Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.21.73 (urokinase-type plasminogen activator)
 10,685 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Plasminogen activator inhibitor-1 (PAI-1) is a serine protease inhibitor that inhibits both tissue-type and urokinase-type plasminogen activators. Expression of PAI-1 is regulated by growth factors, cytokines, and hormones. To determine the molecular mechanisms involved in the basal expression of the rat PAI-1 gene, we have analyzed the cis-acting sequences and the trans-acting factors involved in the transcription of this gene in the HTC rat hepatoma cell line. DNase I protection analyses revealed eight regions within the first 764 base pairs of 5'-flanking sequence that interact specifically with HTC cell nuclear proteins. The proteins that bind to five of the eight footprinted sites were identified as PEA3-, Sp1-, and CTF/NF-1-like proteins using competition electrophoretic mobility shift assays. The expression of fusion genes containing progressive 5' deletions of the rat PAI-1 promoter linked to the chloramphenicol acetyltransferase reporter gene were analyzed in transient transfection experiments in HTC cells. These studies demonstrated the Sp1 and CTF/NF-1 sites to be important for transcriptional activation. Two of the footprinted sites contain the sequence 5'-TTTGn(n)TCAAT-3' and were shown in competition electrophoretic mobility shift assays to bind the same or related protein(s). Sequences containing these sites, from -764 to -628 base pairs, and from -266 to -188 base pairs, were identified in functional studies as repressor elements of transcription.
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PMID:Regulatory sequences and protein-binding sites involved in the expression of the rat plasminogen activator inhibitor-1 gene. 160 87

We have studied the regulation of urokinase-type plasminogen activator gene expression by cAMP in LLC-PK1 cells. We found a cAMP responsive region 3.4 kb upstream of the transcription initiation site, which comprised three protein-binding domains designated A, B, and C. Domains A and B both contain a sequence, TGACG, homologous to a consensus cAMP response element (CRE; TGACGTCA). Effective cAMP-mediated induction was achieved when these two domains were linked with domain C, which by itself did not confer cAMP responsiveness to a heterologous promoter nor contained CRE-like sequence, suggesting a functional cooperation among these domains. Results of competition studies using gel retardation and DNase I footprinting assays suggest that there is a protein-protein interaction between a CRE binding protein and a domain C binding protein. In gel retardation assays, binding of a nuclear protein to domains A and B was strongly augmented by addition of the catalytic subunit of cAMP-dependent protein kinase, whereas the protein binding to domain C was slightly inhibited, suggesting that protein phosphorylation is involved in the regulation of protein-DNA interaction.
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PMID:Macromolecular interaction on a cAMP responsive region in the urokinase-type plasminogen activator gene: a role of protein phosphorylation. 215 33

In LLC-PK1 cells, the urokinase-type plasminogen activator (uPA) gene is induced by two of the major signal transduction pathways, the protein kinase C (PKC) and the cAMP-dependent protein kinase (PKA) pathways. We have analyzed the chromatin structure of 26 kb of the uPA gene locus and have shown that PKA activation but not PKC activation induce major chromatin structural alterations in the uPA gene promoter. In uninduced cells, several DNase I hypersensitive (HS) sites were detected in the 5' and 3' flanking regions but not in the transcribed region. Two of the sites correspond to previously characterized regulatory sites: a cAMP responsive site at nucleotide position -3500 with respect to the initiation site, and the PEA3/AP1 site at -2100 that mediates PKC activation. After the activation of PKA but not PKC, a strong HS site was induced at -2600. Functional analysis of this region revealed cAMP responsive activity. Chromatin structural alterations again brought about specifically by PKA but not by PKC were were also detected in the upstream of the promoter by topoisomerase I cleavage site analysis, with two prominent sites appearing at -2800 and -3300. These results suggest that the strong cAMP induction of the uPA gene requires structural alterations that permit cooperative interactions between the multiple cAMP responsive sites.
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PMID:Activation of cAMP-dependent protein kinase alters the chromatin structure of the urokinase-type plasminogen activator gene promoter. 812 5

