Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.14.14.3 (luciferase)
 38,195 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Regulation of the human type 1 plasminogen activator inhibitor (PAI-1) promoter by transforming growth factor-beta (TGF beta) was studied. An 800-base pair fragment from the PAI-1 promoter and 5'-flanking region was fused to the firefly luciferase reporter gene and transfected into Hep3B human hepatoma cells. Treatment of the cells with TGF beta induced luciferase activity by more than 50-fold. Transfection studies using constructs with 5' or 3' deletions through this region revealed that two sequences were important in the TGF beta response. The first sequence was located in the proximal promoter (-49 to -87) and mediated an 11-fold induction with TGF beta, while the second more distal region (-636 to -740) contained two sequences which together mediated a 50-fold or greater response. Sequence comparison indicated that both of the responsive regions contained sequences with high homology to the AP-1 consensus binding site. Moreover, gel retardation analysis experiments demonstrated that both sequences bound a common nuclear protein, and that an oligonucleotide containing a consensus AP-1 sequence was able to compete for the binding of this common protein. Thus, the response of the PAI-1 gene to TGF beta is mediated by at least two separate regions, and both of these regions contain DNA sequences homologous to the AP-1 binding site.
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PMID:Identification of regulatory sequences in the type 1 plasminogen activator inhibitor gene responsive to transforming growth factor beta. 174 1

Experiments were designed to clarify the role of several proteins, junB, retinoblastoma protein (RB), and the transforming growth factor-beta (TGF-beta) receptors that are potential intermediates in TGF-beta activation of the alpha 2(I) collagen promoter. Treatment of NIH-3T3 cells with TGF-beta increased the activity of a transiently transfected murine alpha 2(I) collagen promoter (nucleotides -350 to +54) fused to a luciferase reporter gene 9-fold. Cotransfection of a junB stimulated the basal activity of the alpha 2(I) collagen promoter 93-fold, respectively. Expression of antisense junB RNA attenuated the effect of TGF-beta. Simian virus 40 large T antigen, an inhibitor RB function, did not prevent TGF-beta effects on the alpha 2(I) collagen promoter. A chimeric receptor containing the extracellular domain of the colony-stimulating factor-1 receptor and the intracellular domain of the type I TGF-beta receptor enhanced alpha 2(I) collagen promoter activity 4.8-fold, whereas a similar chimera containing the type II receptor intracellular domain had much weaker effects. Similar results were obtained with a plasminogen activator inhibitor-1 promoter, previously shown to be activated by TGF-beta through AP-1 elements. We conclude that TGF-beta activates the alpha 2(I) collagen and plasminogen activator inhibitor-1 promoters in NIH-3T3 cells through junB and the type I TGF-beta receptor kinase domain.
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PMID:Requirements for transforming growth factor-beta regulation of the pro-alpha 2(I) collagen and plasminogen activator inhibitor-1 promoters. 787 14

Retinoic acid (RA)-treated HL-60 cells were used as a model to study differentiation of granulocytic leukemias. RA induces these cells to mature into granulocytes and to decrease growth. Mediators of these RA effects have not been identified definitively, but transforming growth factor-beta (TGF-beta) has been implicated in regulating proliferation and differentiation of myelogenous leukemic cells. The role of TGF-beta in RA-dependent differentiation and cessation of growth was examined by adding neutralizing anti-TGF-beta IgG to RA-treated HL-60 cells, followed by assessing cell growth and markers of granulocytic differentiation over 5 days. After addition of neutralizing anti-TGF-beta IgG, growth of RA-treated HL-60 cells was maintained at control levels, but granulocytic differentiation continued. These experiments demonstrated that the antiproliferative activity of RA was TGF-beta dependent but that differentiation was not. Because most cell types secrete TGF-beta in a biologically inactive complex, a TGF-beta-dependent effect requires cells to activate the latent form of TGF-beta. Active and total TGF-beta levels were quantitated in media harvested from control and RA-treated cells using a luciferase-based bioassay for TGF-beta activity. Similar levels of total TGF-beta were observed between control and RA-treated cells. RA-treated cells produced active TGF-beta (18-24 pg/ml) after 1, 2, and 3 days of treatment, whereas negligible levels were produced by control cultures. Activation of endogenous latent TGF-beta by RA-treated cells occurred through a plasmin-independent mechanism.
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PMID:Effects of endogenously activated transforming growth factor-beta on growth and differentiation of retinoic acid-treated HL-60 cells. 856 60

