Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.6 (RNA polymerase)
 34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The biological role of platelet-derived growth factor (PDGF)-AA in lung morphogenesis was investigated by incubating embryonic lung explants with phosphorothioate antisense PDGF-A oligonucleotides, which decreased PDGF-AA but not PDGF-BB protein content. Antisense PDGF-A oligonucleotides inhibited DNA synthesis. This inhibitory effect of antisense PDGF-A was reversed by the addition of exogenous PDGF-AA but not PDGF-BB. Morphometric analyses of antisense-treated cultures showed a significant reduction in lung size. The number of terminal buds of the lung explants was significantly decreased by antisense PDGF-A oligonucleotides. PDGF-AA but not PDGF-BB attenuated the inhibitory effect of antisense PDGF-A on early lung branching. Sense PDGF-A had no effect on DNA synthesis and early lung branching. Reverse transcriptase-polymerase chain reaction analysis revealed PDGF-A mRNA expression in the epithelial component of the embryonic lung, while message for PDGF alpha-receptor was expressed in the mesenchyme. Incubation of explants with neutralizing PDGF-AA antibodies also reduced DNA synthesis and early branching morphogenesis. We conclude that PDGF-AA and its receptor represent an important epithelial-mesenchymal interaction which plays a critical role in early lung branching morphogenesis.
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PMID:PDGF-AA and its receptor influence early lung branching via an epithelial-mesenchymal interaction. 767 19

Collagen deposition and myofibroblast proliferation beneath the epithelial basement membrane in patients with asthma is now increasingly recognized, although the molecular pathogenesis remains obscure. We have evaluated messenger ribonucleic acid (mRNA) expression of the profibrotic cytokine, platelet-derived growth factor-beta (PDGF-beta), in alveolar macrophages obtained following fibreoptic bronchoscopy and bronchoalveolar lavage in patients with asthma. Three subject groups were studied: 1) asthmatics using regular inhaled glucocorticoid medication (ASTST, n = 9), 2) asthmatics using intermittent inhaled beta 2-agonist therapy only (ASTBR, n = 10); 3) nonasthmatic control volunteers (n = 10). Alveolar macrophage mRNA was extracted and PDGF-beta mRNA quantified by reverse-transcriptase polymerase chain reaction (PCR) (RT-PCR) and expressed as the ratio to that of a control gene glyceraldehyde-3-phosphate dehydrogenase (GAPDH). There were no significant differences in PDGF-beta mRNA expression between the groups, or between all asthmatic (n = 19) and control subjects. Furthermore, there was no correlation between alveolar macrophage PDGF-beta mRNA expression and airway spirometry, or duration of glucocorticoid usage or dose. Thus, in contrast to other fibrotic lung diseases, we found little evidence of enhanced expression of PDGF-beta mRNA in alveolar macrophages in clinically stable bronchial asthma.
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PMID:Platelet-derived growth factor-beta mRNA in human alveolar macrophages in vivo in asthma. 787 66

We have investigated the effect of phorbol 12-myristate 13-acetate (PMA) on platelet-derived growth factor (PDGF) B-chain gene transcription as well as on mRNA stability in cultured human mesangial cells. Addition of actinomycin to cells stimulated with PMA decreases steady state levels of PDGF-B chain mRNA analysed by solution hybridization assay. PDGF-B chain gene transcription was also assayed directly by measuring elongation of transcripts in isolated nuclei followed by hybridization of labeled RNA transcripts to a cDNA encoding for PDGF-B chain. Our data show that PMA induces PDGF-B chain gene transcription by approximately 2-fold. alpha-Amanitin, an RNA polymerase II inhibitor, blocked transcription by more than 70%. In addition, we determined the effect of PMA on the halflife of PDGF-B chain mRNA directly by pulse chase method. In human mesangial cells, the PDGF-B chain mRNA exhibited halflife of approximately 105 min. In the presence of PMA, the halflife of PDGF-B chain mRNA was reduced to approximately 72 min. These studies indicate that regulation of PDGF-B chain gene by PMA in human mesangial cells involves a coordinate effort at the level of transcription and mRNA stability.
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PMID:Platelet-derived growth factor B-chain gene expression in mesangial cells: effect of phorbol ester on gene transcription and mRNA stability. 787 95

