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)

A proinflammatory cytokine cascade, including IL-1 alpha, IL-1 beta, TNF-alpha, IL-6, and IL-8, is activated in response to infection or immunologic insult. Besides their immunologic effects, several of these mediators stimulate bone resorption and inhibit bone formation. Osteocalcin, the most abundant noncollagenous protein present in bone, is an osteoblast-specific product whose production closely correlates with bone formation, and which has also been implicated in control of bone resorption. IL-1 and TNF have previously been shown to down-regulate osteocalcin production in vitro and in vivo, although the mechanism of this inhibition is unknown. In the present studies, IL-1 beta and TNF-alpha both inhibited 1,25-dihydroxyvitamin D3-stimulated production of osteocalcin protein and mRNA by ROS 17/2.8 osteosarcoma cells, whereas IL-6 had no effect on protein and only weakly inhibited mRNA. To determine if down-regulation was exerted at the transcriptional level, an osteocalcin promoter-chloramphenicol acetyltransferase (CAT) fusion gene was constructed (PHOC-CAT). After transient transfection of PHOC-CAT into ROS 17/2.8 osteosarcoma cells, reporter CAT activity was up-regulated by vitamin D at concentrations above 10(-12) M. In screening studies, TNF-alpha (-57%) and IL-6 (-37%) inhibited vitamin D-stimulated osteocalcin transcription, whereas IL-1 alpha, IL-1 beta, and IL-8 had no effect. Other immune cytokines and growth factors, including IL-2, IL-3, IL-7, and M-CSF, also failed to regulate osteocalcin transcription. Despite their lack of promoter regulation, IL-1 alpha and IL-1 beta also stimulated PGE2 production by ROS 17/2.8, further confirming the ability of the host cell to respond to these mediators. In dose-response experiments, down-regulation by TNF-alpha was significant at concentrations as low as 0.14 pM (0.1 U/ml), whereas approximately 10(4)-fold higher concentration of IL-6 was required to exert a similar effect. TNF-alpha-mediated down-regulation was unaffected by indomethacin. These data demonstrate that of these cytokines, TNF-alpha alone potently down-regulates osteocalcin promoter function, whereas IL-1 acts post-transcriptionally, possibly by reducing mRNA stability. Heterogeneity therefore exists among the proinflammatory cytokines with respect to the level at which control of osteocalcin expression is exerted.
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PMID:Proinflammatory cytokines tumor necrosis factor-alpha and IL-6, but not IL-1, down-regulate the osteocalcin gene promoter. 130 41

We have examined the effects of several PGs on the synthesis and release of the atrial natriuretic peptide (ANP) in vivo and in vitro. PGF2 alpha infusion in anesthetized rats resulted in a significant increase in plasma immunoreactive (ir) ANP levels in vivo despite effecting only modest changes in hemodynamics. The PGs were also effective at promoting irANP secretion in primary cultures of neonatal rat atrial and ventricular cardiocytes. PGF2 alpha increased irANP release with half-maximal induction seen at approximately 10(-8) M; PGE2 was somewhat less effective and prostacyclin (PGI2) was without effect. The PGs also increased ANP mRNA levels in these cells, suggesting that these agents exert a major effect on the synthesis as well as the secretion of the prohormone. Transient expression analysis of atrial cells transfected with 2,500 bp of human (h) ANP 5' flanking sequence linked to a chloramphenicol acetyltransferase (CAT) reporter demonstrated that PGF2 alpha (10(-5) M) increased hANP promoter activity approximately twofold relative to the control. PGF2 alpha had no effect on the promoterless control (pSV0-lamin CAT). Treatment of cultured atriocytes with high concentrations of a cyclooxygenase inhibitor resulted in a significant suppression of ANP secretion in vitro and a truncation of the plasma ANP response to volume infusion in vivo. Taken together these studies support a role for PGs as regulators of cardiac ANP synthesis and secretion, and suggest an additional mechanism whereby eicosanoids may act to control cardiovascular and renal homeostasis.
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PMID:Prostaglandins regulate the synthesis and secretion of the atrial natriuretic peptide. 214 68

