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 mouse myosin light-chain 1A (MLC1A) gene, expressed in the atria of the adult heart, is one of the first muscle genes to be activated when skeletal as well as cardiac muscles form in the embryo. It is also transcribed in skeletal muscle cell lines at the onset of differentiation. Transient transfection assays of mouse skeletal muscle cell lines with DNA constructs containing MLC1A promoter fragments fused to the chloramphenicol acetyltransferase (CAT) gene show that the first 630 bp of the promoter is sufficient to direct expression of the reporter gene during myotube formation. Two E boxes located at bp -76 and -519 are necessary for this regulation. MyoD and myogenin proteins bind to them as heterodimers with E12 protein and, moreover, transactivate them in cotransfection experiments with the MLC1A promoter in nonmuscle cells. Interestingly, the effect of mutating each E box is less striking in primary cultures than in the C2 or Sol8 muscle cell line. A DNA fragment from bp -36 to -597 confers tissue- and stage-specific activity to the herpes simplex virus thymidine kinase promoter in both orientations, showing that the skeletal muscle-specific regulation of the MLC1A gene is under the control of a muscle-specific enhancer which extends into the proximal promoter region. At bp -89 is a diverged CArG box, CC(A/T)6AG, which binds the serum response factor (SRF) in myotube nuclear extracts, as does the wild-type sequence, CC(A/T)6GG. Both types of CArG box also bind a novel myotube-enriched complex which has contact points with the AT-rich part of the CArG box and adjacent 3' nucleotides. Mutations within the CArG box distinguish between the binding of this complex and binding of SRF; only SRF binding is directly involved in the specific regulation of the MLC1A gene in skeletal muscle cell lines.
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PMID:A skeletal muscle-specific enhancer regulated by factors binding to E and CArG boxes is present in the promoter of the mouse myosin light-chain 1A gene. 762 50

Troponin I (TnI) is a muscle-specific protein involved in the calcium-mediated contraction of striated muscle. Three TnI isoforms have been identified, each encoded by a separate gene and expressed in specific striated muscles in the adult. The slow isoform gene (TnIs) is transcriptionally regulated during skeletal muscle development such that its expression in the adult is restricted to muscle fibers innervated by a slow nerve. To delineate regions of this gene that are responsive to information imparted by the slow nerve, we generated transgenic mice carrying -4,200 to +12 bp of the human TnIs gene linked to the bacterial chloramphenicol acetyltransferase (CAT) coding region. By Northern blot analysis, we detected transgene transcripts only in muscles containing slow-twitch fibers. CAT histochemical analysis revealed that expression of the transgene is restricted solely to slow-twitch fibers as characterized by type I myosin heavy-chain (MyHC) expression. Using regeneration as a model for neural influenced expression, we show that this gene construct also contains sequences necessary to respond to cues from the central nervous system.
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PMID:The human troponin I slow promoter directs slow fiber-specific expression in transgenic mice. 762 19

DNA-binding proteins containing the basic helix-loop-helix (bHLH) domain have been implicated in lineage determination and the regulation of specific gene expression in a number of cell types. By oligonucleotide screening of an adipocyte cDNA expression library, we have identified a novel member of the bHLH-leucine zipper transcription factor family designated ADD1. ADD1 mRNA is expressed predominantly in brown adipose tissue in vivo and is regulated during both determination and differentiation of cultured adipocyte cell lines. ADD1 can function as a sequence-specific transcriptional activator in that it stimulates expression of a chloramphenicol acetyltransferase vector containing multiple ADD1 binding sequences but is unable to activate the myosin light-chain enhancer, which contains multiple binding sites for another bHLH factor, MyoD. ADD1 can also activate transcription through a binding site present in the 5'-flanking region of the fatty acid synthetase gene which is expressed in a differentiation-dependent manner in adipose cells. These data suggest that ADD1 plays a role in the regulation of determination- and differentiation-specific gene expression in adipocytes.
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PMID:ADD1: a novel helix-loop-helix transcription factor associated with adipocyte determination and differentiation. 833 13

We have studied gene activation via CArG boxes in the context of myogenesis. The proximal CArG box of the human cardiac actin gene (HCA1) stimulates transcription from the herpes simplex virus thymidine kinase (TK) promoter in a tissue-specific fashion. Thus in transient transfection assays, when the expression of chloramphenicol acetyltransferase (CAT) from p(HCA1)4 TKCAT is compared to that derived from p(M1)4 TKCAT which contains an inactive mutated version (M1) of the HCA1 element, high levels of expression are seen in C2 mouse myoblasts and myotubes, and in the T4 myoblast cell line derived from the C3H10T1/2 cell line by 5-azacytidine treatment, whereas only low levels of expression are seen in the mouse L fibroblast cell line. The parental C3H10T1/2 cell line shows intermediate levels of expression. A similar situation is seen in stably transfected cell lines. Gene activation via CArG boxes was also analyzed in the course of myogenic conversion of C3H10T1/2 cells treated with 5-azacytidine. Our results indicate that activation of the CAT gene from the HCA1 element is slightly posterior to the appearance of the first MyoD1 and myogenin transcripts, concomitant with the appearance of cardiac alpha-actin transcripts, but clearly precedes the accumulation of myosin light-chain 1a transcripts and the appearance of troponin T-positive cells. These results further establish that CArG boxes can be seen as muscle-specific cis-acting regulatory element prior to terminal differentiation.
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PMID:Activation of gene expression via CArG boxes during myogenic differentiation. 845 94

