of the genome, were shown still able to initiate viral replication. Furthermore, HDV cDNA monomer, separated from plasmid sequences by restriction enzyme digestion, also could be used to initiate viral RNA replication. The competence of HDV cDNA alone to direct viral RNA production suggested the presence of cryptic internal promoter-like elements. Such elements actually demonstrated by the chloramphenicol acetyltransferase assay. Therefore, HDV cDNA, in contrast to that of viroids, could be used to initiate viral RNA replication in monomeric form. This observation simplified the use of HDV cDNA for studying viral biology in transfection systems.
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PMID:Hepatitis delta virus cDNA monomer can be used in transfection experiments to initiate viral RNA replication. 821 49

The precursor peptide of TRH (prepro-TRH) contains five copes of pro-TRH linked by other peptide sequences. These peptides are coprocessed with TRH in the median eminence of the hypothalamus and released into the portal circulation, rendering this family of peptides available to act at the level of the anterior pituitary. Therefore, we tested the potential bioactivity of one cryptic peptide, prepro-TRH amino acids 160-169 [prepro-TRH-(160-169)], in a TRH-responsive pituitary cell line (GH3). In a heterologous TSH expression assay, we found that prepro-TRH-(160-169) stimulated TSH beta gene promoter activity in a time- and dose-dependent manner; moreover, the effect of prepro-TRH-(160-169) was more rapid and of greater magnitude than that of TRH on TSH beta-directed chloramphenicol acetyltransferase synthesis. In the same cells, we found that prepro-TRH-(160-169) stimulated PRL synthesis and secretion, but the effect was similar in magnitude and duration to that of TRH. The effect of prepro-TRH-(160-169) appears to be additive to that of TRH, suggesting that prepro-TRH-(160-169) may act through a mechanism separate from that of TRH. Thus, prepro-TRH-(160-169) has potent endocrinological effects at the level of the genome.
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PMID:A cryptic peptide from the preprothyrotropin-releasing hormone precursor stimulates thyrotropin gene expression. 834 17

Plasmid constructs containing a putative Trypanosoma cruzi rRNA promoter and transcription start point upstream from the bacterial chloramphenicol acetyltransferase (CAT) reporter gene were transfected into cultured T. cruzi epimastigotes to verify the presence of a promoter activity. Constructs bearing the putative promoter and a 3' trans-splicing acceptor site in the proper orientation yielded approx. two orders of magnitude greater CAT expression than that previously observed with the T. cruzi spliced leader (SL) gene promoter. In contrast, similar constructs lacking the known 3' splice site yielded reduced but readily measurable expression suggesting that sequences near the promoter may function as cryptic 3' splice sites. A repeated sequence upstream from the putative basal rRNA promoter in a position analogous to rRNA gene enhancer elements in other eukaryotes did not enhance expression from the T. cruzi rRNA promoter. Finally, these constructs were functional in some but not all T. cruzi isolates, and were inactive in other kinetoplastid species, suggesting that the T. cruzi rRNA promoter may have a limited host range.
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PMID:Transient expression mediated by the Trypanosoma cruzi rRNA promoter. 853 93

The bacterial gene chloramphenicol acetyltransferase (CAT) is a widely used reporter in both in-vitro and in-vivo studies of genetic regulation. We have recently generated novel rat transgenic lines carrying an arylalkylamine N-acetyltransferase (AA-NAT) promoter-reporter construct in which CAT (with associated SV40 small-t antigen sequence) is the reporter. In addition to the predicted transgene transcript (1.9 kb), we identified an abundant 1.5 kb transcript which derives from an alternative splicing event that utilises a cryptic splice donor site located within the CAT gene. The native CAT open reading frame (ORF) is lost in the 1.5 kb transcript, and a western analysis has shown that protein deriving from an aberrant open reading frame is not expressed at detectable levels.
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PMID:Use of a cryptic splice donor site in the chloramphenicol acetyltransferase (CAT)-SV40 small-t antigen cassette generates alternative transcripts in transgenic rats. 1085 70

