UNIPROT:P00750 - FACTA Search

Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P00750 (PLA)
16,800 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Single-chain urokinase (scuPA), the unique form of urokinase secreted by cells, is converted to an active two-chain molecule through the cleavage of a single peptide bond by plasmin and other specific proteinases. Although scuPA may express limited enzymatic activity, its contribution to plasminogen activation on cell surfaces remains uncertain. Further, although it is well known that scuPA binds to a specific extracellular urokinase-type plasminogen activator receptor, the effect of this interaction on the enzymatic activity of scuPA has not been described. In the present paper we report that the binding of scuPA to cellular an to recombinant soluble urokinase-type plasminogen activator receptors (suPAR) increases its catalytic activity as measured by the cleavage of a urokinase-specific chromogenic substrate. suPAR increased the Vmax of scuPA 5-fold with little change in its Km. suPAR also stimulated the plasminogen activator activity of scuPA by decreasing its Km for Glu-plasminogen from 1.15 microM to 0.022 microM and by increasing the kcat of this reaction from 0.0015 to 0.022 s-1. Preincubation of scuPA with suPAR also enhances its susceptibility to inhibition by plasminogen activator inhibitor type 1, consistent with exposure of its catalytic site. The activity of scuPA bound to suPAR is not accompanied by cleavage of scuPA, which continues to migrate as a single band in SDS-polyacrylamide gel electrophoresis under reducing conditions. Moreover, suPAR increases the plasminogen activator activity of a plasmin-insensitive variant, scuPA (scuPA-Glu158), as well. Enhancement of scuPA activity by suPAR is both prevented and reversed by its aminoterminal fragment (amino acids 1-135), which competes for receptor binding, suggesting that continued binding to the receptor is required for expression of scuPA's enzymatic activity. Thus, our data suggest that scuPA may undergo a reversible transformation between a latent and an active state. The urokinase receptor may induce or stabilize scuPA in its active conformation, thereby contributing to the initiation of plasminogen activation on cell surfaces.
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PMID:Enhancement of the enzymatic activity of single-chain urokinase plasminogen activator by soluble urokinase receptor. 761 42

Altered coronary artery expression of plasminogen activator (PA) system components may predispose to thrombosis and modulate the vascular response to injury. By immunohistochemistry, we studied the expression of PAs (tPA and uPA), their major physiological inhibitor (PAI-1), and a receptor for uPA (uPAR) in human coronary arteries with either pure fibrointimal proliferation (n = 15) or developed atherosclerotic plaques (n = 10). Overall, the degree of staining showed the following rank order: PAI-1 > tPA > uPAR > uPA. A similar pattern was seen in two normal coronary arteries. There were no significant differences in the extent of staining in any vascular compartment between atherosclerotic arteries and those with only fibrointimal proliferation. However, the ratio of intimal to medial expression of tPA (P = .001) and uPAR (P = .004) was significantly increased in atherosclerotic arteries, with a similar trend for uPA (P = .069) but not for PAI-1 (P = .73). Four of 10 atherosclerotic arteries had higher uPAR expression in the intima than in the media, whereas none of the 15 arteries with only fibrointimal proliferation had this pattern (P < .01). Dual labeling studies demonstrated colocalization of all four PA system components in endothelial cells, smooth muscle cells, and macrophages, with a predominance of PAI-1. Thus, coronary arteries with a wide range of vascular pathology express an abundance of antifibrinolytic potential with enhanced local expression of profibrinolytic proteins, mainly within atherosclerotic plaques.
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PMID:Plasminogen activator system in human coronary atherosclerosis. 767 Sep 59

