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:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In fibroblasts derived from human adipose tissue, aromatase induction is observed after exposure to 1 microM cortisol in the presence of serum or platelet-derived growth factor (PDGF). Progesterone suppresses this induction in a dose-dependent manner, 10 microM resulting in complete inhibition. A reduced cortisol concentration (0.1 microM) concomitantly reduces the progesterone concentration required for effective inhibition (10-100 nM). This effect of progesterone is specific, as neither the release of cellular enzymes nor aromatase induction by dibutyryl-cAMP, which acts independently from cortisol, are affected. However, the inhibitory effect of progesterone requires its presence throughout the induction period. Kinetic studies in intact cells reveal a reduced number of aromatase active sites upon progesterone treatment, whereas progesterone at near-physiological concentration (100 nM) does not inhibit aromatase activity in isolated microsomes. Semi-quantitative reverse transcriptase PCR analysis shows reduced amounts of aromatase mRNA in progesterone-treated cells, indicating specific inhibition of the glucocorticoid-dependent pathway of aromatase induction. The inhibitory effect of progesterone is not blocked by the anti-progestin ZK114043, excluding action via progesterone receptors and indicating competition for the glucocorticoid receptor. Progesterone must be considered a potential physiological inhibitor of glucocorticoid-dependent aromatase induction in adipose tissue. It is proposed that it is a suppressor of aromatase induction in adipose tissue in premenopausal women.
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PMID:Progesterone inhibits glucocorticoid-dependent aromatase induction in human adipose fibroblasts. 984 69

Recent research suggests that antidepressants exert their clinical action in depression via the restoration of glucocorticoid receptor (GR) function with a subsequent normalization of the altered feed-back regulation of the hypothalamic-pituitary adrenocortical (HPA) system. We, therefore, studied the effects of amitriptyline, a standard antidepressant, and of the glucocorticoid dexamethasone, which has recently been reported to possess antidepressive properties, on glucocorticoid receptor mRNA (GR-mRNA) derived from blood cells of healthy male volunteers. Whole blood samples were exposed in vitro for 24 h to amitriptyline and dexamethasone, the mRNA was extracted, transcripts of the 'house-keeping gene' glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and the GR-gene were subjected to reverse transcriptase-polymerase chain reaction (RT-PCR) and semiquantitatively determined by subsequent densitometry. In a concentration of 10 nM, amitriptyline induced a significant increase in GR-mRNA (GR/GAPDH ratio) to 186 +/- 31% of the control condition, while a concentration of 10 microM of amitriptyline resulted in an increase of GR-mRNA (GR/GAPDH ratio) to 165 +/- 36%. Dexamethasone also up-regulated blood cell GR-mRNA (GR/GAPDH ratio) levels at a concentration of 10 nM to 184 +/- 29%, whereas an incubation with 10 microM apparently resulted in toxic effects on blood cells with a decreased amount of total mRNA samples recovered. In conclusion, we here show an increase of GR-mRNA in human blood cells after treatment with amitriptyline and dexamethasone, pointing to a direct action of these substances on GR-gene expression in a human system.
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PMID:Regulation of glucocorticoid receptor-mRNA in human blood cells by amitriptyline and dexamethasone. 1040 68

A cDNA library from plasma membrane glucocorticoid receptor-enriched (mGR(++)) S- 49 mouse T lymphoma cells was screened with full-length rat intracellular GR (iGR) cDNA, BUGR-2 antibody, and PCR amplimers to portions of the mouse GR cDNA. One or two single-base substitutions resulting in amino acid changes (which do not incapacitate the receptor) were found in all but one clone: Val437 --> Gly (located in the first zinc finger), and Glu546 --> Gly (in the steroid-binding domain). Two previously unidentified exon 1 variants (1D and 1E), and two of three previously reported variants (1A, 1B) were found to be spliced onto the common exon 2. Exon 1D- and 1E-containing transcripts were confirmed by direct sequencing of amplimers from reverse transcriptase-coupled PCR. RNase protection studies revealed that one of these transcripts was expressed in mGR(++) cells only, but not in two mGR-less (mGR(--) S-49, and AtT-20 mouse pituitary) cell lines. These studies suggest that at least four promoters may be responsible for the control of GR (iGR and mGR) types in mouse lymphoma cells.
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PMID:Multiple glucocorticoid receptor transcripts in membrane glucocorticoid receptor-enriched S-49 mouse lymphoma cells. 1041 43