Human cytomegalovirus (HCMV) is a common cause of morbidity and mortality in immunosuppressed patients, especially transplant recipients. In this population, infection is frequently due to reactivation of latent virus. The major immediate early promoter of HCMV controls production of immediate early gene products, which are both trans- and cis-active and are responsible for reactivation. Activation of this promoter is therefore a crucial step in regulation of reactivation infection. It is known that there are cAMP-response elements in the HCMV major immediate early promoter. We hypothesized that prostaglandins (PG), like PGE2, which are known to increase cAMP, as well as cytokines known to be released during acute inflammation, may be important in the regulation of this promoter and thus in reactivation of HCMV. To examine this, we transfected pCAT760, a plasmid containing the major immediate early promoter of HCMV upstream of a chloramphenicol acetyltransferase (CAT) gene, into THP-1 cells. These cells were subsequently stimulated with PGE2 and/or one of a variety of cytokines. We found that PGE2, tumor necrosis factor (TNF)-alpha, and interleukin (IL)-1 beta each upregulated the HCMV major immediate early promoter. TNF-alpha, IL-1 beta, IL-6, and IL-10 were each synergistic or additive with PGE2 in upregulating the promoter. Since PGE2 and the cytokines are all products of activated macrophages, we suggest that acute inflammation and macrophage activation may predispose to reactivation of latent HCMV.
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PMID:Synergistic activation of the human cytomegalovirus major immediate early promoter by prostaglandin E2 and cytokines. 945 65

Prostaglandin E2 (PGE2) enhances transcription of the human dopamine beta-hydroxylase (DBH) gene in human neuroblastoma SK-N-BE(2)C cells. To identify a PGE2-responsive cis-acting element in the human DBH gene, serial deletion constructs of the human DBH 5'-upstream region fused to the chloramphenicol acetyltransferase (CAT) reporter gene were transiently transfected into SK-N-BE(2)C cells. Treatment of the transformed cells with PGE2 increased CAT expression two- to threefold in all constructs except where the promoter region was shortened beyond position -114 bp. There are several cis-regulatory elements in the region between -262 and -114 bp from the transcription initiation site that include a cyclic AMP response element (CRE) and a putative AP1 sequence. We presupposed that the CRE and AP1 might be candidates for PGE2 stimulation, and therefore, used site-directed mutagenesis to change the CRE and AP1 motives and test which of the two elements mediated the transcriptional enhancement. Only a specific mutation within the CRE sequence abolished the PGE2 effect. In addition, cotransfection with an expression vector expressing PKA inhibitor resulted in the specific blockage of the PGE2 effect on DBH gene expression. Northern blot analysis revealed that the increase in DBH gene transcription caused by PGE2 results in elevated DBH mRNA levels. Gel-retardation and competition assays confirmed that the binding of nuclear factors to the CRE site is sequence specific. Our data, therefore, indicate that PGE2 enhances the transcription of the human DBH gene. The effect is mediated by the CRE motif through activation of PKA.
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PMID:Stimulation of human DBH gene expression by prostaglandin E2 in human neuroblastoma SK-N-BE(2)C cells. 948 16

