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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)

The 5' flanking regions of the mouse and pig urokinase plasminogen activator (uPA) genes were sequenced and sequence homology interrupted by repeat elements was found to extend to -4.6kb in pig and -6.6kb in mouse. A transient transfection procedure was devised for the murine macrophage cell line RAW264. Pig uPA promoter-CAT constructs were more active than mouse constructs in this assay. This contrast may involve sequence differences within 100 bp of the transcription start site. The selective deletion of distal regions of the promoter (greater than 2.6 kb upstream), and of a conserved element, 5'-AGGAGGAAATGAGG-TCA-3' around -2 kb greatly reduced the activity of reporter constructs in RAW264 cells. Electrophoretic mobility shift assays using the latter sequence identified a single nuclear protein complex. This element has been referred to as PEA3/AP1-like, but the complex did not comigrate with either AP1 or known proteins that bind polypurines (including the macrophage-specific factor PU-1) and was not competed by AP1 or polypurine oligonucleotides. uPA promoters contain multiple AP1 and AP2-like DNA sequences, which were recognised by nuclear proteins expressed constitutively in RAW264 cells. They also contain multiple binding sites for NF kappa B but activated NF kappa B was not expressed in RAW264 cells. The conserved, transcribed 5' non-coding sequences were also required for maximal gene expression. Hence, the uPA promoter contains multiple weak cis-acting elements distributed over 7.0 kb 5' to the translation start site.
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PMID:Constitutive expression of the urokinase plasminogen activator gene in murine RAW264 macrophages involves distal and 5' non-coding sequences that are conserved between mouse and pig. 176 14

The fosB gene encodes a nuclear protein that shows a high degree of homology with c-Fos in several of the known functionally crucial domains, e.g., the leucine zipper and the DNA-binding site, but shows considerable divergence in other regions. Here, we report that FosB, when placed under the control of a constitutive promoter, exhibits clear transforming properties in focus assays using mouse NIH3T3 or rat 208F fibroblasts. The transforming potential of FosB is considerably stronger than that of a corresponding c-fos construct and resembles that of viral fos genes. Using chimeric fos/fosB constructs we show that the C-terminal half of FosB is responsible for these stronger transforming properties, apparently by giving rise to significantly higher levels of protein as compared with the corresponding c-fos sequence. Surprisingly, substitution of the N-terminus of Fos with that of FosB decreases its transforming potential. These differences in the transforming potential are not related to DNA or protein expression, but rather seem to reflect differences in the molecular function(s) encoded in the N-terminal halves of Fos and FosB protein. Both, fosB- and v-fos transformed cells show increased expression of a number of endogenous genes, including c-jun, transin, alpha 1(III) collagen and tissue plasminogen activator. Transactivation by FosB and v-fos of the c-jun and alpha 1(III) collagen gene promoters and of a 3 x TRE-tk chimeric promoter could be shown in transient CAT assays. v-Fos, but not FosB-transformed cells, also show elevated levels of urokinase and plasminogen activator inhibitor mRNAs, pointing to potential differences in the gene regulatory properties of the two Fos family members.
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PMID:fosB is a transforming gene encoding a transcriptional activator. 190 95

The effects of glucagon, heparin, urokinase, thromboxane synthetase inhibitor (OKY-046), and the free radical scavengers, superoxide dismutase and catalase (SOD + CAT), on the viability of ischemic intestine were evaluated based on various parameters measured. The mucosal blood flow, the fluorescence pattern, and the histopathological findings in a rabbit model with 3.5 hr total vascular occlusion of a short small intestine indicated that glucagon improved the ischemic intestine. Glucagon increased, tremendously, the mucosal blood flow by 112% in the ischemic intestine compared with that of 25% in the nonischemic intestine. This indicated that vascular spasm, not reperfusion injury or thrombosis, played the initial role in the progression of transmural bowel necrosis. In addition, the outcome in the viability of the ischemic intestine was not detected by the fluorescence technique but was able to be detected through the mitochondrial morphology under the electron microscope.
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PMID:Does glucagon improve the viability of ischemic intestine? 226 88

The urokinase-type plasminogen activator plays a central role in tissue remodeling by controlling the synthesis of the extracellular matrix-degrading plasmin. Urokinase expression is transcriptionally regulated by a variety of cytokines including TNF-alpha. The present study was undertaken to identify key transcription factor binding sites in the urokinase promoter necessary for the TNF-alpha-dependent induction of urokinase expression. TNF-alpha treatment of a squamous cell carcinoma cell line, UM-SCC-1, which produces no detectable TNF-alpha, led to a dose-dependent increase in urokinase secretion, thus reflecting a more abundant mRNA. Transient transfections of UM-SCC-1 cells with a CAT reporter driven by 5' deletion fragments of the urokinase promoter indicated that a sequence spanning -2109 to -1870, which contained binding sites for AP-1 and PEA3 was required for the stimulation by TNF-alpha. Mutation of an AP-1 binding site at -1967 and a PEA3 motif at -1973 completely abrogated the inductive effect of TNF-alpha on urokinase promoter activity. Mobility shift assays indicated the presence of a jun-containing factor(s) which bound specifically to the AP-1 sequence present in the urokinase promoter. The amount and/or activity of this factor(s) was greatly enhanced by TNF-alpha treatment. UM-SCC-1 cells transiently transfected with a CAT reporter driven by 3 tandem AP-1 binding sites demonstrated increased CAT activity following TNF-alpha treatment. Thus, the induction of urokinase expression by TNF-alpha is likely to involve the altered expression and/or activity of transcription factors which bind to the AP-1 and PEA3 target sequences in the urokinase promoter.
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PMID:Stimulation of urokinase expression by TNF-alpha requires the activation of binding sites for the AP-1 and PEA3 transcription factors. 762 64