The small leucine-rich proteoglycan biglycan is involved in several physiological and pathophysiological processes through the ability of its core protein to interact with other extracellular matrix molecules and transforming growth factor-beta (TGF-beta). To learn more about the regulation of biglycan core protein expression, we have cloned and sequenced 1218 base pairs from the 5'-flanking region of the human biglycan gene, demonstrated functional promoter activity, and investigated the molecular mechanisms through which various agents modulate its transcriptional activity. Sequencing revealed the presence of several cis-acting elements including multiple AP-2 sites and interleukin-6 response elements, a NF-kappaB site, a TGF-beta negative element, and an E-box. The TATA and CAAT box-lacking promoter possesses many features of a growth-related gene, e.g. a GC-rich immediate 5' region, many Sp1 sites, and the use of multiple transcriptional start sites. Transient transfections of the tumor cell lines MG-63, SK-UT-1, and T47D with various biglycan 5'-flanking region-luciferase reporter gene constructs showed that the proximal 78 base pairs are sufficient for full promoter activity. Several agents among them interleukin-6, and tumor necrosis factor-alpha. were capable of altering biglycan promoter activity. However, in MG-63 cells, TGF-beta1 failed to increase either activity of the biglycan promoter constructs or specific transcription from the endogenous biglycan gene. Since TGF-beta1 also did not alter the stability of cytoplasmic biglycan mRNA as determined from Northern analysis after inhibition of transcription with 5,6-dichloro-1beta-D-ribofuranosylbenzimidazole, an as yet unidentified nuclear post-transcriptional mechanism was considered responsible for the TGF-beta effect in this cell type. These results might help to elucidate the molecular pathways leading to pathological alterations of biglycan expression observed in atherosclerosis, glomerulonephritis, and fibrosis.
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PMID:Transcriptional regulation of the human biglycan gene. 866 74

The kinases and regulatory proteins that convey signals initiated by transforming growth factor-beta (TGF-beta) to the nucleus are poorly characterized. To study the role of the extracellular signal-regulated kinase (ERK) pathway in this process, we transiently transfected NIH 3T3 fibroblasts with TGF-beta-responsive luciferase reporter genes and expression vectors designed to interrupt this kinase cascade. Mitogen-activated protein (MAP) kinase phosphatase-1 and a dominant negative MAP/ERK kinase 1 mutant reduced stimulation of plasminogen activator inhibitor-1 (PAI-1) promoter activity by TGF-beta1 from 11.5- to 4-fold and 4.9-fold, respectively. Similar results were observed with the type I collagen promoters. TGF-beta1 increased ERK1 activity 4.5-fold at 5 min and 3. 1-fold at 3 h, while Jun kinase and p38 activity were not affected. Cotransfection of a dominant negative mutant of the small G protein, Rac, but not dominant negative Ras, Cdc42, or Rho mutants, reduced the effects of TGF-beta1 on the PAI-1 promoter by approximately half. In support of a role for Rac in signaling by TGF-beta, GTP binding to Rac was increased 3.7-fold following exposure of NIH 3T3 cells to TGF-beta1 for 3 min. These findings indicate that TGF-beta1 modulates gene expression partly through ERK and Rac in NIH 3T3 cells.
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PMID:Extracellular signal-regulated kinase and the small GTP-binding protein, Rac, contribute to the effects of transforming growth factor-beta1 on gene expression. 866 31

We have examined the expression of the alpha 1(IV) collagen gene in murine proximal tubular cells (MCT) to better understand how it is regulated in parenchymal cells. Transcriptional activity was examined using luciferase reporters driven by the alpha 1(IV) promoter and varying lengths of 5'-flanking sequences. The minimal bidirectional promoter showed low intrinsic activity in MCT cells, but addition of upstream sequences increased luciferase expression. Maximal activity resided within the first 1,200 bp upstream. A minigene construct was generated by placing a portion of the alpha 1(IV) first intron downstream from the promoter region. The intronic sequences significantly decreased activity of the promoter in MCT cells and 3T3 fibroblasts but greatly enhanced expression in murine parietal yolk sac (PYS) endodermal cells. Addition of transforming growth factor-beta (TGF-beta) to MCT cultures elevated the levels of secreted type IV collagen. Treatment of either transiently or stably transfected MCT cells with TGF-beta produced an increase in the levels of expression of all of the reporters tested. These data support the hypothesis that cell-specific regulation of alpha 1(IV) collagen is dependent upon downstream sequences, which act to decrease the expression of type IV collagen in tubular epithelium. The activity of the alpha 1(IV) collagen gene in proximal tubular cells is increased by TGF-beta, which acts on the domain(s) embedded within the intergenic bidirectional promoter.
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PMID:Transforming growth factor-beta modulation of the alpha 1(IV) collagen gene in murine proximal tubular cells. 876 Feb 52

