Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.3.1.28 (chloramphenicol acetyltransferase)
 5,100 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have studied the 5' upstream sequences required for the transcriptional regulation of the hamster gene encoding the intermediate filament protein, vimentin. Although vimentin is regarded as the intermediate filament protein of mesothelial tissue, it is also produced in most cultured cells. The human mammary carcinoma cell line, MCF-7, belongs to the exceptions. It contains no vimentin, and the complete upstream promoter region is inactive in this particular cell line. By using transient transfection of chimeric constructs into MCF-7 and HeLa cells, and subsequent chloramphenicol acetyltransferase assays, we were able to show the presence of two negative control regions flanking a double AP-1 enhancer element. Our data indicate that these elements exert their effect irrespective of orientation and position, suggesting that they are silencers. In vitro footprinting assays, gel mobility assays and Southwestern (protein-DNA) blotting revealed the presence of trans-acting factors interacting with both silencer elements. The silencing effect was particularly pronounced in MCF-7 cells, although DNA-binding proteins are present in HeLa cells as well.
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PMID:Identification of two silencers flanking an AP-1 enhancer in the vimentin promoter. 148 48

The peripherin gene, which encodes a neuronal-specific intermediate filament protein, is transcriptionally induced with a late time course when nerve growth factor (NGF) stimulates PC12 cells to differentiate into neurons. We have studied its transcriptional regulation in order to better understand the neuronal-specific end steps of the signal transduction pathway of NGF. By 5' deletion mapping of the peripherin promoter, we have localized two positive regulatory elements necessary for full induction by NGF: a distal positive element and a proximal constitutive element within 111 bp of the transcriptional start site. In addition, there is a negative regulatory element (NRE; -179 to -111), the deletion of which results in elevated basal expression of the gene. Methylation interference footprinting of the NRE defined a unique sequence, GGCAGGGCGCC, as the binding site for proteins present in nuclear extracts from both undifferentiated and differentiated PC12 cells. However, DNA mobility shift assays using an oligonucleotide probe containing the footprinted sequence demonstrate a prominent retarded complex in extracts from undifferentiated PC12 cells which migrates with slower mobility than do the complexes produced by using differentiated PC12 cell extract. Transfection experiments using peripherin-chloramphenicol acetyltransferase constructs in which the footprinted sequence has been mutated confirm that the NRE has a functional, though not exclusive, role in repressing peripherin expression in undifferentiated and nonneuronal cells. We propose a two-step model of activation of peripherin by NGF in which dissociation of a repressor from the protein complex at the NRE, coupled with a positive signal from the distal positive element, results in depression of the gene.
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PMID:Nerve growth factor-induced derepression of peripherin gene expression is associated with alterations in proteins binding to a negative regulatory element. 158 54

The gfa gene encodes glial fibrillary acidic protein, an intermediate filament protein found almost exclusively in astrocytes. Transient transfection studies with a chloramphenicol acetyltransferase reporter gene were used to identify regions of the gfa gene responsible for its expression. Three regions, A, B, and D, were found to be important. The D region is located near the basal promoter, while A and B are next to each other about 1500 bp further upstream. The regions contain several sequences homologous to binding sites of known transcription factors, and in addition, each contains an identical novel 10-bp motif. The A, B, and D regions act in a cell-specific manner; when joined to the SV 40 early promoter, they enhance transcription in the glial cell line U251, but not in the nonglial cell line HepG2. Consistent with this observation, the DNase I footprint produced in these regions by nuclear extract from U251 cells differs from that produced by an extract from HepG2 cells. The B region appears to be the most active of the three, as by itself it stimulates strong cell-specific transcription, whereas addition of the other two regions has little effect. When the B region is at its normal distance from the basal promoter, deletion of D severely reduces transcription, but when B is placed near the promoter, D is unimportant. This suggests that the D region may function primarily to promote interactions that bring B close to the promoter.
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PMID:Multiple interacting sites regulate astrocyte-specific transcription of the human gene for glial fibrillary acidic protein. 191 4

To understand astrocyte-specific transcription, we have been studying the human gfa gene. This gene encodes glial fibrillary acidic protein (GFAP), an intermediate filament protein expressed primarily in astrocytes. A survey of the gfa 5' flanking region showed it to contain several segments that contribute to expression of a chloramphenicol acetyltransferase reporter gene in transfected cells. The most active of these was the 124-bp B region, which spans bp -1612 to -1489. We have now used site-directed mutagenesis to analyze this region in greater detail, and show that the B region itself contains several important elements. The most crucial of these is a consensus AP-1 sequence, the binding site for the Fos and Jun families of transcription factors. The presence of members of both these families in the glial fibrillary acidic protein-expressing U251 cell line used for our transfection studies was verified by gel mobility-shift experiments. This is the first demonstration of the functioning of a specific transcription factor site for astrocytes, and provides a focus for future studies of glial fibrillary acidic protein regulation during development and reactive gliosis.
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PMID:Analysis of a segment of the human glial fibrillary acidic protein gene that directs astrocyte-specific transcription. 851 62