This report examines the effect of polyunsaturated fatty acids (PUFA) on lipogenic gene expression in cultured 3T3-L1 adipocytes. Arachidonic acid (20:4, n-6) and eicosapentaenoic acid (20:5, n-3) suppressed mRNAs encoding fatty acid synthase (FAS) and S14, but had no effect on beta-actin. Using a clonal adipocyte cell line containing a stably integrated S14CAT fusion gene, oleic acid (18:1, n-9), arachidonic acid (20:4, n-6) and eicosapentaenoic acid (20:5, n-3) inhibited chloramphenicol acetyltransferase (CAT) activity with an ED50 of 800, 50, and 400 microM, respectively. Given the high potency of 20:4, n-6, its effect on adipocyte gene expression was characterized. Arachidonic acid suppressed basal CAT activity, but did not affect glucocorticoid-mediated induction of S14CAT expression. The effect of 20:4, n-6 on S14CAT expression was blocked by an inhibitor of cyclooxygenase implicating involvement of prostanoids. Prostaglandins (PGE2 and PGF2alpha at 10 microM) inhibited CAT activity through a pertussis toxin-sensitive Gi/Go-coupled signalling cascade. Our results suggest that 20:4, n-6 inhibits lipogenic gene expression in 3T3-L1 adipocytes through a prostanoid pathway. This mechanism of control differs from the polyunsaturated fatty acid-mediated suppression of hepatic lipogenic gene expression.
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PMID:Arachidonic acid inhibits lipogenic gene expression in 3T3-L1 adipocytes through a prostanoid pathway. 968 35

The mechanism by which the proto-oncogene, c-fos, is up-regulated in response to PGE2 in the mouse osteoblastic (MC3T3-E1) cell line was investigated using RT-PCR. c-fos messenger RNA up-regulation by dmPGE2 is rapid, starting 10 min post stimulation, and transient. The specific protein kinase A (PKA) inhibitor, H89, inhibited c-fos induction. Moreover, down-regulation of protein kinase C (PKC) activity by chronic TPA treatment had no effect on the induction of c-fos by dmPGE2. We conclude that up-regulation of c-fos by dmPGE2 is primarily dependent on PKA in MC3T3-E1 osteoblasts. In S49 lymphoma wild-type but not S49 cyc- cells, which are deficient in cAMP signaling, dmPGE2 up-regulates c-fos and increases cell growth compared with unstimulated cells. Thus in S49 lymphoma cells, c-fos induction by PGE2 is also dependent on cAMP signaling. The minimal c-fos promoter region required for dmPGE2-induced expression was identified by transfecting c-fos promoter deletion constructs coupled to the chloramphenicol acetyltransferase (CAT) reporter gene into Vero cells. Transfection of a plasmid containing 99 bp c-fos proximal promoter was sufficient to direct c-fos/CAT expression following stimulation with dmPGE2. Because induction of c-fos is mediated by cAMP, these data are consistent with activation of c-fos via the CRE/ATF cis element.
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PMID:Prostaglandin E2-induced up-regulation of c-fos messenger ribonucleic acid is primarily mediated by 3',5'-cyclic adenosine monophosphate in MC3T3-E1 osteoblasts. 1061 50

In congenital heart disease with left- or right-sided obstruction, prostaglandin E (PGE)1 or PGE2 is infused to maintain ductus arteriosus (DA) patency. We hypothesized that transfection of the DA with PGE synthase would lead to a greater production of PGE2 in situ and, hence, patency of the DA. The cDNA for human prostaglandin synthase was sequenced and ligated into a eukaryotic expression vector. The negative control was created by ligating the cDNA encoding the bacterial protein chloramphenicol acetyltransferase into the same plasmid. Transfection (600 microg DNA) was achieved in lambs within the first 24 h of life using the hemagglutinating virus of Japan (HVJ)-liposome transfection method with a custom-made, basket-weave-perforated catheter. Echocardiography was performed to assess DA patency until the time of sacrifice. To confirm expression of the transgene, PGE2 concentration was measured in organ culture of the DA by immunoassay and by Western immunoblotting of homogenized DA tissue. Patency of the DA was demonstrated by color Doppler in all the lambs (7/7) in which the PGE synthase was delivered, whereas functional closure was seen in the control group (6/6). The PGE2 concentration in the culture medium of the explanted DA in the treatment group was 3-fold higher than that of the control groups. Western immunoblotting confirmed the presence of PGE synthase in the treatment group. Gene transfer of PGE synthase to the DA is feasible and will maintain patency for at least 1 wk.
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PMID:Gene transfer of prostaglandin synthase maintains patency of the newborn lamb arterial duct. 1618 5