Progesterone stimulates differentiation and inhibits the growth of endometrial tissue. Also, progesterone reduces plasminogen activator (PA) activity, which implies reduced turnover of extracellular matrix proteins in the secretory phase. To elucidate the mechanism responsible for reduced PA activity, primary cultures of human endometrial stromal cells were stimulated with estradiol and progesterone. Conditioned media were assayed for urokinase-type and tissue-type PA (u-PA and t-PA, respectively), PA inhibitor-1 (PAI-1), and PA activity. Binding of [125I]u-PA and [125I]u-PA:PAI-1 complex to the u-PA receptor and clearance of these ligands were studied. The PA activity of conditioned medium decreased after stimulation with progesterone, and this was secondary to a decrease in u-PA, but not t-PA, and an increase in PAI-1. Northern blot analysis showed induction of PAI-1 messenger ribonucleic acid, whereas the content of u-PA messenger ribonucleic acid was not influenced. Furthermore, the number of free u-PA receptor-binding sites was increased by estradiol and progesterone. The stromal cells degraded complexed u-PA more efficiently than free u-PA, and degradation of both ligands was inhibited by colchicine, chloroquine, and methylamine. Degradation was increased after hormone treatment, and this was apparently due to increased ligand binding, because neither ligand affinity nor the relative rate of degradation was increased. Increased expression of u-PA receptor-binding sites was not regulated on the transcriptional level, but may result from posttranslational mechanisms, such as decreased turnover of the receptor. Activation of plasminogen by receptor bound u-PA initiates a cascade of proteolytic events in the extracellular matrix that is important during tissue proliferation. Our data suggest that differentiated endometrial stroma in the secretory phase regulates extracellular proteolysis by increased elimination of u-PA through increased release of PAI-1 and increased u-PA receptor density.
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PMID:Progesterone stimulates degradation of urokinase plasminogen activator (u-PA) in endometrial stromal cells by increasing its inhibitor and surface expression of the u-PA receptor. 767 23

Using immunohistochemistry and in-situ hybridization, we studied the expression of the components of the plasminogen activation system during progression to malignant melanoma with fresh melanocytic lesions. Expression of these components is confined to late stages of melanoma. t-PA expression is limited to rare cases of metastatic melanoma. The other components are frequently expressed concomitantly in the same tumour. Urokinase (u-PA) is expressed in stromal cells and only in tumour cells at invasive foci, urokinase receptor (u-PAR) in tumour cells, plasminogen activator inhibitor type I (PAI-1) in the intratumoral extracellular matrix and plasminogen activator inhibitor type II (PAI-2) in tumour cells and stromal cells. In order to investigate the role of u-PAR as a prognostic marker, we have developed an assay for quantitation of the receptor. As a first step towards structural investigations, we have determined the disulfide cross-links of the first domain of uPAR.
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PMID:Studies on functional and structural role of urokinase receptor and other components of the plasminogen activation system in malignancy. 774 86

Urokinase plasminogen activator (uPA) is a serine protease involved in cancer invasion and metastasis. uPA mediates its action while attached to a membrane-bound receptor (uPAR). In this investigation we show that uPAR levels correlate with uPA levels in human breast cancers. uPAR levels, however, do not correlate with other components of the plasminogen activator system such as tissue-type plasminogen activator (t-PA), PAI-I or PAI-2. In addition, uPAR levels showed no correlation with tumor size, axillary-node status or estrogen-receptor status. On the basis of an optimum cut-off point, patients with breast cancers containing high levels of uPAR had a worse prognosis than patients with low levels of the receptor. However, as a prognostic marker in breast cancer, uPAR was not as strong as uPA. Our results are consistent with data from model systems suggesting that both uPA and uPAR are necessary for metastasis.
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PMID:Urokinase plasminogen activator and urokinase plasminogen activator receptor in breast cancer. 776 29