The DNA regulatory element(s) involved in beta-myosin heavy chain (beta-MHC) induction by the physiological stimulus of mechanical overload have not been identified as yet. To delineate regulatory sequences that are required for mechanical overload induction of the beta-MHC gene, transgenic mouse lines were generated that harbor transgenes containing serial deletions of the human beta-MHC promoter to nucleotides -293 (beta 293), -201 (beta 201), and -141 (beta 141) from the transcription start site (+1). Mechanically overloaded adult plantaris and soleus muscles contained 11- and 1.9-fold increases, respectively, in endogenous beta-MHC-specific mRNA transcripts (Northern blot) compared with sham-operated controls. Expression assays (chloramphenicol acetyltransferase specific activity) revealed that only transgene beta 293 expression was muscle specific in both fetal and adult mice and was induced in the plantaris (10- to 27-fold) and soleus (2- to 2.5-fold) muscles by mechanical overload. Histochemical staining for myosin adenosinetriphosphatase activity revealed a fiber-type transition of type II to type I in the overloaded plantaris and soleus muscles. These transgenic data suggest that sequences located between nucleotides -293 and +120 may be sufficient to regulate the endogenous beta-MHC gene in response to developmental signals and to the physiological signals generated by mechanical overload in fast- and slow-twitch muscles.
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PMID:Muscle-specific and inducible expression of 293-base pair beta-myosin heavy chain promoter in transgenic mice. 885 92

While the TR4 orphan receptor (TR4) is able to repress the expression of its target genes via its interaction with the direct repeat 1-hormone response element (DR1-HRE) and DR2-HRE, we now report that TR4 can also induce the transcriptional activity of the reporter gene containing a DR4-HRE via chloramphenicol acetyltransferase assay. Electrophoretic mobility shift assay and Scatchard analysis reveal a strong binding affinity (dissociation constant = 2 nM) between TR4 and DR4-HRE. The induction mediated by TR4 was detected not only in the synthetic DR4-HRE but also in some genes, such as rat alpha-myosin heavy-chain and S14 genes, containing the DR4 or DR4-like motif, which have been suggested to be the response elements for a thyroid hormone receptor. Our data also demonstrate this TR4-mediated gene induction is TR4 dose- and DR4 sequence-dependent. Together, our data suggest that DR4-HRE can be a positive regulatory element for TR4, which may be able to induce the transcriptional activity of the genes containing such positive HREs.
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PMID:Identification of direct repeat 4 as a positive regulatory element for the human TR4 orphan receptor. A modulator for the thyroid hormone target genes. 911 96

Using nuclei isolated from neonatal cardiomyocytes, we have mapped the DNase I hypersensitive sites (DHSs) residing within the 5'-upstream regions of the hamster cardiac myosin heavy-chain (MyHC) gene. Two cardiac-specific DHSs within the 5 kb upstream region of the cardiac MyHC gene were identified. One of the DHSs was mapped to the -2.3 kb (beta-2.3 kb) region and the other to the proximal promoter region. We further localized the beta-2.3 kb site to a range of 250 bp. Multiple, conserved, muscle regulatory motifs were found within the beta-2.3 kb site, consisting of three E-boxes, one AP-2 site, one CArG motif, one CT/ACCC box and one myocyte-specific enhancer factor-2 site. This cluster of regulatory elements is strikingly similar to a cluster found in the enhancer of the mouse muscle creatine kinase gene (-1256 to -1050). The specific interaction of the motifs within the beta-2.3 kb site and the cardiac nuclear proteins was demonstrated using gel mobility-shift assays and footprinting analysis. In addition, transfection analysis revealed a significant increase in chloramphenicol acetyltransferase activity when the beta-2.3 kb site was linked to a heterologous promoter. These results suggest that previously undefined regulatory elements of the beta-MyHC gene may be associated with the beta-2.3 kb site.
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PMID:Multiple muscle-specific regulatory elements are associated with a DNase I hypersensitive site of the cardiac beta-myosin heavy-chain gene. 935 23

In vivo, vascular smooth muscle (VSM) cells change their contractile phenotype toward a more proliferative phenotype during the pathogenesis of vascular diseases. Because these dedifferentiated VSM cells may gradually regain contractile functions, we aimed to identify signaling pathways that result in an increased expression of contractile proteins in VSM cells. In vitro, serum and thrombin induced a reversible upregulation of smooth muscle myosin heavy-chain (SM-MHC) in cultured neonatal rat VSM cells. Cotransfection of a SM-MHC-promoter chloramphenicol acetyltransferase-construct with dominant-negative N17Ras or N17Raf or treatment with the mitogen-activated/ERK-activating kinase (MEK) inhibitor PD 98059 concentration dependently decreased the serum- or thrombin-induced SM-MHC promoter activity. Consistently, the serum- or thrombin-induced phosphorylation of MEK and extracellular signal-regulated kinase 1/2 (ERK1/2) coincided with a MEK-dependent nuclear accumulation of phosphorylated ERK1/2 and subsequent nuclear phosphorylation of the transcription factors c-myc and Elk-1. A 5'-deletion analysis of cis-elements within the SM-MHC promoter demonstrated that a conserved region (nucleotide -1346 to -1102) was required for both cell type-specific expression and serum- or thrombin-induced upregulation of the SM-MHC promoter in VSM cells. Within this region, 2 CArG-boxes, a GC-rich element, and a CTF/NF-1 site are critical positively acting cis-elements for the serum- or thrombin-induced upregulation of SM-MHC. We conclude that the serum- or thrombin-induced differentiation requires an intact Ras/Raf/MEK/ERK signaling cascade, nuclear translocation of activated ERK1/2, phosphorylation of transcription factors, and several cis-elements within the SM-MHC promoter.
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PMID:ERK1/2-dependent contractile protein expression in vascular smooth muscle cells. 1262 57


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