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)

A 1.5-kb genomic DNA fragment situated upstream from the quail tyrosine hydroxylase (TH) gene transcription site was isolated. This upstream region starts 15 bp from the translation initiation site. It contains two canonical TATA boxes, at positions -37 and -297, three putative glucocorticoid-responsive elements, at positions -1487, -1329, and -1268, and one putative cyclic AMP (cAMP)-responsive element (CRE) at position -53, as well as a putative negative regulatory element consensus sequence at position -735. The consensus POU-Oct site is partly conserved. Comparison of the 5' flanking sequences of quail and mammalian (bovine, human, and rat) TH genes revealed a strong sequence conservation within the 230 nucleotides upstream of the TATA box, with a distinct conservation of the CRE region. Constructs in which the bacterial chloramphenicol acetyltransferase (CAT) reporter gene was linked to promoter stretches of increasing lengths were transfected into three cell lines, two of them originating from quail and rat neural crest and the third derived from mouse fibroblasts. Reporter gene expression was specifically high in the quail and rat neural crest-derived cells compared to the fibroblast cell line. The physiological activity of this putative quail CRE was analyzed further in transfected neural crest cells of quail origin. Both cAMP analogues and agents that enhance intracellular cAMP increased CAT activity. The physiological relevance of this finding is sustained by the presence, in quail nuclear extracts, of a protein(s) that binds CRE consensus sequences.
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PMID:The quail tyrosine hydroxylase gene promoter contains an active cyclic AMP-responsive element. 809 61

The aim of the present study was to examine the intracellular localization of ectopically expressed proteins of the bacterial reporter gene in neurons of transgenic mice. In the brain of transgenic mice that carry a chimeric gene composed of the human tyrosine hydroxylase gene promoter and the bacterial gene encoding chloramphenicol acetyltransferase (CAT), the expressed CAT protein was previously found in catecholaminergic neurons as well as in non-catecholaminergic neurons expressing CAT ectopically. In the gene-expressing catecholaminergic neurons, the CAT protein was formerly detected in the axon terminals. In the present study, we immunocytochemically showed that the ectopically expressed protein was localized in cell bodies and axon terminals of the granule cells of the dentate gyrus. This suggests that the ectopically expressed protein of the bacterial gene is transported in the axons to their terminals.
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PMID:Axonal transport of protein products of the bacterial reporter gene in the brain of transgenic mice. 872 13

Transcriptional regulation of the rat tyrosine hydroxylase (TH) gene by prostaglandin E2 (PGE2) was investigated in human neuroblastoma SK-N-BE(2)C cells. Prostaglandins increased intracellular cAMP in the presence of 3-isobutyl-1-methylxanthine (IBMX), a cAMP phosphodiesterase inhibitor. Among the prostaglandins tested for their cAMP raising property PGE2 was the most effective. The results suggest that the cells express adenylyl cyclase-linked prostanoid receptors that have a higher affinity for PGE2 than for any other naturally occurring prostaglandin. The treatment of cells with PGE2 increased the TH gene expression approximately 2-fold, even though the cAMP accumulation induced by PGE2 alone was almost negligible. Simultaneous treatment with PGE2 and IBMX enhanced the gene expression concomitantly with a marked accumulation of cAMP. Transient transfection assays with 5' upstream serially deleted constructs of the rat TH gene promoter region fused to the chloramphenicol acetyltransferase (CAT) gene revealed that a cAMP response element (CRE) located at -45 to -38 from the start of the TH gene was essential for the enhancement of TH gene expression by PGE2. Site-directed mutagenesis and specific deletion within the sequence of the CRE motif abolished the transcriptional enhancement by PGE2. In addition, a protein kinase A (PKA) inhibitor, H89, specifically blocked the PGE2 effect on TH gene expression. Northern blot analysis revealed that the increase in TH gene transcription with PGE2 is associated with an elevated TH mRNA level. Gel retardation and competition assays confirmed that the binding of nuclear factors to the CRE site was sequence specific and was augmented by PGE2. Our data indicate that PGE2 enhances transcription of the TH gene mediated by the CRE motif through the activation of PKA. They also suggest that the signal flow from the adenylyl cyclase-linked prostanoid receptor to the nucleus is efficient although cAMP accumulation is not prominent.
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PMID:Transcriptional enhancement of tyrosine hydroxylase by prostaglandin E2 in SK-N-BE(2) C cells. 880 26

