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)

Clinical and experimental data indicate that activated oxygen species interfere with vascular endothelial cell function. Here, the impact of extracellular oxidant injury on the fibrinolytic response of cultured human umbilical vein endothelial (HUVE) cells was investigated at the protein and mRNA levels. Xanthine (50 microM) and xanthine oxidase (100 milliunits), which produces the superoxide anion radical (O2-) and hydrogen peroxide (H2O2), was used to sublethally injure HUVE cells. Following a 15-min exposure, washed cells were incubated for up to 24 h in serum-free culture medium. Tissue-type plasminogen activator (t-PA) antigen, plasminogen activator inhibitor-1 (PAI-1) antigen, and PAI-1 activity were determined in 1.25 ml of conditioned medium and t-PA and PAI-1 mRNA in the cell extracts of 2 x 10(6) HUVE cells. Control cells secreted 3.9 +/- 1.3 ng/ml (mean +/- S.D., n = 12) within 24 h. Treatment with xanthine/xanthine oxidase for 15 min induced a 2.8 +/- 0.4-fold increase (n = 12, p less than 0.05) of t-PA antigen secretion after 24 h. The t-PA antigen was recovered predominantly in complex with PAI-1. The oxidant injury caused a 3.0 +/- 0.8-fold increase (n = 9, p less than 0.05) in t-PA mRNA within 2 h. Total protein synthesis was unaltered by xanthine/xanthine oxidase. The oxidant scavengers superoxide dismutase and catalase, in combination, abolished the effect of xanthine/xanthine oxidase on t-PA secretion and t-PA mRNA synthesis. Xanthine/xanthine oxidase treatment of HUVE cells did not affect the PAI-1 secretion in conditioned medium nor the PAI-1 mRNA levels in cell extracts. Thus extracellular oxidant injury induces t-PA but not PAI-1 synthesis in HUVE cells.
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PMID:Modulation of the fibrinolytic response of cultured human vascular endothelium by extracellularly generated oxygen radicals. 173 Jun 19

In conditioned medium (CM) from cultured human endothelial cells, two forms of plasminogen-activator inhibitor (PA-inhibitor) can be demonstrated: a fast-acting active form and an immunologically related, inactive form. Evidence is presented that endothelial cells produce active PA-inhibitor which is rapidly inactivated upon secretion into the medium. This inactivation can, at least partly, be prevented by culturing cells with excess of tissue-type plasminogen activator (t-PA). This results in the formation of large amounts of t-PA-PA-inhibitor complex at the cost of accumulation of inactive PA-inhibitor. No complex was detectable when inactive PA-inhibitor preparations were incubated with t-PA either in the absence or in the presence of cells. Furthermore, in cell extracts, predominantly functionally active PA-inhibitor was present. PA-inhibitor derived from the t-PA-PA-inhibitor complex showed an Mr approx. 4000 lower by polyacrylamide-gel electrophoresis than that of the inactive form. The rapid inactivation seems to be confined to newly synthesized molecules, since PA-inhibitor molecules in CM are inactivated much more slowly (even with cells or cell homogenates) than necessary to explain the excessive production of inactivated PA-inhibitor by cells. It could not be prevented by inhibitors of oxidative processes, like butylated hydroxytoluene, dithiothreitol, superoxide dismutase and catalase.
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PMID:Rapid inactivation of the plasminogen-activator inhibitor upon secretion from cultured human endothelial cells. 310 1

