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)

Adipose tissue and skeletal and heart muscle, which exhibit insulin-stimulated glucose uptake, express a specific, insulin-responsive glucose transporter. Previously, a cDNA (GT2) encoding this protein was isolated from a mouse 3T3-L1 adipocyte library and was sequenced. Here we report the isolation and characterization of the corresponding mouse gene designated GLUT4. The GLUT4 gene spans 7 kilobases and consists of 11 exons and 10 introns. The start site of transcription was mapped 180 nucleotides upstream of the initial methionine codon. The GLUT4 promoter contains four potential binding sites for the nuclear transcription factor Sp1 as well as a CCAAT box. DNase I footprinting of the GLUT4 promoter with nuclear extracts from undifferentiated and differentiated 3T3-L1 cells revealed that a differentiation-specific nuclear factor binds in the region at position -258 relative to the start site of transcription. Purified CCAAT/enhancer binding protein (C/EBP) was found to bind at the same position. Transient cotransfection into 3T3-L1 preadipocytes of a GLUT4 promoter-chloramphenicol acetyltransferase gene construct that contains the C/EBP binding site, together with a C/EBP expression vector, revealed that C/EBP trans-activates the GLUT4 promoter. We suggest that C/EBP plays an important role in tissue-specific, as well as metabolic, regulation of the insulin-responsive glucose transporter gene.
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PMID:Mouse insulin-responsive glucose transporter gene: characterization of the gene and trans-activation by the CCAAT/enhancer binding protein. 240 78

The NGFI-A gene encodes a "zinc-finger" protein that is rapidly induced by nerve growth factor (NGF) in PC12 rat pheochromocytoma cells. The complete exon/intron organization and nucleotide sequence of the rat NGFI-A gene have been determined. The gene spans 3789 nucleotides (nt) and is interrupted by a single intron at nt 588. All three zinc-finger DNA-binding domains are contiguously coded for within the 3' exon; this is in contrast to the structure described by others for the Xenopus laevis transcription factor TFIIIA gene. To analyze the transcription of this gene, we have determined the transcription start site and nucleotide sequence of the 5' flanking region. Transfection of PC12 cells with a fragment from the 5' flanking region linked to the chloramphenicol acetyltransferase (CAT) gene revealed that it contains an element which imparts an NGF-inducible phenotype to the normally silent CAT gene. Several regions with homologies to recognizable sequence elements are present in this fragment, including a TATA box at nt -27, serum response elements at nt -84, -106, -370, and -408, a cAMP-responsive element at nt -140, and a transcription factor Sp1-binding site at nt -286. These results establish the genomic structure of this mammalian multifinger protein and demonstrate that an NGF-responsive element lies upstream of the NGFI-A gene.
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PMID:Structure of the NGFI-A gene and detection of upstream sequences responsible for its transcriptional induction by nerve growth factor. 249 4

We have examined the control sequences for the late promoter function of simian virus 40 (SV40) in COS-1 cells which produce SV40 T antigen constitutively. Plasmids were constructed by cloning mutant late promoter segments upstream from sequences coding for the bacterial chloramphenicol acetyltransferase (CAT) gene, and were converted to "double-origin" type by inserting functional replication origin segments downstream from the CAT gene for replicative competence when necessary. The late promoter activity was determined by transient expression assay of the CAT mRNA and enzyme activity levels following DNA-mediated gene transfer into COS-1 cells. We find that the minimal replication origin and the 21-bp repeat containing T antigen and transcription factor Sp1 binding sites, respectively, are dispensable for late promoter function provided that one copy of the 72-bp repeat enhancer is present. We have mapped within the 72-bp repeat the major late promoter component in a 68-bp fragment (located between nucleotides 205 and 272), and found an overlapping 55-bp fragment (located between nucleotides 179 and 234) to have about one-fifth of the late promoter activity. Both the 68- and 55-bp fragments lack some of the core sequence elements required of the 72-bp repeat for transcriptional enhancer activity, and lack the ability to enhance the activity of the SV40 early promoter. The results suggest that the organization of functional units of the 72-bp repeat required for transcriptional enhancement of the early promoter is different from that required for late promoter function. The 21-bp repeat was found to have some late promoter activity located within the origin-distal copy in the absence of the 72-bp repeat. In association with the 21-bp repeat, the otherwise dispensable origin-proximal 22-bp of the 72-bp repeat containing activator protein AP-1 binding site augmented late promoter activity by three- to fourfold.
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PMID:Functional anatomy of the simian virus 40 late promoter. 283 21