We report expression of the wt1 (Wilms' tumor) gene by cultured human melanoma cells. Using RNA polymerase chain reaction analysis, wt1 transcripts were detected in 7 of 9 melanoma cell lines but not in 5 normal melanocyte strains. In Northern blot analysis, steady-state wt1 mRNA levels were found in 2 of 4 melanoma lines but not in normal melanocytes. Sequence analysis of the wt1 cDNA expressed by melanoma cell line WM 902-B revealed the presence of 4 previously published splice variants but no evidence for mutations in the coding region. Previous work has shown that WT1 modulates transcription after binding to the early growth response (EGR)-1 sites present in the platelet-derived growth factor (PDGF)-A chain promoter; the PDGF-A chain gene is known to be expressed by various melanoma cell lines. Based on these findings, we studied the relationship of wt1 and PDGF-A chain gene expression in melanoma cell lines. Co-expression of the wt1 and the PDGF-A chain genes was observed in 2 melanoma cell lines with mutated p53 but not in 2 melanoma cell lines with wild-type p53; this result is consistent with a previous report showing that, in the context of absent or mutated p53, WT1 acts as a transcriptional activator, whereas in the presence of wild-type p53 it acts as a repressor.
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PMID:Expression of the wt1 Wilms' tumor gene by normal and malignant human melanocytes. 792 8

Bovine vascular smooth muscle cells (SMC) express the urokinase-type plasminogen activator receptor (u-PAR) claimed to be important in cell invasion. Receptor numbers and affinity are regulated by thrombin and several other mitogens involved in SMC proliferation. We investigated the effects of these mitogens on u-PAR mRNA levels. On continuous thrombin stimulation the u-PAR message in SMC was 10 +/- 2.3-fold elevated reaching a maximum between 6 and 9 hours and declining to control values within 48 hours. Thrombin present for 30 minutes on the cell surface produced similar effects. Stimulation with the thrombin receptor activation peptide S-F-L-L-R-N representing the NH2-terminus of the tethered ligand also increased u-PAR mRNA levels with an identical time course. D-Phe-Pro-Arg-chloromethyl ketone (PPACK) active site blocked thrombin and the catalytically inactive thrombin mutant S205A did not affect u-PAR mRNA levels. Thrombin stimulation also resulted in a 2 +/- 0.2-fold transient increase in thrombin receptor mRNA preceding the rise in u-PAR message. Transforming growth factor beta 1 (TGF beta 1) and platelet-derived growth factor (PDGF) showed similar time courses for the elevation of u-PAR mRNA levels with a maximal 5.5 +/- 0.9 and 12 +/- 2.5-fold increase, respectively. Basic fibroblast growth factor (bFGF) and phorbol myristate acetate (PMA) showed a more prolonged effect increasing u-PAR mRNA levels 8 +/- 2.0-fold and 12.3 +/- 2.5-fold, respectively, within 6 hours but remaining 5 to 10-fold elevated at 48 hours. In order to decide if the u-PAR mRNA increase was due to message stabilization or a consequence of transcriptional activation we used the RNA polymerase II inhibitor 5,6-dichloro-1-beta-D-ribofuranosyl benzimidazole (DRB) during the stimulation experiments. u-PAR mRNA levels on TGF beta 1 stimulation of SMC decayed after the addition of DRB indicating that enhancement of transcriptional activity was involved in the induction. In contrast, the time course of u-PAR mRNA elevation on thrombin, bFGF, and PMA stimulation was not significantly altered in the presence of DRB suggesting that in these latter cases u-PAR mRNA message accumulation was at least in part due to mRNA stabilization. Increased transcriptional activity, mRNA stabilization and expression of u-PAR protein on the SMC surface in response to growth factors may facilitate enhanced cell surface protease activity, cell migration, and development of atheromatous lesions.
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PMID:Effect of thrombin, the thrombin receptor activation peptide, and other mitogens on vascular smooth muscle cell urokinase receptor mRNA levels. 794 25