Renal 11beta-hydroxysteroid dehydrogenases (11beta-HSDs) are subject to modulation by various endogenous factors. 11beta-HSDs convert glucocorticoids into inactive 11-ketones and thereby determine tissue levels of active glucocorticoids and thus the extent of glucocorticoid receptor (GR) and mineralocorticoid receptor (MR) activation. As such, modulation of the activity of renal 11beta-HSDs may contribute to the cascade of regulatory events involved in renal electrolyte water handling. We investigated whether renal 11beta-HSDs are modulated by elevated circulating angiotensin II. In rats infused for 2 wk with angiotensin II (250 ng/[kg x min] subcutaneously), plasma angiotensin II, aldosterone, and corticosterone were raised 5.1-, 10.7-, and 2.3-fold, respectively, compared with control rats. Angiotensin II infusion raised corticosterone 11beta-oxidation 1.46- and 1.35-fold in renal cortical proximal and distal tubules (enriched by Percoll centrifugation), respectively, but had no effect on 11beta-HSD1 and 11beta-HSD2 mRNA levels (semiquantitative reverse transcriptase polymerase chain reaction), except for distal tubular 11beta-HSD1 mRNA, which was decreased to 50%. In vitro treatment of freshly isolated tubules with angiotensin II for 45 min prior to assessment of 11beta-HSD activity showed no direct acute effects of angiotensin II on tubular corticosterone 11beta-oxidation. The enhanced renal tubular corticosterone 11beta-oxidation in vivo may partly protect renal GR and MR from elevated plasma corticosterone on angiotensin II infusion.
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PMID:Effect of angiotensin II on rat renal cortical 11beta-hydroxysteroid dehydrogenase. 1121 53

To test the hypothesis that changes in the expression of the glucocorticoid receptor (GCR) and the beta(2)-adrenoceptor (beta(2)-AR) contribute significantly to the abnormal glucose metabolism in skeletal muscle from patients with Type II diabetes, we have examined (1) the levels of total GCR (alpha+beta isoforms), the alpha/alpha 2 isoform of GCR and beta(2)-AR mRNAs in skeletal muscle from insulin-resistant patients with Type II diabetes (n=10) and healthy controls (n=15), and (2) the effects of 8 weeks of intensive treatment on the whole-body glucose disposal rate and on total GCR, alpha/alpha 2 GCR and beta(2)-AR mRNA levels in diabetic patients. The total glucose disposal rate was measured by the euglycaemic hyperinsulinaemic (2 m-units x min(-1) x kg(-1)) clamp technique, and mRNA levels were assessed by reverse transcriptase-PCR and HPLC for separation of standard and unknown and quantification. Mean levels of total GCR and alpha/alpha 2 GCR mRNAs were increased in patients with Type II diabetes when compared with control subjects [total GCR, 2.06+/-0.30 and 1.47+/-0.10 amol/microg of total RNA respectively (P=0.09); alpha/alpha 2 GCR mRNA, 1.69+/-0.31 and 0.92+/-0.09 amol/microg of total RNA respectively (P=0.02)], whereas mRNA levels of the beta isoform of GCR (total GCR minus alpha/alpha 2 GCR) were decreased (P=0.006). beta(2)-AR mRNA levels were comparable in diabetic patients and control subjects (0.53+/-0.05 and 0.45+/-0.02 amol/microg of total RNA respectively; P=0.2). Intensive treatment for 8 weeks was associated with improved glycaemic control (P=0.019), and during the clamp a 75% (P=0.001) increase in the whole-body insulin-stimulated glucose disposal rate was demonstrated. Total GCR (P=0.005), alpha/alpha 2 GCR (P=0.005) and beta(2)-AR (P=0.03) mRNA levels all decreased significantly after intensive insulin treatment. A close correlation was found between increments in glucose uptake during intensive treatment and decrements in skeletal muscle total GCR mRNA (r=0.95, P<0.001; multiple regression analysis), and between glucose uptake and alpha/alpha 2 GCR m RNA levels (r=0.88, P<0.001; simple correlation). In conclusion, the abnormal regulation of GCR mRNA is likely to play a significant role in the insulin resistance observed in obese patients with Type II diabetes.
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PMID:Increments in insulin sensitivity during intensive treatment are closely correlated with decrements in glucocorticoid receptor mRNA in skeletal muscle from patients with Type II diabetes. 1167 59