Three overlapping genomic clones that contain the 5'-terminal portion of the human c-erbB-2 gene (ERBB2) were isolated. The promoter region was identified by nuclease S1 mapping with c-erbB-2 mRNA. Seven transcriptional start sites were identified. DNA sequence analysis showed that the promoter region contains a "TATA box" and a "CAAT box" about 30 and 80 base pairs (bp), respectively, upstream of the most downstream RNA initiation site. Two putative binding sites for transcription factor Sp1 were identified about 50 and 110 bp upstream of the CAAT box, and six GGA repeats were found between the CAAT box and the TATA box. This region had strong promoter activity when placed upstream of the bacterial chloramphenicol acetyltransferase gene and transfected into monkey CV-1 cells. These data indicate that the promoter of the human c-erbB-2 protooncogene is different from that of the protooncogene c-erbB-1 (epidermal growth factor receptor gene), which does not contain either a TATA box or a CAAT box. Comparison of the promoter sequences and activities of the two protooncogenes should be helpful in analysis of the regulatory mechanism of expression of their gene products, which are growth-factor receptors.
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PMID:Characterization of the promoter region of the human c-erbB-2 protooncogene. 288 35

Previous studies have shown that differentiation of 3T3-L1 preadipocytes leads to the activation of transcription of an unidentified gene which encodes a 4.9-kilobase (kb) mRNA. Several cDNAs that include the complete sequence of this mRNA were obtained and used to isolate and characterize the gene. Analysis of the nucleotide and amino acid sequences of both cDNA and genomic clones revealed that the gene encodes the mouse stearoyl-CoA desaturase (SCD), an enzyme known to be expressed upon differentiation of 3T3-L1 preadipocytes. The predicted amino acid sequence (355 residues) of the mouse 3T3-L1 adipocyte SCD exhibits 92% identity to that of the rat liver SCD. There is also a high degree of nucleotide sequence identity between the mouse and rat mRNAs in their unusually long approximately 3.5-kb 3'-untranslated regions. Mice fed a diet containing unsaturated triacylglycerides express SCD mRNA only in adipose tissue, whereas mice starved and refed a fat-free diet, express SCD mRNA in both liver and adipose tissue. The mouse gene for the desaturase spans approximately 15 kb and contains 6 exons and 5 introns with all intron-exon junctions conforming to the GT/AG splicing rule. As determined by S1 nuclease mapping and primer extension analysis, the transcriptional initiation site maps 152 nucleotides upstream from the initiation methionine codon. A canonical promoter "TATA" box is located 30 base pairs upstream of the Cap site. A typical "CCAAT" box sequence is not present in the adjacent 5'-flanking region; however, there is a GC-rich sequence (at nucleotide -215) similar to the binding site for the nuclear transcription factor Sp1. Upstream from the transcriptional initiation site are elements with homology (approximately 75%) to the putative fat-specific transcriptional element FSE2 and core consensus sequences for cAMP and glucocorticoid regulatory elements. A chimeric construct, containing 363 base pairs of 5'-flanking sequence and 30 nucleotides of 5'-untranslated sequence of the mouse SCD gene ligated to the bacterial chloramphenicol acetyltransferase gene, was transfected into 3T3-L1 cells. When cells were induced to differentiate into adipocytes, expression of the SCD chloramphenicol acetyltransferase gene increased approximately 63-fold, suggesting that the SCD promoter region contains elements that mediate the response to adipogenic agents which induce differentiation.
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PMID:Differentiation-induced gene expression in 3T3-L1 preadipocytes. Characterization of a differentially expressed gene encoding stearoyl-CoA desaturase. 290 62

The 5'-end of the human transforming growth factor-beta 1 gene (TGF-beta 1) was isolated from a human leukocyte genomic DNA library. Analysis of the transcriptional start sites of human TGF-beta 1 mRNAs by S1 mapping and primer extension revealed two major start sites 271 nucleotides from one another; several minor sites were also identified. DNA sequence analysis showed that the promoter region contains neither a "TATA" box nor a "CAAT" box, is very G+C rich, and contains 11 CCGCCC repeats. Seven putative binding sites for the transcription factor Sp1 were also identified. To determine the location of sites that may be important for the function of the TGF-beta 1 promoter, we joined the 5'-end of the TGF-beta 1 gene to the coding region for chloramphenicol acetyltransferase. The chimeric gene produced high levels of chloramphenicol acetyltransferase activity in transfected HT-1080, AKR-2B, and A-549 cells. Sequences responsible for both promotion and inhibition of transcription were located in the region extending from 1400 to 300 base pairs upstream of the first major TGF-beta 1 transcriptional start site. The 130-base pair fragment located between 453 and 323 base pairs upstream of this start site contains positive regulatory activity in all cells tested. A second promoter activity was identified in the region between the two major transcriptional start sites. These findings revealed a complex pattern of regulation of human TGF-beta 1 gene expression.
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PMID:Characterization of the promoter region of the human transforming growth factor-beta 1 gene. 290 28