Tyrosine hydroxylase (TH) gene transcription rate is increased in rat adrenal medulla after administration of muscarinic agonists. In order to study this muscarinic regulation of TH gene expression in more detail, we have generated a rat pheochromocytoma PC18 cell line that stably expresses the mouse m1 muscarinic acetylcholine receptor. Treatment of this cell line, designated PC18/m1-13, with carbachol leads to rapid increases in phosphatidylinositol turnover and intracellular [Ca2+]i; these increases are totally blocked by the muscarinic antagonist atropine. Carbachol produces no changes in cAMP levels or protein kinase A activity in PC18/m1-13 cells. TH mRNA levels in PC18/m1-13 cells increase approximately 3-fold after 6 h of treatment with carbachol. This induction of TH mRNA is also completely inhibited by simultaneous treatment with atropine. Transient transfection assays using a TH gene promoter-chloramphenicol acetyltransferase (TH-CAT) construct demonstrate that sequences within the most proximal 272 bp of the TH gene 5'-flanking region are responsive to carbachol in PC18/m1-13 cells. Studies using TH-CAT constructs with site-directed mutations within the TH gene promoter indicate that the responsiveness of the promoter to carbachol is mediated primarily by the cAMP response element; however, the AP1 site also participates to a lesser extent in this response. The carbachol-mediated stimulation of TH gene promoter activity is partially inhibited by down-regulation of protein kinase C (PKC) or by treatment with the Ca2+/calmodulin-dependent protein kinase inhibitor, KN62. These results are consistent with the hypothesis that agonist occupation of m1 muscarinic receptors stimulates the TH gene via signal transduction pathways that are initiated by activation of PKC and Ca2+/calmodulin-dependent protein kinase, leading to activation of transcription factors that interact with the TH CRE and AP1 sites.
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PMID:Regulation of tyrosine hydroxylase gene expression by the m1 muscarinic acetylcholine receptor in rat pheochromocytoma cells. 884 12

Membrane depolarization of PC12 cells using 50 mM KCl leads to induction of tyrosine hydroxylase (TH) mRNA. This induction of TH mRNA is apparently due to increased TH gene promoter activity mediated by the influx of Ca2+. In PC12 cells transiently transfected with a chimeric gene expressing chloramphenicol acetyltransferase (CAT) driven by the proximal TH gene 5'-flanking region, 50 mM KCl increases TH gene promoter activity 3-4-fold. Promoter analysis utilizing TH-CAT constructs containing mutagenized sequences indicates that this response to the depolarization-mediated influx of Ca2+ is primarily dependent on both the TH cAMP-responsive element (CRE) and TH activating protein-1 (AP1) site. Minimal promoter constructs that contain a single copy of either the TH CRE or TH AP1 site fused upstream of the TH gene basal promoter are only modestly responsive or nonresponsive, respectively, to depolarization. However, both these constructs are strongly responsive to the calcium ionophore, A23187. Gel shift assays indicate that TH AP1 complex formation is dramatically increased after treatment with either 50 mM KCl or A23187. Using antibodies to transcription factors of the Fos and Jun families, we show that the nuclear proteins comprising the inducible TH AP1 complex include c-Fos, c-Jun, JunB, and JunD. In cAMP-responsive element binding protein (CREB)-deficient cell lines that express antisense RNA complementary to CREB mRNA, the response of the TH gene promoter to cyclic AMP is dramatically inhibited, but the response to A23187 remains robust. This result indicates that transcription factors other than CREB can participate in the Ca2+-mediated regulation of the TH gene. In summary, our results support the hypothesis that regulation of the TH gene by Ca2+ is mediated by mechanisms involving both the TH CRE and TH AP1 sites and that transcription factors other than or in addition to CREB participate in this response.
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PMID:Tyrosine hydroxylase gene promoter activity is regulated by both cyclic AMP-responsive element and AP1 sites following calcium influx. Evidence for cyclic amp-responsive element binding protein-independent regulation. 903 29

