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)

Oncogenic activation of ras results in changes in the transcription of several genes leading to uncontrolled cell growth. In this paper, we demonstrate that transformation of fibroblast cells by the ras oncogene leads to transcriptional repression of the smooth muscle alpha-actin promoter. Transient transfection analysis of plasmids containing the 5' upstream region of the human alpha-actin gene fused to human growth hormone or bacterial chloramphenicol acetyltransferase coding sequences into Rat-2 and ras-transformed Rat-2 (HO6) cells indicates that alpha-actin promoter is repressed in ras-transformed cells. In addition, stable rat fibroblast cell lines expressing human growth hormone or beta-galactosidase under the control of alpha-actin promoter exhibit repressed reporter gene activity following transformation by the ras oncogene. alpha-Actin promoter-driven beta-galactosidase activity is derepressed in revertants of ras-transformed stable cell lines. This revertant cell line expresses elevated levels of ras p21 protein and is resistant to retransformation by Ki and Ha-ras oncogenes. The revertant may have either a defective target protein whose activity is essential for the transforming activity of ras or an activated tumor suppressor gene which can suppress the activity of ras. These results indicate that smooth muscle alpha-actin promoter activity is a sensitive marker to follow phenotypic changes following transformation by ras and subsequent reversion. The advantages of this alpha-actin promoter-reporter gene assay system to screen for drugs that inhibit the transforming activity of ras, either directly or indirectly, are discussed.
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PMID:Regulation of smooth muscle alpha-actin promoter in ras-transformed cells: usefulness for setting up reporter gene-based assay system for drug screening. 145 76

To assess the role of cis-acting elements within the smooth muscle alpha-actin gene in smooth muscle cells (SMC), we transfected chicken smooth muscle alpha-actin promoter-chloramphenicol acetyltransferase gene fusion plasmids into SMC derived from rat and chicken aortas. In marked contrast to effects in chicken skeletal myoblasts and fibroblasts, p122CAT (positions -122 to +19), containing two conserved CArG elements, elicited a modest increase in chloramphenicol acetyltransferase reporter activity in chicken SMC. Addition of upstream sequences between -122 and -151 (p151CAT) increased activity in adult chicken SMC. Addition of sequence between positions -151 and -257 (p257CAT) resulted in a 7-fold increase in chloramphenicol acetyltransferase activity over that of p151CAT in rat SMC, but not in chicken SMC. A genomic clone encoding the rat smooth muscle alpha-actin gene was isolated, and the 5'-flanking region was partially characterized. Comparison of primary sequence between rat and chicken promoters showed a conserved E box motif at position -214 in the chicken gene and at position -213 in the rat gene. Results of these studies demonstrate that regions upstream of the conserved CArG elements exert potent regulatory effects on transcription and that SMC require different cis-acting elements than other cell types to transcriptionally regulate this gene.
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PMID:Elements of the smooth muscle alpha-actin promoter required in cis for transcriptional activation in smooth muscle. Evidence for cell type-specific regulation. 173 Jun 87

Vasoconstrictors such as arginine vasopressin (AVP) and angiotensin II (Ang II) have been shown to increase protein and mRNA levels of smooth muscle alpha-actin (SM-alpha-actin) in vascular smooth muscle cells. In the same cells, platelet-derived growth factor (PDGF) decreased SM-alpha-actin protein and mRNA. The rat SM-alpha-actin promoter that has recently been isolated contains two E-boxes and three CC(A/T)6GG (CArG) elements. To examine regulation of the SM-alpha-actin promoter, a 765-bp region of the rat SM-alpha-actin gene was ligated into chloramphenicol acetyltransferase (CAT)-containing vectors and transfected into rat aortic vascular smooth muscle cells. Stimulation of cells with either AVP or Ang II increased CAT activity 5- to 10-fold. PDGF was able to completely block the AVP-induced increase in CAT activity. To identify regions of the promoter responsible for both the AVP stimulation and PDGF inhibition of promoter activity, a series of truncation mutants were prepared and transfected into vascular smooth muscle cells. Truncation of both E-boxes and the most distal CArG element did not qualitatively alter either AVP-induced stimulation of CAT activity or PDGF inhibition. However, removal of the middle CArG element resulted in a loss of AVP stimulation. These studies indicate that the AVP-induced elevation and PDGF-induced inhibition of SM-alpha-actin levels in vascular smooth muscle cells are mediated at least in part through regulation of the SM-alpha-actin promoter. The critical region of the promoter mediating this effect involves at a minimum one of the CArG elements.
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PMID:Regulation of smooth muscle alpha-actin promoter by vasopressin and platelet-derived growth factor in rat aortic vascular smooth muscle cells. 795 49

Advanced glycation end products (AGEs) stimulate synthesis of extracellular matrix (ECM) in a receptor-mediated manner on mesangial cells. In the present study, we examined the transcriptional regulation of the gene for type IV collagen [(IV)collagen], which is one of the major components of mesangial sclerosis, after stimulation of AGEs on mesangial cells. The methylation pattern of the promoter/enhancer region of (IV)collagen gene was similar in AGE-treated and control cells. AGEs significantly increased the transcriptional activity of the (IV)collagen gene, as measured by transient transfection assays using the reporter gene construct containing (IV)collagen promoter/enhancer and the chloramphenicol acetyltransferase gene. AGEs also increased smooth muscle alpha-actin mRNA levels as well as its transcriptional activity. Nuclear factor binding of the promoter of (IV)collagen gene was stimulated by AGEs. Furthermore, AGEs dramatically decreased the mRNA levels of (IV)collagen promoter binding protein (MSW), a larger subunit of DNA replication complex, AP1. These results suggest that AGEs increase expression of (IV)collagen gene by modulating the levels of promoter binding proteins. These transcriptional events may play a critical role in ECM accumulation in response to AGEs.
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PMID:Advanced glycation end products modulate transcriptional regulation in mesangial cells. 888 74

E-box/basic helix-loop-helix (bHLH)-dependent regulation of promoters for skeletal muscle-specific genes is well established, but similar regulation of smooth muscle-selective promoters has not been reported. Using transient transfection assays of smooth muscle alpha-actin (SMalphaA) promoter-chloramphenicol acetyltransferase (CAT) reporter constructs in rat vascular smooth muscle cells (SMCs) and L6 skeletal myotubes, we identified two activator elements, smE1 and smE2, with sequences corresponding to E-box (5'-CAnnTG-3') motifs. In L6 myotubes, 4-bp mutations of smE1 or smE2 E-box motif alone completely abolished promoter activity. In contrast, mutation of smE1 and smE2 was required to reduce promoter activity in SMCs. Supershift analyses identified a myogenin-containing complex as the predominant smE1 and smE2 binding activity in skeletal muscle, and myogenin overexpression transactivated the promoter. Supershift analyses with SMC extracts demonstrated that the bHLH protein upstream stimulatory factor (USF) bound smE1, and USF overexpression transactivated the promoter in an smE1-dependent manner. In summary, our results provide novel evidence implicating E-box elements in directing expression of the SMalphaA promoter through distinct bHLH factor complexes in skeletal vs. smooth muscle.
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PMID:Differential activation of the SMalphaA promoter in smooth vs. skeletal muscle cells by bHLH factors. 1036 6