The plasminogen activator (PA)/plasmin system is thought to be involved in processes such as tumor invasion and wound healing, during which epithelial and mesenchymal cells come close together. However, information on regulation of the PA/plasmin system during epithelial-mesenchymal interactions is scarce. Therefore, we examined the in vitro modulation of the production and activity of the components of the PA/plasmin system in squamous carcinoma cells (SCC-4) and normal human keratinocytes in relation to cell density and the presence or absence of fibroblasts (3T3 cells). There was an inverse relation between cell density and mRNA expression for urokinase-type plasminogen activator (u-PA) and u-PA receptor in both SCC-4 cells and keratinocytes. In addition, such a relation was found for plasminogen activator inhibitor types 1 (PAI-1) and 2 (PAI-2) in SCC-4 monocultures, but not in keratinocyte monocultures. In contrast to monocultures, variation of cell density did not affect the mRNA expression of the components of the PA/plasmin system in cocultures of SCC-4 cells or keratinocytes with 3T3 cells. However, the relative expression of mRNAs in co-cultures was clearly different from that in monocultures, especially at low cell density. For most of the components of the PA/plasmin system, a decrease in mRNA expression and u-PA receptor protein was observed at most cell densities, whereas for PAI-1 only in keratinocytes a marked increase was documented. Zymography of supernatants revealed that the levels of both free u-PA and PA-PAI were increased in SCC-4/3T3 co-cultures, whereas in keratinocytes/3T3 co-cultures, only levels of the PA-PAI complex were increased, while the amount of free u-PA activity decreased. This occurred despite the increase u-PA immunoreactivity and was probably caused by the markedly elevated levels of immunoreactive PAI-1. The results of the present study reveal that the production and synthesis of various components of the PA/plasmin system in keratinocytes and SCC-4 cells depend on the density of epithelial cells and are modulated by fibroblasts, probably through a direct cell-cell or cell-matrix contact. Fibroblast-induced modulations are similar in keratinocytes and SCC-4 cells except for the regulation of PAI-1, which is markedly enhanced only in keratinocytes. This suggests that the modulation of PA activity in the direct microenvironment may be different under physiologic and pathologic conditions.
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PMID:Differential regulation of plasminogen activation in normal keratinocytes and SCC-4 cells by fibroblasts. 786 Oct 5

Urokinase (u-PA)-mediated cell surface plasminogen activation is required for cellular tissue invasion. This invasion occurs in environments rich in plasminogen activator inhibitors (PAIs), which efficiently inhibit receptor-bound two-chain u-PA. Single-chain u-PA (scu-PA) was recently found to efficiently initiate cell surface plasminogen activation, and we herein describe the interaction of scu-PA with PAI type 2 (PAI-2). In the fluid phase (no cells) the plasminogen-activating activities of both scu-PA and Glu158-scu-PA (a plasmin non-activatable variant of scu-PA) were inhibited in a concentration-dependent manner by recombinant human PAI-2. This inhibition occurred with both forms of scu-PA remaining as single-chain molecules throughout the interactions. Although scu-PA did not form SDS-stable complexes with PAI-2, preincubation of scu-PA with 125I-PAI-2 demonstrated a dose-dependent inhibition of SDS-stable complex formation between 125I-PAI-2 and subsequently added two-chain u-PA. This indicates that although a "stable intermediate" type complex between scu-PA and PAI-2 was not detected, there was a physical association between the two molecules that shared at least some determinants with the two-chain u-PA-PAI-2 complex. In contrast, Glu158-scu-PA bound to u-PA receptors on monocytes was only minimally inhibited by a large molar excess of PAI-2. These data suggest that the initiation of cell surface plasminogen activation may involve the partitioning of scu-PA between PAI-2 (a "negative modulator") and the u-PA receptor (a "positive modulator") and that the enzymatic activity of receptor-bound scu-PA may allow initiation of cell surface proteolysis even in PAI-2-rich environments. A model along these lines is presented.
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PMID:Differential inhibition of soluble and cell surface receptor-bound single-chain urokinase by plasminogen activator inhibitor type 2. A potential regulatory mechanism. 790 91