Each of 11 tumors tested produced a factor that markedly suppressed the ability of macrophages to release H2O2 or O.2- in response to phorbol myristate acetate or zymosan. Four of seven normal cell types produced a similar activity, which was 3.5-7 times lower in titer than that in tumor cell-conditioned medium (TCM), and which was much more rapidly reversible in its effects. TCM caused 50% inhibition of H2O2 release when it was present in the medium for 48 h at a concentration of 13%, or when 100% TCM was present in the medium for 18 h. The H2O2-releasing capacity of macrophages incubated in TCM only returned to control levels by 6 d after its removal. TCM prevented augmentation of H2O2-releasing capacity by lymphokines. The titer of suppressive activity in TCM depended on both the concentration of tumor cells and the duration of their incubation. TCM did not augment the activity of catalase, myeloperoxidase, glutathione peroxidase, or glutathione reductase or the content of glutathione within macrophages, suggesting that decreased synthesis rather than increased catabolism was responsible for reduced secretion of H2O2. Suppression of the release of H2O2 or O.2- by TCM appeared to be a relatively specific effect, in that TCM increased macrophage spreading and adherence to glass while exerting little influence on rates of phagocytosis, synthesis of protein, or secretion of lysozyme, plasminogen activator, or arachidonic acid and its metabolites. Thus, tumor cells and some normal cells can secrete a factor that selectively deactivates macrophage oxidative metabolism.
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PMID:Suppression of macrophage oxidative metabolism by products of malignant and nonmalignant cells. 715 14

The first defined sequential epitope of the tissue plasminogen activator (t-PA) was determined by a monoclonal antibody against a synthetic peptide segment corresponding to peptide sequence 341-354 of t-PA. This segment was selected by computer assisted epitope prediction. Balb/c mice were immunized with catalase-peptide and tripalmitoyl-S-glyceryl-cys-teinyl-seryl-peptide conjugates. A monoclonal antibody derived from this immunization was reactive with native recombinant t-PA (rt-PA) and reduced carboxymethylated recombinant t-PA (RCM rt-PA). The sequential epitope was detected by Pepscan method using overlapping octa- and nonapeptides. By fine epitope mapping with tetra-, penta-, hexa- and heptapeptides the epitope was minimized to the pentapeptide EEEQK (347-351). Replacement set analysis confirmed the importance of this amino acid sequence, especially of the amino acid E348, for antibody binding. Functional assays of rt-PA were not affected by this antibody indicating that the epitope has no influence on the enzymatic center and the binding site of the inhibitor. The analysis demonstrates that the predicted recognition site of the monoclonal antibody 17-134/11 is exposed on the surface of the native rt-PA molecule.
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PMID:Pentapeptide identified as a monoclonal antibody binding site in the serine-protease domain of t-PA. 752 35

The effects of the E1A protein of adenovirus-5 on the differentiation program of F9 teratocarcinoma cells were examined by the stable introduction of plasmids that expressed wild-type or mutated forms of E1A. Constitutive expression of plasmids for most of the mutant E1As induced loss of expression of the cell-surface antigen SSEA-1 and the enhanced expression of genes specific for the differentiated phenotype of F9 cells, such as genes for laminin B1, tissue-type plasminogen activator (tPA) and type IV collagen, as well as the altered cell morphology that is associated with the differentiated state. However, such changes were not observed in the case of genes for mutant proteins from which a conserved region (CR1) of E1A had been deleted. Furthermore, no significant induction of expression of the c-jun gene or transactivation of the c-jun-CAT reporter gene were observed when the sequence that encodes CR1 of E1A had been deleted. A palindromic sequence element (DRE) of the c-jun promoter was essential for the E1A-mediated up-regulation of the c-jun gene. These results imply that CR1 is required for activation of the c-jun gene and that it is implicated in the growth arrest, expression of parietal endoderm-specific functions and the orderly differentiation of F9 cells.
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PMID:Induction by adenovirus-5 E1A of the differentiation phenotype of F9 teratocarcinoma cells involves a conserved region (CR1) of E1A. 774 80