Glial cell line-derived neurotrophic factor (GDNF), a member of the transforming growth factor-beta family of growth factors, was first identified by its ability to promote the survival of midbrain dopaminergic neurons in culture. We demonstrate that GDNF treatment of several neuroblastoma cell lines leads to dose-dependent tyrosine phosphorylation of the RET receptor and that other transforming growth factor-beta family members are not able to activate the RET receptor. GDNF treatment of neuroblastoma cells also results in increased transcription of an Elk luciferase reporter gene, suggesting that GDNF activates the mitogen-activated protein kinase signal transduction pathway.
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PMID:Glial cell line-derived neurotrophic factor signals through the RET receptor and activates mitogen-activated protein kinase. 879 76

Lineage analysis studies in the avian embryo have identified two types of smooth muscle cells (SMCs) in the tunica media of large elastic arteries; one that originates within the cardiac neural crest and is ectoderm in origin (Ect) and another that arises from local mesenchyme of mesodermal origin (Mes). To determine if differences in primary embryonic lineage can give rise to SMCs with stable differences in growth and differentiation properties, we isolated Ect and Mes SMCs from the Day 14 chick embryo aorta. We report that despite different primary embryonic origins, Ect and Mes SMCs express nearly identical levels of seven SMC differentiation markers in vitro, consistent with their common smooth muscle developmental fates in vivo. By contrast, Ect SMCs displayed a greater capacity for growth in serum-free medium than Mes SMCs, but only under conditions permitting short-range cell-cell interactions. Most of the peptide growth factors tested that might account for serum-independent growth (PDGF-AA, PDGF-BB, basic FGF, EGF, or activin) stimulated DNA synthesis to similar extents in Ect and Mes SMCs. However, we found dramatic, lineage-dependent differences in SMC responses to transforming growth factor-beta (TGF-beta). Exposure to TGF-beta1 (0.4 to 400 pmole/liter) consistently increased DNA synthesis in Ect SMCs, whereas in paired cultures of Mes SMCs, TGF-beta1 was growth inhibitory. In SMC cultures transfected with p3TP-lux, a luciferase reporter controlled by the TGF-beta1-response elements of the human PAI-1 promoter, TGF-beta1 (120 pM) produced 12 ± 2-fold increases in luciferase activity in Ect SMCs and only 3 ± 1.5-fold increases in Mes SMCs. Analysis of TGF-beta receptor phenotypes by Northern blot, radioligand binding, and crosslinking assays showed that Ect and Mes SMCs expressed similar levels of types I, II, and III TGF-beta receptors. However, using a polyclonal antibody specific for the chick type II TGF-beta receptor subunit, we demonstrate that Mes SMCs produce a fully glycosylated form of this protein while Ect SMCs elaborate only an unglycosylated type II TGF-beta receptor. These results show that Ect and Mes SMCs exhibit lineage-dependent differences in growth and receptor-mediated transcriptional responses to at least one important class of SMC morphogens and growth modifiers, e.g., the TGF-betas. Our findings suggest that different SMC populations within a common vessel wall may respond in lineage-dependent ways to signals that direct formation of the tunica media in the embryo and to factors involved in the progression of vascular disease later in life.
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PMID:Smooth Muscle Lineage Diversity in the Chick Embryo 881 40