Since recent studies demonstrated that platelet-derived growth factor (PDGF) induces vascular smooth muscle cell (SMC) proliferation by stimulating polyamine synthesis, we examined whether the transcellular transport of L-ornithine, the cationic amino acid precursor of polyamines, could regulate the mitogenic response of PDGF. Treatment of SMC with PDGF stimulated DNA and putrescine synthesis, and this was enhanced further by increasing the extracellular concentration of L-ornithine. The potentiating effect of L-ornithine was reversed by the competitive inhibitor of cationic amino acid transport, methyl-L-arginine, or by preventing putrescine formation with alpha-difluoromethylornithine. Cationic amino acid uptake by SMC was Na+-independent and was mediated by both a high and low affinity carrier system. Treatment of SMC with PDGF initially (0-2 h) decreased basic amino acid transport, while longer exposures (6-24 h) progressively increased uptake. Kinetic studies indicated that PDGF-induced inhibition was associated with a decrease in affinity for cationic amino acids, while the stimulation was mediated by an increase in transport capacity. Endogenous PDGF released by collagen-activated platelets likewise up-regulated cationic amino acid transport in SMC. Reverse transcriptase-polymerase chain reaction detected the presence of mRNA encoding two distinct cationic amino acid transporter (CAT) proteins, CAT-1 and CAT-2B. Treatment of SMC with PDGF strongly induced the expression CAT-2B mRNA and modestly elevated the level of CAT-1 mRNA. These results demonstrate that PDGF-induced polyamine synthesis and SMC mitogenesis are dependent on the transcellular transport of L-ornithine. The capacity of PDGF to up-regulate the transport of L-ornithine by inducing the expression of the genes for CAT-1 and CAT-2B may modulate its mitogenic effect by providing SMC with the necessary intracellular precursor for polyamine biosynthesis.
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PMID:Platelet-derived growth factor regulates vascular smooth muscle cell proliferation by inducing cationic amino acid transporter gene expression. 866 68

We have investigated the regulation of p27kip1, a cyclin-dependent kinase inhibitor, in BALB/c 3T3 cells during growth factor-stimulated transition from quiescence (G0) to a proliferative (G1) state. The level of p27kip1 protein falls dramatically after mitogenic stimulation and is accompanied by a decrease in cyclin E associated p27kip1, as well as a transient increase in cyclin D1-associated p27kip1 that later declines concomitantly with the loss of total p27kip1. Analysis of metabolically labelled cells revealed that cyclin D2, cyclin D3, and cdk4 were also partnered with p27kip1 in quiescent BALB/c 3T3 cells and that this association decreased after platelet-derived growth factor (PDGF) treatment. Furthermore, the decline in p27kip1 and reduced association with cyclin D3, initiated by the addition of PDGF but not plasma-derived factors, suggested that these changes are involved in competence, the first step in the exit from G0. Synthesis of p27kip1 as determined by incorporation of [35S]methionine was repressed upon mitogenic stimulation, and PDGF was sufficient to elicit this repression within 2 to 3 h. Pulse-chase experiments demonstrated the reduced rate of synthesis was not the result of an increased rate of degradation. Full repression of p27kip1 synthesis required the continued presence of PDGF and failed to occur in the presence of the RNA polymerase inhibitor 5,6-dichlorobenzimidazole riboside. These characteristics demonstrate that repression was a late effect of PDGF and was consistent with our finding that conditional expression of activated H-ras did not affect synthesis of p27kip1. Northern (RNA) analysis of p27kip1 mRNA revealed that the repression was not accompanied by a corresponding decrease in p27kip1 mRNA, suggesting that the PDGF-regulated decrease in p27kip1 expression occurred through a translational mechanism.
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PMID:Repression of p27kip1 synthesis by platelet-derived growth factor in BALB/c 3T3 cells. 875 33

To clarify whether in vivo expression of growth factors in the glomerulus is induced in a hypertensive animal model, we investigated the expression of platelet-derived growth factor (PDGF) B chain, transforming growth factor beta (TGF-beta), and angiotensin II (Ang II) type 1 receptors in glomeruli of spontaneously hypertensive rats (SHR) and Wistar-Kyoto (WKY) rats. We also investigated the effects of treatment with cilazapril, and angiotensin I-converting enzyme inhibitor, on this expression. First, the expression of PDGF B chain, TGF-beta, and Ang II receptors from the glomerulus were investigated using the reverse-transcriptase polymerase chain reaction in SHR and WKY rats. Although there was no significant difference in PDGF B chain, TGF-beta, and Ang II receptors from the glomerulus were investigated using the reverse-transcriptase polymerase chain reaction in SHR and WKY rats. Although there was no significant difference in PDGF B chain, TGF-beta or Ang II receptor expression between SHR and WKY rats at the age of 7 weeks, the PDGF B chain expression of 16-week-old SHR was significantly higher (4.4-fold) than that of age-matched WKY rats. Next, we administered oral cilazapril at a dose of 10 mg/kg to 13-week-old SHR daily for 3 weeks. The systolic blood pressure in SHR treated with cilazapril was significantly lower than that in control SHR. After administration of cilazapril for 3 weeks, we examined the in vivo expression of growth factors and Ang II receptors in the glomerulus. The PDGF B chain expression was suppressed by treatment with cilazapril (2.5-fold) as compared with nontreated SHR. No alteration in TGF-beta or Ang II receptor expression was detected. We did not find any histological changes in the kidneys of SHR, WKY rats or cilazapril-treated SHR, and cilazapril treatment did not suppress the glomerular size. These findings indicate that the expression of PDGF B chain in the glomerulus preceded the appearance of histological changes in SHR and that the administration of cilazapril inhibited the expression of PDGF B chain without affecting the glomerular size. This suggests that angiotensin I-converting enzyme inhibitors directly suppress the Ang II-induced PDGF B chain promotion in the glomerulus of SHR at the established hypertensive stage.
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PMID:The angiotensin I-converting enzyme inhibitor, cilazapril inhibits the platelet-derived growth factor B chain expression in glomeruli of spontaneously hypertensive rats. 886 81