DNA polymerase beta (beta-pol) is a housekeeping enzyme considered to be involved in DNA repair in vertebrate cells. We cloned a fragment of genomic DNA spanning the first two exons of the human beta-pol gene and approximately 11 kilobases of the flanking region. The segment just 5' of the transcription start site can direct expression of the bacterial chloramphenicol acetyltransferase (CAT) gene in HeLa cells. A sequence containing only 113 base pairs of flanking DNA has promoter activity, and various constructs containing up to 4.8 kilobases of flanking sequence are expressed at a similar level, indicating that with this assay the important regulatory elements are located within or proximal to the approximately 100-bp core promoter. S1 nuclease mapping was used to show that transcription of the transfected genes is initiated at the same position as the endogenous beta-pol gene. The region upstream of the transcription start site is G + C rich and contains neither CAAT nor TATA boxes, but does have three decanucleotide elements matching high affinity binding sites for the RNA polymerase II transcription factor Sp1. Extending 5' from position -39 and surrounded by Sp1 consensus binding elements, there is a 10-nucleotide sequence with perfect dyad symmetry, GTGACGTCAC. Similar sequences are found in a number of cellular and viral promoters, including several adenovirus promoters. Experiments to test whether the core beta-pol promoter is activated by the adenovirus early region products showed that cotransfection with an adenovirus expression plasmid strongly activates expression of the beta-pol promoter.
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PMID:Human beta-polymerase gene. Structure of the 5'-flanking region and active promoter. 318 28

Genomic DNA fragments corresponding to the promoter region of the human transferrin receptor were linked to either the full-length receptor cDNA or to the bacterial enzyme chloramphenicol acetyltransferase. These constructs were transfected into mouse and human cells, respectively. Gene expression was monitored 40-48 hours after transfection. Bal31 exonuclease was employed to produce 5' to 3' deletions of the promoter region. Deletion of DNA between -86 and -70 upstream of the receptor's mRNA start site resulted in a greater than 80% reduction in apparent promoter activity. DNA sequencing of the 150 bp upstream of the start site revealed that the promoter region contained several sequence elements more than 90% homologous to the consensus sequence for binding of the transcription factor Sp1. In addition, an 11 bp sequence identical to a segment of the enhancers of polyoma virus and adenovirus was located between -80 and -70. Internal deletions confirmed that this enhancer homologue was critical for full promoter activity. A 66 bp fragment encompassing the -80/-70 element augmented gene expression when the fragment was placed in either orientation upstream of the remainder of the transferrin receptor promoter.
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PMID:The promoter region of the human transferrin receptor gene. 338 45

Fragments of human genomic DNA corresponding to the promoter region of the gene for the transferrin receptor have been cloned upstream of the bacterial gene for chloramphenicol acetyltransferase and these constructs used to assess promoter activity following transfection into a human rhabdomyosarcoma cell line. Progressive 5' deletions as well as internal linker-substitution constructs support a critical role in gene expression of a sequence element approximately 70 bp upstream of the mRNA start site. In this region, the receptor gene was found to contain 11bp that are identical to a segment of the enhancers of polyoma virus and adenovirus. A fragment encompassing this element was shown to increase gene expression when the fragment was placed in either orientation upstream of the remainder of the transferrin receptor promoter but the same fragment did not activate an enhancer-less SV40 promoter. Removal from within the receptor promoter of three potential binding sites for the transcription factor Sp1 did not decrease the promoter's activity.
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PMID:Deletional analysis of the promoter region of the human transferrin receptor gene. 342 6

Several lines of evidence have suggested that the regulation of type I collagen gene transcription is complex and that important regulatory elements reside 5' to, and within, the first intron of the alpha 1(I) gene. We therefore sequenced a 2.3-kilobase HindIII fragment that encompasses 804 base pairs of 5' flanking sequence, the first exon, and most of the first intron of the alpha 1(I) human collagen gene. A 274-base-pair intronic sequence, flanked by Ava I sites (A274), contained a sequence identical to a high-affinity decanucleotide binding site for transcription factor Sp1 and a viral core enhancer sequence. DNase I protection experiments indicated zones of protection that corresponded to these motifs. When A274 was cloned 5' to the chloramphenicol acetyltransferase (CAT) gene, driven by an alpha 1(I) collagen promoter sequence, and expression was assessed by transfection, significant orientation-specific inhibition of CAT activity was observed. This effect was most apparent in chicken tendon fibroblasts, which modulate their level of collagen synthesis in culture. We propose that normal regulation of alpha 1(I) collagen gene transcription results from an interplay of positive and negative elements present in the promoter region and within the first intron.
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PMID:Regulatory elements in the first intron contribute to transcriptional control of the human alpha 1(I) collagen gene. 348 May 16


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