Lineage analysis studies in the avian embryo have identified two types of smooth muscle cells (SMCs) in the tunica media of large elastic arteries; one that originates within the cardiac neural crest and is ectoderm in origin (Ect) and another that arises from local mesenchyme of mesodermal origin (Mes). To determine if differences in primary embryonic lineage can give rise to SMCs with stable differences in growth and differentiation properties, we isolated Ect and Mes SMCs from the Day 14 chick embryo aorta. We report that despite different primary embryonic origins, Ect and Mes SMCs express nearly identical levels of seven SMC differentiation markers in vitro, consistent with their common smooth muscle developmental fates in vivo. By contrast, Ect SMCs displayed a greater capacity for growth in serum-free medium than Mes SMCs, but only under conditions permitting short-range cell-cell interactions. Most of the peptide growth factors tested that might account for serum-independent growth (PDGF-AA, PDGF-BB, basic FGF, EGF, or activin) stimulated DNA synthesis to similar extents in Ect and Mes SMCs. However, we found dramatic, lineage-dependent differences in SMC responses to transforming growth factor-beta (TGF-beta). Exposure to TGF-beta 1 (0.4 to 400 pmole/liter) consistently increased DNA synthesis in Ect SMCs, whereas in paired cultures of Mes SMCs, TGF-beta 1 was growth inhibitory. In SMC cultures transfected with p3TP-lux, a luciferase reporter controlled by the TGF-beta 1-response elements of the human PAI-1 promoter, TGF-beta 1 (120 pM) produced 12 +/- 2-fold increases in luciferase activity in Ect SMCs and only 3 +/- 1.5-fold increases in Mes SMCs. Analysis of TGF-beta receptor phenotypes by Northern blot, radioligand binding, and crosslinking assays showed that Ect and Mes SMCs expressed similar levels of types I, II, and III TGF-beta receptors. However, using a polyclonal antibody specific for the chick type II TGF-beta receptor subunit, we demonstrate that Mes SMCs produce a fully glycosylated form of this protein while Ect SMCs elaborate only an unglycosylated type II TGF-beta receptor. These results show that Ect and Mes SMCs exhibit lineage-dependent differences in growth and receptor-mediated transcriptional responses to at least one important class of SMC morphogens and growth modifiers, e.g., the TGF-betas. Our findings suggest that different SMC populations within a common vessel wall may respond in lineage-dependent ways to signals that direct formation of the tunica media in the embryo and to factors involved in the progression of vascular disease later in life.
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PMID:Smooth muscle lineage diversity in the chick embryo. Two types of aortic smooth muscle cell differ in growth and receptor-mediated transcriptional responses to transforming growth factor-beta. 883 Jul 42

We recently identified a bipartite element in the rat osteocalcin (OC) promoter that confers synergistic induction by fibroblast growth factor receptor 2 (FGF2) and cAMP. A GCAGTCA motif (OCFRE) at -146 to -138 in the OC promoter is necessary for synergy and participates in a FGF2-regulated DNA-protein interaction. We have isolated the FGF-regulated component of this transcriptional response for detailed study. Two or three copies of the OC promoter fragment -154 to -113 with the intact OCFRE confer 10- or 30-fold FGF2-inductive responses, respectively, on the unresponsive basal promoter 92 OCLUC (luciferase reporter) in MC3T3-E1 cells; a single copy is insufficient. As in the native context, induction depends upon an intact OCFRE motif (mutant GCATTTA motifs unresponsive). FGF receptor 1 can mediate activation; expression of this receptor in L6 cells (no endogenous FGF receptors) permits FGF2 induction of (OCFRE)2 92 OCLUC. FGF2 induction of (OCFRE)2 92 OCLUC in MC3T3-E1 cells is not recapitulated by platelet-derived growth factor-BB, epidermal growth factor, insulin-like growth factor I, or transforming growth factor-beta (< 10% the activity of FGF2). OCFRE activation is not inhibited by kinase inhibitors H-89, wortmannin, staurosporine, KN-62, or H-7. However, the phosphoprotein phosphatase inhibitors okadaic acid (OKA), calyculin A, and vanadate decrease FGF induction of (OCFRE)2 92 OCLUC or (OCFRE)3 92 OCLUC, without inhibiting induction of the interstitial collagenase promoter. OKA and calyculin A do not decrease OCFRE DNA-protein interactions, suggesting that important protein-protein interactions are phosphatase regulated. These data provide evidence that; 1) FGF receptors elaborate transcriptional activation signals that functionally differ from those of other receptor tyrosine kinases; 2) an OKA-sensitive phosphatase participates in FGF receptor-dependent activation of the OCFRE; and 3) two transcriptional activation signals are initiated by FGF receptor activation in MC3T3-E1 cells, reflected in the divergent sensitivities of OCFRE and interstitial collagenase promoter induction to OKA and vanadate.
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PMID:The rat osteocalcin fibroblast growth factor (FGF)-responsive element: an okadaic acid-sensitive, FGF-selective transcriptional response motif. 884 19


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