It is established that the medically significant yersiniae require the presence of physiological levels of Ca2+ (ca. 2.5 mM) for sustained growth at 37 degrees C and that this nutritional requirement is mediated by a shared ca. 70-kb Lcr plasmid. The latter also encodes virulence factors (Yersinia outer membrane proteins [Yops] and V antigen) known to be selectively synthesized in vitro at 37 degrees C in Ca(2+)-deficient medium. In this study, cells of Yersinia pestis KIM were first starved for Ca2+ at 37 degrees C to prevent synthesis of bulk vegetative protein and then, after cell division had ceased, pulsed with [35S]methionine. After sufficient chase to ensure plasminogen activator-mediated degradation of Yops, the remaining major radioactive peptides were separated by conventional chromatographic methods and identified as Lcr plasmid-encoded V antigen and LcrH (and possibly LcrG), ca. 10-kb Pst plasmid-encoded pesticin and plasminogen activator, ca. 100-kb Tox plasmid-encoded fraction 1 (capsular) antigen and murine exotoxin, and chromosomally encoded antigen 4 (pH 6 antigen) and antigen 5 (a novel hemin-rich peptide possessing modest catalase activity but not superoxide dismutase activity). Also produced at high concentration was a chromosome-encoded GroEL-like chaperone protein. Accordingly, the transcriptional block preventing synthesis of bulk vegetative protein at 37 degrees C in Ca(2+)-deficient medium may not apply to genes encoding virulence factors or to highly conserved GroEL (known in other species to utilize a secondary stress-induced sigma factor).
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PMID:Major stable peptides of Yersinia pestis synthesized during the low-calcium response. 841 35

The first temperature-dependent proteins (expressed at 37 degrees C, but not 26 degrees C) to be identified in Yersinia pestis were antigens 3 (fraction 1), 4 (pH 6 antigen), and 5 (hereafter termed KatY). Antigens 3 and 4 are now established virulence factors, whereas little is known about KatY, except that it is encoded chromosomally, produced in abundance, possesses modest catalase activity, and is shared by Yersinia pseudotuberculosis, but not Yersinia enterocolitica. We report here an improved chromatographic method (DEAE-cellulose, calcium hydroxylapatite, and Sephadex G-150) that yields enzymatically active KatY (2,423 U/mg of protein). Corresponding mouse monoclonal antibody 1B70.1 detected plasminogen activator-mediated hydrolysis of KatY, and a polyclonal rabbit antiserum raised against outer membranes of Y. pestis was enriched for anti-KatY. A sequenced approximately 16-kb Y. pestis DNA insert of a positive pLG338 clone indicated that katY encodes an 81.4-kDa protein (pI 6.98) containing a leader sequence of 2.6 kDa; the deduced molecular mass and pI of processed KatY were 78.8 kDa and 6. 43, respectively. A minor truncated variant (predicted molecular mass of 53.6 kDa) was also expressed. KatY is similar (39 to 59% identity) to vegetative bacterial catalase-peroxidases (KatG in Escherichia coli) and is closely related to plasmid-encoded KatP of enterohemorrhagic E. coli O157:H7 (75% identity). katY encoded a putative Ca2+-binding site, and its promoter contained three homologues to the consensus recognition sequence of the pCD-encoded transcriptional activator LcrF. rbsA was located upstream of katY, and cybB, cybC, dmsABC, and araD were mapped downstream. These genes are not linked to katG or katP in E. coli.
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PMID:Molecular characterization of KatY (antigen 5), a thermoregulated chromosomally encoded catalase-peroxidase of Yersinia pestis. 1032 12

Phospholipases A(2) (PLA(2)s) catalyzing the hydrolysis of phospholipids form a family of proteins with diverse physiological and pharmacological properties. While there have been several reports on the cloning of PLA(2) cDNAs, very few studies have been carried out on the PLA(2) genes and, most importantly, no information has been available on the gene structure and function of group I venom PLA(2). This study, on the PLA(2) gene from a spitting cobra, besides being the very first report on any venom group I PLA(2) gene, constitutes the missing link in the biology and evolution of phospholipases. The 4-kb gene consists of four exons and three introns and resembles the human pancreatic PLA(2) gene. However, the size of intron 3 in particular is much smaller than that in the pancreatic gene. Interestingly, the information for the toxic and most of the pharmacological properties of the venom PLA(2) can be attributed to the end of exon 3 and the whole of exon 4 of the gene. This functional delineation fits in well with the theory of adaptive evolution exhibited by the venom PLA(2)s. We also show that the mammalian pancreatic and elapid PLA(2)s have similar paths of evolution (probably following gene duplication) from a common ancestral gene. Venom group II phospholipases, although evolved from the same ancestor, diverged early in evolution from the group I PLA(2) genes. Intriguingly, CAT reporter gene assays and DNase 1 footprinting studies on the promoter and its deletion constructs using CHO and HepG2 cell lines identified the possible involvement of cis elements such as Sp1, AP2, gamma-IRE, and (TG)(12) repeats in the expression of the gene in a tissue-specific manner.
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PMID:Structure and phylogeny of the venom group I phospholipase A(2) gene. 1088 14