The HER2/neu (c-erbB2) protooncogene, which encodes a transmembrane receptor (p185neu), contributes to tumor cell invasion/metastasis through mechanism(s) which are, at present, poorly defined. Since basement membrane degradation is a prerequisite for tumor progression, we undertook a study to determine if the expression of urokinase, a key protease implicated in extracellular matrix proteolysis, was regulated by this oncogene. Stable overexpression of a cDNA encoding HER2/neu in H460 lung cancer cells led to elevated secretion of urokinase which was a consequence of a higher level of protease mRNA. Transfection of the HER2/neu-overexpressing B 104-1 cells with a CAT reporter construct driven by the urokinase promoter, gave rise to increased CAT activity when compared with parental NIH3T3 cells, which have low levels of HER2/neu, suggesting that the protooncogene can enhance urokinase promoter activity. Since the enhanced expression of HER2/neu results in increased tumor invasion/metastasis (1), these data suggest that, at least in vitro, HER2/neu-induced expression of urokinase may contribute to tumor progression in p185neu-positive cancers.
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PMID:Up-regulation of urokinase-type plasminogen activator expression by the HER2/neu proto-oncogene. 765 95

We undertook a study to determine if the serine-threonine kinase-encoding v-mos oncogene regulated the expression of the urokinase-type plasminogen activator. An expression vector encoding v-mos, but not a kinase-inactive mutant, stimulated urokinase promoter activity in CAT assays employing a squamous cell carcinoma cell line. The induction of urokinase promoter activity by v-mos was mediated, in part, via an increased AP-1 activity since (a) mutation of 2 AP-1 binding sites (at -1967 and -1885), or the co-expression of a transactivation domain-lacking c-jun mutant reduced the induction of the urokinase promoter by v-mos and (b) expression of v-mos increased the activity of a CAT reporter driven by three AP-1 tandem repeats. The stimulation of the urokinase promoter by v-mos was partially countered by co-expression of an ERK1/ERK2-inactivating phosphatase. Western blotting and zymographic analysis indicated that v-mos-transformed NIH3T3 cells (MSV NIH-3T3) secreted more urokinase compared with NIH3T3 cells and this was associated with a higher level of activated ERK1 and ERK2. Expression of a catalytically-inactive MAPKK mutant reduced the activity of a urokinase promoter-driven CAT reporter in the MSV NIH-3T3 cells. In conclusion, the data herein indicate that urokinase expression is regulated by v-mos through a MAPKK-dependent signaling pathway.
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PMID:Regulation of urokinase-type plasminogen activator expression by the v-mos oncogene. 854 21

The urokinase-type plasminogen activator contributes to tissue remodeling by controlling the synthesis of the extracellular matrix-degrading plasmin. We undertook a study to determine the role of the extracellular signal-regulated kinases (ERKs) in the regulation of urokinase-type plasminogen activator expression in a squamous cell carcinoma cell line (UM-SCC-1) that contains a transcriptionally activated urokinase-type plasminogen activator gene. Transient transfection studies using a CAT reporter driven by the urokinase-type plasminogen activator promoter, which had progressive 5' deletions or which had been point-mutated, indicated the requirement of binding sites for AP-1 (-1967) and PEA3 (-1973) for its maximal activation. Expression of a mutant jun protein, which lacks the transactivation domain, caused a dose-dependent repression of a CAT reporter driven by either the urokinase-type plasminogen activator promoter or three tandem AP-1 repeats upstream of a thymidine kinase minimal promoter indicating the importance of AP-1-binding transcription factor(s) in the regulation of urokinase-type plasminogen activator synthesis. Mobility shift assays with UM-SCC-1 nuclear extract revealed binding of fos and junD proteins to an oligonucleotide spanning the AP-1 site at -1967. In-gel kinase assays indicated the constitutive activation of ERK1, which regulates fos synthesis via phosphorylation of p62TCF, but not ERK2, in UM-SCC-1 cells. Moreover, the expression of a dominant-negative ERK1, but not ERK2, repressed urokinase-type plasminogen activator promoter activity. Similarly, interfering with the function of the c-raf serine-threonine kinase, which lies upstream of ERK1, by the expression of a kinase-inactive c-raf repressed the activity of a CAT reporter driven by either the urokinase-type plasminogen activator promotor or tandem AP-1 repeats. These data suggest that urokinase-type plasminogen activator expression in UM-SCC-1 cells is regulated partly by an ERK1, but not ERK2, -dependent signaling pathway.
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PMID:Regulation of urokinase-type plasminogen activator expression by an ERK1-dependent signaling pathway in a squamous cell carcinoma cell line. 876 47