The mechanism of peritoneal fibrosis in patients on continuous ambulatory peritoneal dialysis is poorly understood. The production of basic fibroblast growth factor (bFGF) by human peritoneal mesothelial cells cultured in high glucose medium was investigated, and the behavior of peritoneal fibroblasts, as well as the inhibitory effect of prednisolone, was assessed. Reverse transcriptase-PCR and immunocytochemistry showed the expression of glucocorticoid receptors in mesothelial cells. The semiquantitative reverse transcriptase-PCR showed that high glucose medium (4.0%) increased bFGF mRNA by 2.5-fold relative to control medium (0.1% glucose), with 83% suppression of the increase by 1 microM prednisolone. The bFGF protein level in culture supernatant was also increased by 1.5-fold in high glucose medium, with this change showing 45% suppression by 1 microM prednisolone. These effects of prednisolone were prevented by a glucocorticoid receptor antagonist (RU486) in a concentration-dependent manner. The proliferation of peritoneal fibroblasts was increased 1.9-fold by the supernatant of mesothelial cells cultured in high glucose medium, with 85% suppression by 1 microM prednisolone and suppression to 16% below basal proliferation by an anti-bFGF neutralizing antibody (10 microg/ml), whereas proliferation showed a concentration-dependent increase on addition of an anti-transforming growth factor beta-neutralizing antibody. Recombinant bFGF (50 to 1000 pg/ml) likewise caused a concentration-dependent increase of peritoneal fibroblast proliferation and fibronectin release by these cells was also increased (at 50 to 5000 pg/ml). These results suggest the potential importance of bFGF for initiation of peritoneal fibrosis and the possible efficacy of glucocorticoids for preventing such fibrosis in patients receiving peritoneal dialysis.
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PMID:Glucose and prednisolone alter basic fibroblast growth factor expression in peritoneal mesothelial cells and fibroblasts. 1172 49

The human glucocorticoid receptor isoforms GRalpha and GRbeta are generated by alternative splicing. Upon hormone binding, GRalpha regulates positively or negatively transcription. In particular, it represses numerous genes encoding pro-inflammatory mediators by inhibiting the transcription factors activator protein (AP)-1 and nuclear factor (NF)-kappaB. The observation that GRbeta, which does not bind the hormone, may act as a dominant negative receptor is subject to controversy. Because GRbeta must be more abundant than GRalpha to act as such, we evaluated the relative amounts of GRalpha and GRbeta in COS-1, A549 and HeLa cells using a monoclonal antibody that recognises the two isoforms equally well on western blots. Messenger RNA levels of GRalpha and GRbeta were compared by reverse transcriptase polymerase chain reaction analysis. To gain insight into the possible function of GRbeta, we examined the ability of overexpressed GRbeta to alter transcription of glucocorticoid, AP-1 and NF-kappaB inducible reporter genes using transient transfection in COS-1 and A549 cells. Subcellular localisation of GRbeta was determined in A549 cells by immunofluoresence microscopy. Data indicate that GRalpha is the predominant endogenous isoform in A549 and HeLa cells. GRbeta became the major form after transfection with the corresponding expression vector and translocated into cell nuclei even in the absence of hormone. Overexpression of GRbeta inhibited glucocorticoid-induced transcription markedly in COS-1 cells but weakly in A549 cells. We found that GRbeta did not act as a dominant negative modulator of GRalpha for repression of AP-1 and NF-kappaB activities. In fact, both GRbeta and GRalpha inhibited hormone-independently these activities by 25-60%. This property was not shared by the closely related mineralocorticoid receptor. Our results suggest that overexpression of either GRalpha or GRbeta may have an anti-inflammatory effect.
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PMID:Overexpression of the human glucocorticoid receptor alpha and beta isoforms inhibits AP-1 and NF-kappaB activities hormone independently. 1202 37