Prolonged stress is associated with the induction of tyrosine hydroxylase (TH) gene expression in rat adrenal medulla. We have used transgenic mice expressing a transgene encoding 4.5 kb of rat TH gene 5' flanking region fused upstream of the structural gene encoding chloramphenicol acetyltransferase (CAT) to test whether cold exposure or immobilization stress regulates TH gene expression in mouse adrenal gland. Exposure of mice to cold for 3 days increases adrenal TH protein and enzymatic activity. Cold exposure also increases adrenal TH-CAT expression two- to threefold. Immobilization stress induces mouse adrenal TH-CAT expression after either one immobilization or multiple immobilizations. TH-CAT expression increases transiently after a single immobilization, but after multiple immobilizations the induction of TH-CAT is sustained for at least 24 h. TH protein and TH enzymatic activity in mouse adrenal gland are elevated 2.8-fold and 1.5-fold, respectively, after seven immobilizations, but are not increased after either one, two, or three immobilizations. These results indicate that cold exposure and immobilization stress induce adrenal TH gene expression at least partially by stimulating the transcription rate of the TH gene. Furthermore, as observed in the rat, multiple mechanisms apparently regulate adrenal TH gene transcription rate and TH enzyme induction depending on whether mice are subjected to a single immobilization or multiple immobilizations. Our results indicate that these transgenic mice are an excellent model system to study the molecular mechanisms regulating TH gene expression in adrenal medulla.
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PMID:Induction of tyrosine hydroxylase protein and a transgene containing tyrosine hydroxylase 5' flanking sequences by stress in mouse adrenal gland. 904 52

We used a catecholaminergic neuron-like cell line (CATH.a cells) as a model system to investigate the likelihood that pituitary adenylate cyclase-activating polypeptide (PACAP) may participate in the regulation of specific gene expression in catecholaminergic neurons. Analysis by reverse transcriptase-PCR amplification revealed the presence in these cells of type I PACAP receptors, with a short isoform, together with a heavier so-called Hop splice variant. PACAP38 and PACAP27 enhanced, in a dose-dependent manner, both cyclic AMP formation and phosphoinositide breakdown, with EC50 values of, respectively, 0.6 x 10(-10) and 2 x 10(-9) M. These peptides, in addition, also elevated [Ca2+]i by mobilizing intracellular calcium pools. Vasoactive intestinal peptide (VIP) was approximately 1,000-fold less potent in stimulating cyclic AMP (with EC50 = 2 x 10(-7) M) and failed to change the turnover of phosphoinositides and to alter [Ca2+]i. Both forms of PACAP, as well as forskolin, stimulated transcriptional induction of tyrosine hydroxylase (TH) and c-fos promoters fused to a chloramphenicol acetyltransferase (CAT) reporter gene in transiently transfected cells (p < 0.01 vs. controls). Induction of CAT activity linked to both TH and c-fos promoters was obliterated upon coexpression of a dominant inhibitory mutant (Mt-RAB) of cyclic AMP-dependent protein kinase. We conclude that CATH.a cells do express functional PACAP type I receptors, the activation of which impinges on TH and c-fos transcription according to a process that is primarily dependent on the cyclic AMP-PKA pathway.
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PMID:Pituitary adenylate cyclase-activating polypeptide triggers dual transduction signaling in CATH.a cells and transcriptionally activates tyrosine hydroxylase and c-fos expression. 908 43