Extracellular matrix (ECM) remodeling is essential for normal development and tissue repair. Although many roles for extracellular proteinases in the breakdown of ECM have been established, the regulations of these proteinases in human tissue are not fully understood. Inflammatory cytokines have been implicated in the regulation of several matrix metalloproteinases. To determine whether these mediators have a similar effect on fibrinolysis and the remodeling of the fibrin provisional matrix, we examined the role of cytokines on the regulation of urokinase-type plasminogen activator (uPA) and tissue-type plasminogen activator (tPA) in human skin. In this report, we show that interleukin-8 (IL-8), but not other cytokines tested, is a potent inducer of the 38-kDa uPA in organ-cultured human skin. In addition, the uPA inhibitor, PAI-1, was not affected by IL-8. When primary epidermal human keratinocytes were treated with IL-8, 55-kDa pro-uPA was significantly induced in the conditioned medium. The mRNA expression of uPA in the keratinocytes was found to be constitutively elevated and was not affected by IL-8. To support such a notion, activation of the 5'-flanking promoter of the human uPA gene was measured using the CAT reporter assay. Consistent with the results of mRNA measurement, the promoter is constitutively active in keratinocytes and is not affected by IL-8. In contrast, the promoter construct is neither active in the dermal fibroblasts nor stimulated by the cytokine. This differential transactivation of uPA gene in these cells indicates that keratinocyte-specific factors may govern the basal expression of the gene. These results indicate a complex regulation of uPA expression in epidermal cells.
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PMID:IL-8-stimulated expression of urokinase-type plasminogen activator in human skin and human epidermal cells. 1217 88

Several microbial pathogens augment their invasive potential by binding and activating human plasminogen to generate the proteolytic enzyme plasmin. Yeast cells and cell wall proteins (CWP) of the human pathogenic fungus Candida albicans bound plasminogen with a K(d) of 70 +/- 11 nM and 112 +/- 20 nM respectively. Bound plasminogen could be activated to plasmin by mammalian plasminogen activators; no C. albicans plasminogen activator was detected. Binding of plasminogen to CWP and whole cells was inhibited by epsilon ACA, indicating that binding was predominantly to lysine residues. Candida albicans mutant strains defective in protein glycosylation did not show altered plasminogen binding, suggesting that binding was not mediated via a surface lectin. Binding was sensitive to digestion by basic carboxypeptidase, implicating C-terminal lysine residues in binding. Proteomic analysis identified eight major plasminogen-binding proteins in isolated CWP. Five of these (phosphoglycerate mutase, alcohol dehydrogenase, thioredoxin peroxidase, catalase, transcription elongation factor) had C-terminal lysine residues and three (glyceraldehyde-3-phosphate dehydrogenase, phosphoglycerate kinase and fructose bisphosphate aldolase) did not. Activation of plasminogen could potentially increase the capacity of this pathogenic fungus for tissue invasion and necrosis. Although surface-bound plasmin(ogen) degraded fibrin, no direct evidence for a role in invasion of endothelial matrix or in penetration and damage of endothelial cells was found.
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PMID:Candida albicans binds human plasminogen: identification of eight plasminogen-binding proteins. 1262 18


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