We have cloned a cDNA coding for a novel steroid receptor co-activator protein termed SRAP from a rat prostate library. Although the nucleotide sequence of the SRAP has 78.2% identity to that of the human steroid receptor RNA activator (SRA), a novel RNA molecule which was reported to act as an RNA transcript without being translated into protein [Lanz, McKenna, Onate, Albrecht, Wong, Tsai, Tsai and O'Malley (1999) Cell 97, 17-27], the cDNA of SRAP is capable of generating a functional protein. Glutathione S-transferase pull-down assays showed that SRAP associates with the partial androgen receptor (AR) protein composed of a DNA-binding domain and an activation function 2. Luciferase assays demonstrated that SRAP enhances the transactivation activity of the AR, the glucocorticoid receptor and the peroxisome proliferator-activated receptor gamma(1) in a ligand-dependent manner. Using a green fluorescent protein (GFP) fusion-protein construct, we demonstrated in vivo translation of the GFP-SRAP fusion protein in HeLa cells co-transfected with pSG5AR and reporter gene in the presence of 5 alpha-dihydrotestosterone (DHT). Co-transfection of the GFP-SRAP fusion protein expression plasmid enhanced the transactivation activity of AR whereas incorporation of mutations in SRAP of the fusion protein resulted in loss of enhancement of the transactivation activity. Northern blot analysis and reverse transcriptase PCR assays showed that SRAP and SRA are expressed in rat and human prostate cancer cell lines respectively. In HeLa cells and the human prostate cancer cells line DU-145, co-transfected with SRAP, the DHT-dependent transactivation activities of AR were not completely inhibited by the anti-androgen flutamide, but the transactivation activities still remained high even in the presence of 5 microM flutamide, suggesting that SRAP may play an important role in enhancing AR activity in prostate cancer.
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PMID:A novel steroid receptor co-activator protein (SRAP) as an alternative form of steroid receptor RNA-activator gene: expression in prostate cancer cells and enhancement of androgen receptor activity. 1235 Feb 25

CYP3A5 is the major CYP3A form in the human lung, and it is inducible by dexamethasone in the human A549 lung adenocarcinoma cell line. In the present study, we characterized the nature and mechanism of this induction process. The induction of CYP3A5 mRNA was assessed by quantitative reverse transcriptase-polymerase chain reaction in A549 cells. About 4-fold induction was detected by nanomolar concentrations of dexamethasone and also by budenoside and beclomethasone dipropionate, glucocorticoids used for the inhalation treatment of bronchial asthma, whereas the CYP3A4 inducers mifepristone (RU486), rifampicin, clotrimazole, and nifedipine were without effect. The glucocorticoid induction was blocked by the glucocorticoid receptor (GR) antagonist RU486. In transient transfection assays to A549 cells, CYP3A5 5' regulatory region was activated by the dexamethasone treatment. In contrast, dexamethasone was unable to induce CYP3A5 transcription in GR-deficient COS-1 cells, but the induction could be achieved after GR cotransfection. The CYP3A5 expression was measured in alveolar macrophages from patients with respiratory diseases. The CYP3A5 expression level was decreased by smoking, but glucocorticoid therapy had no statistically significant effect. In conclusion, CYP3A5 is induced in the A549 cells by glucocorticoids through a GR-mediated pathway, whereas smoking may be able to depress CYP3A5 expression.
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PMID:Regulation of CYP3A5 by glucocorticoids and cigarette smoke in human lung-derived cells. 1253 30

The effect of dexamethasone (DEX) on glucocorticoid receptor (GR)-mediated gene expression was examined in the brain of young and aged rats. Electrophoretic mobility shift assays showed that DEX treatment led to an increase of glucocorticoid response element (GRE) binding activity in aged rats, whereas in young animals GRE binding activity was decreased. Western blot analysis and reverse transcriptase polymerase chain reaction confirmed that, in aged animals, the GR mRNA and the GR protein levels were increased on DEX treatment. The binding activity of GRE activating protein-1 (AP-1) site and cross-competition analysis demonstrated specific pattern of expression during the ageing and DEX treatment, suggesting that GR modulates the activity of transcription factors AP-1 (Fos/Jun proteins) through protein-protein interaction. On the basis of these results, it can be concluded that the composition of transcriptional complexes that bind to GRE and AP-1 regulatory elements changes upon DEX treatment in an age-specific manner.
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PMID:Effects of age and dexamethasone treatment on glucocorticoid response element and activating protein-1 binding activity in rat brain. 1266 92


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