We have previously shown in NIH 3T3 fibroblasts that treatment with 12-O-tetradecanoylphorbol 13-acetate (TPA) or fibroblast growth factor-2 (FGF-2) activates the Ras/Erk signaling pathway in NIH 3T3 fibroblasts, leading to the induction of the urokinase-type plasminogen activator (uPA) gene. In this study, we characterize cis-acting elements involved in this induction. DNase I hypersensitive (HS) site analysis of the uPA promoter showed that two regions were enhanced after TPA and FGF-2 treatment. One was located 2.4kb upstream of the transcription start site (-2.4kb), where a known PEA3/AP1 (AGGAAATGAGGTCAT) element is located. The other was located in a previously undefined far upstream region. Sequencing of this region revealed a similar AP1/PEA3 (GTGATTCACTTCCT) element at -6.9 kb corresponding to the HS site. Deletion analysis of the uPA promoter in transient transfection assays showed that both PEA3/AP1 elements are required for full inducibility, suggesting a synergism between the two elements. When the two sites were inserted together upstream of a minimal promoter derived from the thymidine kinase gene, expression of the reporter gene was more strongly induced by TPA and FGF-2 than with either of the two elements alone. Alone, the -6.9 element was more potent than the -2.4 element. The involvement of AP1 as well as Ets transcription factors was confirmed by examining different promoter constructs containing deletions in either the AP-1 or the PEA3 element, and by using an expression plasmid for dominant negative Ets-2. Electromobility shift analyses using specific antibodies showed that c-Jun and, JunD bind to both elements with or without induction. In addition, ATF-2 binds to the -2.4-kb element even without induction and c-Fos to the -6.9-kb element only after induction. Accordingly, overexpression of c-Fos caused induction from the -6.9-kb element, but reduced induction from the -2.4-kb element. The involvement of the Ets-2 transcription factor was shown by using expression plasmids for wild-type and dominant negative Ets-2.
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PMID:Cooperation of two PEA3/AP1 sites in uPA gene induction by TPA and FGF-2. 940 85

We have analysed in vivo the -2.0 kb enhancer of the human urokinase-type plasminogen activator (uPA) gene in HepG2 cells, in which gene expression can be induced by phorbol esters. The results reveal that, within the regulatory region, the enhancer, the silencer and the minimal promoter become hypersensitive to deoxyribonuclease I (DNase I) upon induction of transcription. The hypersensitivity of the enhancer can be reversed after removal of the inducer. In vivo footprinting analysis indicates that all the cis-acting elements of the enhancer, previously identified in vitro, are occupied in vivo upon 12-O-tetradecanoyl-phorbol-13-acetate (TPA) stimulation of HepG2 cells. Micrococcal nuclease (MNase) cleavage of this region fails to reveal discrete nucleosomal boundaries in vivo in close proximity of the enhancer, either before or after stimulation. Furthermore, this region does not lose its nucleosomal configuration after TPA induction of transcription. An approximately 600 bp long region around the enhancer becomes more, but not fully, accessible to restriction endonucleases upon stimulation. A time-course experiment shows that this accessibility reaches a plateau after a 1 h TPA treatment suggesting the persistent presence of nucleosomes. These results indicate that TPA induces the binding of transcription factors to the uPA enhancer without chromatin remodelling of this region.
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PMID:In vivo analysis of the state of the human uPA enhancer following stimulation by TPA. 1036 54

We have previously shown that urokinase-type plasminogen activator (uPA) is highly expressed in murine C2C12 myoblasts and that antibodies against uPA are able to block both myoblast fusion and differentiation. Here we show the characterization of cis-acting elements in the mouse uPA promoter in vitro which are involved in uPA gene expression in C2C 12 myoblast cells. DNase I hypersensitive (HS) site analysis revealed the presence of three HS sites in myoblasts. Deletion analysis of stably transfected uPA-promoter constructs revealed that at least two of the three HS sites accounted for the high transcriptional expression in C2C12 cells. One was located at -2.4 kb and corresponded to a known PEA3/AP1A element and the other one was located at -4.9 kb and contained a CArG box and a CRE element. So far, no regulatory function had been assigned to this CRE/CArG element. Both HS sites alone were able to activate transcription of a heterologous promoter and showed a cooperative effect when placed together. Electrophoretic mobility-shift assays using myoblast nuclear extracts and specific antibodies demonstrated that cJun, JunD and ATF2 bound to the PEA3/AP1A element, whereas the CRE/CArG element bound SRF. Altogether, these results suggest that high uPA expression in myoblasts is dependent on the cooperation of two regulatory sites in the uPA promoter.
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PMID:Transcriptional regulation of the murine urokinase-type plasminogen activator gene in skeletal myoblasts. 1036 52