Concomitant expression of urokinase type plasminogen activator (uPA) and its surface receptor (uPAR) has been shown to correlate strongly with a more invasive tumor cell phenotype. A highly malignant human epidermoid carcinoma cell line (HEp3) was transfected with a vector capable of expressing an antisense transcript complementary to 300 bases of the 5' end of uPAR, including the ATG codon. Six stably transfected antisense (AS-2, 3, 5, 9, 10, 12) and eight control clones were characterized. All clones produced high levels of uPA activity. Examination of collagenase production and doubling time showed that all of the clones tested produced similar activities. The antisense clones showed a 20-74% reduction in the uPAR sites; the uPAR mRNA level was also reduced. A test of the invasive ability of all clones in a modified chorioallantoic membrane (CAM) showed that invasiveness of the antisense-inhibited clones was directly proportional to the density of surface uPAR. The AS-2 clone, which expressed the lowest number of uPARs showed a significantly reduced level of invasion. The invasiveness of additional AS-inhibited clones was also reduced. Seven control and four AS-inhibited clones were tested for tumorigenicity on CAMs of chick embryos. Inoculation of control cells produced large tumors, while the As clones were non-tumorigenic. AS-2 did not produce tumors even if kept in vivo for up to 10 weeks.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:The effect of antisense inhibition of urokinase receptor in human squamous cell carcinoma on malignancy. 807 94

Several functional defects have been found in neutrophils from leukocyte adhesion deficiency (LAD) patients who fail to express the CD11/CD18 leukoadhesins: Mo1, LFA-1, and p150,95. To better understand the functional defects of LAD neutrophils, we have performed capping experiments. Purified normal or LAD neutrophils were labeled with fluorochrome-conjugated concanavalin A (Con A) or F(ab')2 fragments of antiurokinase-type plasminogen activator receptor (uPAR), anti-Fc gamma RIII (CD16), anti-Mo5, and anti-CD14 antibodies. F(ab')2-labeled cells were capped using a second-step F(ab')2 fragment of an antimurine Fab antiserum. Cells were capped for 30 minutes at 37 degrees C, then observed by fluorescence microscopy. LAD neutrophils were found to be deficient in capping, but not clustering of all of the reagents tested to date. The percent of cells exhibiting capping of Con A, Fc gamma RIII, urokinase receptor, CD14, and Mo5 were 52%, 67%, 70%, 25%, and 64% for normal neutrophils but were only 10%, 5%, 2%, 3%, and 1%, respectively, for LAD neutrophils. Capping of this panel of membrane components in LAD or normal neutrophils was not augmented by the addition of either 10(-5) mol/L colchicine or 10(-7) mol/L FMLP. Because capping requires membrane-to-cytosol communication and an intact microfilament linkage, we suggest that leukoadhesins may play a broad role in promoting the redistribution of membrane components including adherence-related receptors such as Fc gamma RIII and the urokinase receptor.
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PMID:Aberrant capping of membrane proteins on neutrophils from patients with leukocyte adhesion deficiency. 812 56

Fibrinolytic properties of four hybrids of u-PA and t-PA, all containing the u-PA growth factor domain and binding to recombinant human u-PA receptor expressed in CHO cells, were compared. Highest fibrin stimulation was observed with uK2tPA which when compared to t-PA in the rabbit system, had a considerably prolonged circulatory half-life in vivo. Compared to an equimolar dose of t-PA, 0.4 mg/kg uK2tPA caused a similar consumption of alpha 2-antiplasmin and fibrinogen and a considerably greater prolongation of the ex-vivo blood clotting time. Nevertheless, this dose of uK2tPA was inactive in the jugular vein thrombosis assay. This lack of thrombolytic activity is presumably due to the presence of a functional u-PA growth factor domain, which in binding uK2tPA to cellular blood elements possibly retards its penetration into the blood clot and in this manner could neutralize the potential thrombolytic activity of the t-PA kringle 2 and protease domains in uK2tPA.
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PMID:A hybrid plasminogen activator binds to the u-PA receptor and has a reduced thrombolytic potency in vivo. 838 59

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