The urokinase-type plasminogen activator receptor (u-PAR) facilitates extracellular matrix degradation in part by accelerating plasmin formation at the cell surface. We previously reported that u-PAR expression is elevated in colon cancer cell lines characterized by their in vitro invasive capacity. Since, u-PAR expression is increased by a variety of growth factors, which signal through the extracellular signal-regulated kinases 1 and 2 (ERK1/ERK2), we determined if these mitogen-activated protein kinases (MAPKs) regulate u-PAR expression in two cultured colon cancer cell lines. An in-gel kinase assay showed that ERK1 activity was considerably higher in RKO cells, which display > or = 10(5) receptors/cell, than the GEO cells which have approximately 10(4) urokinase receptors per cell. The expression of either an ERK-inactivating phosphatase (CL100), or a kinase-defective ERK1, decreased the activity of a u-PAR promoter-driven CAT reporter in RKO cells. Immune complex kinase assays indicated that the constitutive ERK1 activity in RKO cells was largely a result of an activated MEK1. Further, treatment of RKO cells with a specific inhibitor (PD 098059) of MEK1 activation, which diminished ERK1 activity, reduced the amount of urokinase specifically bound to the cell surface and this was associated with reduced laminin degradation. The expression of a dominant negative c-Raf-1 also reduced u-PAR promoter activity suggesting that MEK1 activation involved an activator at, or upstream, of this serine-threonine kinase. Transfection of the u-PAR-deficient GEO cells with a constitutively activated MEK1 expression construct up-regulated u-PAR promoter activity. Similarly treatment of GEO cells with a phosphatase inhibitor (sodium vanadate) caused a dose-dependent increase in ERK1 activity which paralleled increased cell surface binding of urokinase. Taken together, these data suggest that elevated u-PAR expression, in at least a sub-population of colon cancer, is partly a consequence of a constitutively activated ERK-1-dependent signaling cascade.
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PMID:Elevated urokinase-type plasminogen activator receptor expression in a colon cancer cell line is due to a constitutively activated extracellular signal-regulated kinase-1-dependent signaling cascade. 919 Oct 56

Since c-src overexpression increases colonic cell invasiveness and because both Src activity and urokinase receptor protein are elevated in invasive colon cancers, the present study was undertaken: 1) to determine if a constitutively active Src regulates urokinase receptor expression and 2) to identify required cis-elements and trans-acting factors. SW480 colon cancer cells transfected with an expression plasmid (c-srcY527F) encoding a constitutively active Src protein manifested increased urokinase receptor gene expression and Src activity. Treatment of the src transfectants with a Src-inhibitor (PD173955) reduced urokinase receptor protein levels and laminin degradation. Inasmuch as we recently implicated an upstream region of the urokinase receptor promoter (-152/-135) in constitutive urokinase receptor expression, we determined its role for the induction by src. Whereas the activity of a CAT reporter driven by this region was stimulated by c-srcY527F, the u-PAR promoter mutated at the Sp1-binding motif in the -152/-135 region was not. Nuclear extracts from the src transfectants demonstrated increased Sp1 binding to region -152/-135 compared with those from SW480 cells. Finally, endogenous urokinase receptor protein amounts in 10 colon cancers and corresponding normal colon correlated with Src specific activity. These data suggest that urokinase receptor gene expression is regulated by Src partly via increased Sp1 binding.
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PMID:Transcriptional induction of the urokinase receptor gene by a constitutively active Src. Requirement of an upstream motif (-152/-135) bound with Sp1. 1037 50

The Rel/NF-kappaB transcription factors regulate the expression of many genes. The activity of RelA, a member of the Rel/NF-kappaB transcription factor family, is constitutively activated in the majority of pancreatic adenocarcinomas and cell lines. We report that the urokinase-type plasminogen activator (uPA), one of the critical proteases involved in tumor invasion and metastasis, is overexpressed in pancreatic tumor cells and its overexpression is induced by constitutive RelA activity. The uPA promoter contains an NF-kappaB binding site that directly mediates the induction of uPA expression by RelA. Expression of a dominant-negative IkappaBalpha mutant inhibits kappaB site-dependent transcriptional activation of a uPA promoter-CAT reporter gene. Treating the pancreatic tumor cell lines with the known NF-kappaB inhibitors, dexamethasone and n-tosylphenyalanine chloromethyl ketone (TPCK), abolishes constitutive RelA activity and uPA overexpression. These results show that uPA is one of the downstream target genes induced by constitutively activated RelA in human pancreatic tumor cells, and suggests that constitutive RelA activity may play a critical role in tumor invasion and metastasis. Inhibition of constitutive RelA in pancreatic tumor cells may reduce their invasive and metastatic potential.
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PMID:Overexpression of urokinase-type plasminogen activator in pancreatic adenocarcinoma is regulated by constitutively activated RelA. 1046


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