The ancient drug colchicine has repeatedly been proposed as a novel drug for therapy of pulmonary fibrosis. The present study was undertaken to add to the knowledge on colchicine's antiinflammatory and antifibrotic properties and thus help determine its actual rank in the treatment of pulmonary fibrosis. In vitro cell culture experiments with stimulated and unstimulated normal donor peripheral blood mononuclear cells (PMNC) and a human lung fibroblast cell line (WI-38) were used to determine the effects of colchicine on PMNC cytokine release (interleukin-6 and tumor necrosis factor-alpha) as well as on fibroblast proliferation and collagen synthesis rates. Reverse transcriptase polymerase chain amplifications of alpha 1 (III) collagen were done to detect collagen messenger ribonucleic acid (mRNA) expression. Colchicine did not significantly modulate tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) release of PMNC. Colchicine inhibited fibroblast proliferation and total collagen synthesis significantly at concentrations obtainable in serum in vivo. Transcription of the alpha 1 (III) collagen gene into mRNA continued under colchicine. We conclude that colchicine is a potent in vitro inhibitor of fibroblast functions in terms of proliferation and collagen synthesis. The mechanism of collagen inhibition is more likely an inhibition of cellular collagen secretion than a switch off of collagen mRNA transcription. On the other hand, although colchicine is known to inhibit many leukocyte functions, it is a poor inhibitor of cytokines known to be important for fibrogenesis (e.g. IL-6, TNF-alpha, IL-1, platelet-derived growth factor, and transforming growth factor-beta). This makes colchicine, at least from a theoretical standpoint and as concluded from in vitro studies, a preferable candidate for a combined therapeutic strategy.
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PMID:Antiinflammatory and antifibrotic properties of colchicine: implications for idiopathic pulmonary fibrosis. 895 72

cDNA clones encoding human (h) Grb7 and a previously unknown protein with high homology to hGrb-IR and mGrb10 (where m indicates mouse) were found by screening expressed sequence tag data bases. hGrb7 mRNA expression is greatest in pancreas and restricted to a few other tissues. The second protein termed hGrb-IRbeta/Grb10 contains an intact PH domain and lacks the 80-residue mGrb10 insertion. Expression is greatest in pancreas and muscle but occurs in nearly all tissues. hGrb-IRbeta/Grb10 and hGrb-IR likely arise as alternative mRNA splicing products of a common gene. Reverse transcriptase-coupled polymerase chain reaction shows both mRNAs in muscle. In cells, Grb-IRbeta/Grb10 protein translocates from cytosol to membrane upon insulin stimulation, most likely due to direct interactions with the insulin receptor. These interactions are mediated by the SH2 domain and additional regions of the protein. Studies with mutated receptors and synthetic phosphopeptides show that the hGrb-IRbeta/Grb10 SH2 domain binds at least two sites in the insulin receptor: the kinase activation loop > the juxtamembrane site. hGrb-IRbeta/Grb10 also binds a 135-kDa phosphoprotein in unstimulated 3T3-L1 adipocytes; binding is reduced upon insulin stimulation. In addition, the c-Abl SH3 domain binds Grb-IR/Grb10, whereas Fyn, phosphatidylinositol 3-kinase p85, and Grb2 SH3 domains do not. The site of c-Abl SH3 domain interaction is highly conserved within the Grb-IR/Grb10/Grb7/Grb14 family. hGrb-IRbeta/Grb10 also binds platelet-derived growth factor and epidermal growth factor receptors, suggesting a broader role in the signaling pathways of numerous receptors. We conclude that hGrb-IRbeta/Grb10 is a widely expressed, PH and SH2 domain-containing, SH3 domain-binding protein that functions downstream from activated insulin and growth factor receptors.
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PMID:Human GRB-IRbeta/GRB10. Splice variants of an insulin and growth factor receptor-binding protein with PH and SH2 domains. 900 1


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