Transfection analyses are an informative method to assess the activity of specific promoter or enhancer elements in mammalian cells. Commercially available reporter vectors can be extremely useful investigative tools for such studies. This study reports that the pCAT 3- and pGL3-promoter vectors display cryptic responsiveness to androgens when they contain a DNA insert, while the empty vector, a commonly used negative control, is nonresponsive. Our studies initially aimed to characterize novel androgen-responsive DNA sequences in human genomic DNA through transactivational analyses. An isolated DNA fragment, designated ARC-3, contained three putative androgen response element "half-sites" and was androgen-responsive when cloned into the pCAT3-promoter vector. While we originally believed this to be a novel enhancer element, subsequent analyses of this clone revealed that this vector displays cryptic activity in the presence of an androgen. This was confirmed by cloning several unrelated DNA fragments that did not contain any known classic response elements into the pCAT3-promoter vector, all of which were found to be responsive. The empty vector (negative control) was again nonresponsive. The ARC-3 DNA fragment was also weakly responsive to stimulation when cloned into the pGL3-promotor vector, which is identical to the pCAT3-promoter vector, with the exception of an intron located 5' of the chloramphenicol acetyltransferase gene, and the reporter genes. This work demonstrates that both the pCAT3- and pGL3-promoter vectors are inappropriate to assess androgen-responsive enhancers and emphasizes the importance of the careful selection of reporter vectors and controls when conducting transactivational analysis.
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PMID:Aberrant cryptic responsiveness of the pCAT 3- and pGL3-promoter reporter vectors. 1295 75

The human haptoglobin-related gene (HPR) gene codes for a haptoglobin-related protein (Hpr), a component of trypanosome lytic factor which circulates in plasma in small quantities. Except for the presence of a retrovirus-like element, RTVL-Ia, in intron 1, HPR is 92% identical in sequence to the closely linked human haptoglobin gene (HP) gene coding for haptoglobin. We have explored experimentally in tissue culture and in vivo in mice and in humans the influence of the retroviral-like sequence type Ia (RTVL-Ia) element on HPR expression. Transient expression in HepG2 cells of plasmids carrying the HPR promoter joined by a shortened version of intron 1 to the chloramphenicol acetyltransferase (CAT) vector showed that fragments containing the 5' long terminal repeat (LTR) had no significant effect. In contrast, a gag-pol related part and a pol-env-3'LTR related part of RTVL-Ia decreased expression to 20% and 40% of that in their absence but only when they were in naturally occurring orientation. The latter fragment that contains sequences reminiscent of elements essential for retrovirus viability, such as a splicing acceptor site, TATA box and polyA addition signal sequence, was further tested in site-specific transgenic mice. Similar to in vitro experiment, insertion of this fragment into an HPR transgene in mice reduced HPR expression to 50% compared to a transgene without the insert, but none of the viral sequence motifs appear to explain this effect. Instead, we found within the fragment two cryptic splicing donor sites whose products were present in transgenic mouse and in human liver RNA. Our data suggest that a combination of multiple small effects of RTVL-Ia including aberrant splicing accounts for the low (6%) expression of the present-day HPR relative to HP.
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PMID:An intronic endogenous retrovirus-like sequence attenuates human haptoglobin-related gene expression in an orientation-dependent manner. 1459 71

Highly reduced E. coli strains, MDS40, MDS41, and MDS42, lacking approximately 15% of the genome, were grown to high cell densities to test their ability to produce a recombinant protein with high yields. These strains lack all transposons and insertion sequences, cryptic prophage and many genes of unknown function. In addition to improving genetic stability, these deletions may reduce the biosynthetic requirements of the cell potentially allowing more efficient production of recombinant protein. Basic growth parameters and the ability of the strains to produce chloramphenicol acetyltransferase (CAT) under high cell density, batch cultivation were assessed. Although growth rate and recombinant protein production of the reduced genome strains are comparable to the parental MG1655 strain, the reduced genome strains were found to accumulate significant amounts of acetate in the medium at the expense of additional biomass. A number of hypotheses were examined to explain the accumulation of acetate, including oxygen limitation, carbon flux imbalance, and metabolic activity of the recombinant protein. Use of a non-catalytic CAT variant identified the recombinant protein activity as the source of this phenomenon; implications for the metabolic efficiency of the reduced genome strains are discussed.
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PMID:Expression of two recombinant chloramphenicol acetyltransferase variants in highly reduced genome Escherichia coli strains. 1749 38


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