Basic fibroblast growth factor (FGF-2) mediates numerous important physiological processes, including differentiation and survival of dopaminergic neurons. FGF-2 was found to trigger elevation of tyrosine hydroxylase (TH) gene expression in PC12 cells that was sustained for 1-8 days. FGF-2 induced chloramphenicol acetyltransferase (CAT) reporter activity under control of the TH promoter, indicating that the induction is transcriptionally mediated. The transcriptional activation of TH by FGF-2 was examined using various deletions and point mutations of the 5' flanking region controlling CAT reporter activity. In contrast to the reported mechanisms of transcriptional regulation of TH expression by NGF and phorbol esters, the AP-1 site at -205/-199 was not required for the activation by FGF-2. A construct containing only 60 nucleotides of the promoter was still inducible by FGF-2. However, a construct with a point mutation in the CRE/CaRE was not responsive to induction by FGF-2. These findings indicate that the CRE/CaRE, but not the AP-1, element is required for induction by FGF-2 and point to differences between NGF and FGF-2 in the regulation of TH gene expression.
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PMID:Requirement for cAMP/calcium response element but not AP-1 site in fibroblast growth factor-2-elicited activation of tyrosine hydroxylase gene expression in PC12 cells. 938 81

Recently, a tyrosine hydroxylase (TH)-expressing CNS-derived cell line, CAD, was obtained that is capable of undergoing reversible morphological differentiation. The isolation of the CAD line allowed us to ask whether different DNA regulatory elements direct TH transcription when cells are growing and undifferentiated versus postmitotic and differentiated. To this end, we compared expression of a transiently transfected bacterial chloramphenicol acetyltransferase reporter gene under the transcriptional control of TH 5' flanking DNA in CAD cells grown in the presence and absence of serum. Mutational analysis indicates that CAD cells differently regulate TH transcription depending on their state of differentiation. In both states, the cyclic AMP response element and AP1 site each activate transcription. However, in undifferentiated cells, the dyad/E-box element represses expression by approximately 2.7-fold, whereas it modestly activates transcription in differentiated cells. The role of the dyad/ E-box as a repressor correlates well with the two- to threefold lower amount of endogenous TH protein present in the undifferentiated CAD cells. This study demonstrates the differential use of TH DNA regulatory elements in proliferating, undifferentiated and nonproliferating, differentiated immortalized neuronal cells.
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PMID:Differentiation of a catecholaminergic CNS cell line modifies tyrosine hydroxylase transcriptional regulation. 964 50

The stability of mRNA for tyrosine hydroxylase (TH), the rate-limiting enzyme in catecholamine synthesis, is regulated by oxygen tension in the pheochromocytoma-derived PC12 cell line. We previously identified a pyrimidine-rich 27-base-long protein-binding sequence in the 3'-untranslated region of TH mRNA that is associated with hypoxia-inducible formation of a ribonucleoprotein complex (hypoxia-inducible protein-binding site (HIPBS)). In this study, we show that HIPBS is an mRNA stabilizing element necessary for both constitutive and hypoxia-regulated stability of TH mRNA. The mutations within this sequence that abolish protein binding markedly decrease constitutive TH mRNA stability and ablate its hypoxic regulation. A short fragment of TH mRNA that contains the wild-type HIPBS confers the increased mRNA stability to the reporter chloramphenicol acetyltransferase mRNA. However, it is not sufficient to confer hypoxic regulation. The HIPBS element binds two isoforms of a 40-kDa poly(C)-binding protein (PCBP). Hypoxia induces expression of the isoform 1, PCBP1, but not the isoform 2, PCBP2, in PC12 cells.
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PMID:Regulation of tyrosine hydroxylase mRNA stability by protein-binding, pyrimidine-rich sequence in the 3'-untranslated region. 989 Oct 25


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