Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.4.3 (phospholipase C)
18,461 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Evidence for pleiotropic activation of virulence genes in Listeria monocytogenes is presented. A complementation study of a spontaneous prfA-deletion mutant and analysis of cassette and transposon insertion mutants showed that the gene prfA activates the transcription of four independent genes which code for a phosphatidyl-inositol-specific phospholipase C (gene plcA), listeriolysin O (gene hlyA), a metallo-protease (gene prtA) and a lecithinase (gene prtC). Transcription of prfA is not constitutive. During the growth phase, two peaks of prfA transcript accumulation were observed: the first was during exponential growth, and the second was at the beginning of the stationary phase. In addition, two prfA-specific transcripts of 2.2 kb and 1 kb are detected. Early in exponential growth, prfA is co-transcribed with plcA which lies upstream prfA, giving rise to the 2.2 kb plcA-prfA transcript. In late-exponential growth and at the beginning of the stationary phase, prfA transcripts of 1 kb are predominantly detected. Our results demonstrate that since prfA controls plcA transcription, it also regulates its own synthesis.
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PMID:Pleiotropic control of Listeria monocytogenes virulence factors by a gene that is autoregulated. 166 63

A nonisotopic enzymolysis assay method of phosphatidylinositol kinase (PIK) has been developed. Phosphatidylinositol 4-phosphate (PIP), phosphorylated from phosphatidylinositol (PI) by PIK was hydrolyzed with phosphainositide-specific phospholipase C. The product, inositol 1, 4-bisphosphate (IP2), was separated from inositol 1-phosphate (IP) with Dowex-1 column chromatography. Then, the IP2 was hydrolyzed by alkaline phosphatase to yield PI, and the PI was determined by colorimetry. The PIK activity was defined as PI nmol/mg protein per min. The recovery was 91%, CV = 6.5%
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PMID:[A nonisotopic enzymolysis assay method of phosphatidylinositol kinase]. 166 83

In pancreatic islets the bulk of phosphoinositide-specific phospholipase C (PI-PLC) activity was cytosolic. The soluble enzyme was activated by submicromolar concentrations of Ca2+, independent of calmodulin. It was unaffected by glucose and a series of glycolytic intermediates, including glyceraldehyde 3-phosphate. These observations lend support to the hypothesis that glucose-stimulated inositol triphosphate production in islets may be secondary to and provoked by glucose-mediated Ca2+ influx. All four pyridine nucleotides stimulated PI-PLC. Phosphatidylinositol hydrolysis was also stimulated by dioleine and arachidonic acid, and by the polyamines, putrescine and spermine. Phosphatidylinositol hydrolysis was inhibited by chlorpromazine, tetracaine, ATP, 5'-AMP, inorganic pyrophosphate and by phosphatidylinositol 4,5-bisphosphate, phosphatidylcholine and phosphatidylserine--but not affected by phosphatidylethanolamine. The cyclic nucleotides, cAMP and cGMP had no effect on the enzyme, and GTP-gamma-S did not activate the enzyme event at very low Ca2+ concentrations. The diglyceride lipase inhibitor, RHC 80267, and the cyclooxygenase inhibitor, indomethacin, had no effect on PI-PLC activity.
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PMID:Characteristics of phosphoinositide-specific phospholipase C activity from mouse pancreatic islets. 166 77

1. Phosphatidylinositol 4-phosphate (PtdIns4P) is degraded by isolated membranes from Xenopus laevis oocytes. 2. Incubation of [4-32P]PtdIns4P with membranes yields only radioactive inorganic phosphate, indicating the presence of a phosphomonoesterase. 3. Membranes hydrolyze Ptd[2-3H]Ins4P to produce mainly Ptd[2-3H]Ins in the lipid phase. In this incubation [3H]inositol and inositol monophosphate appear in the water phase. 4. Membrane incubations of Ptd[2-3H]Ins4P carried out in the presence of excess non-radioactive Ins(1,4)P2 allows the trapping of small amounts of [3H]Ins(1,4)P2. These results demonstrate the presence of a phospholipase C. 5. Testing several phosphorylated analogs, it is determined that fructose 1,6-bisphosphate and alpha-glycerophosphate are potent inhibitors of the oocyte PtdIns4P phosphomonoesterase.
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PMID:The hydrolysis of phosphatidylinositol 4-phosphate in membranes of Xenopus laevis oocytes: characteristics of a phosphomonoesterase. 166 8

The respective roles of urokinase plasminogen activator (u-PA) and the u-PA receptor in extracellular matrix degradation was investigated. Human pro-u-PA and the human u-PA receptor were expressed independently by two different mouse LB6 cell lines. The matrix degradation capacity of these cell lines individually or in coculture was studied. Although pro-u-PA-producing cells alone degrade the matrix in the presence of plasminogen, u-PA-receptor producing cells do not. Cocultivation of a small fraction of pro-u-PA-producing cells with the receptor-producing cells increases the rate of matrix degradation at least threefold. By immunoprecipitation it was shown that cocultivation of the two cell lines increases the conversion of the inactive pro-u-PA to the active two chain u-PA. The enhancement of matrix degradation and of pro-u-PA activation requires actual binding of pro-u-PA to its receptor because it is inhibited by u-PA-receptor antagonists. The u-PA receptor must be cell associated, as binding of pro-u-PA to a receptor solubilized from the cell surface with phosphatidyl-inositol specific phospholipase C did not enhance the activation of pro-u-PA in the presence of plasminogen. The finding that activity of u-PA is enhanced when it is bound to its receptor, even when the receptor is produced by a different cell, might have important implications for the mechanisms of u-PA-induced extracellular proteolysis in vivo.
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PMID:Complementation between urokinase-producing and receptor-producing cells in extracellular matrix degradation. 166 3

A human E membrane protein that inhibits lysis by the purified human C5b-9 proteins was isolated and characterized. After final purification, the protein migrated as an 18- to 20-kDa band by SDS-PAGE. Elution from gel slices and functional assay after SDS-PAGE (nonreduced) confirmed that all C5b-9 inhibitory activity of the purified protein resided in the 18- to 20-kDa band. Phosphatidylinositol-specific phospholipase C digestion of the purified protein abolished 50% of its C5b-9 inhibitory activity, and removed approximately 15% of the protein from human E. Western blots of normal and paroxysmal nocturnal hemoglobinuria E revealed an absence of the 18- to 20-kDa protein in the paroxysmal nocturnal hemoglobinuria E cells. The identity of this E protein with leukocyte Ag CD59 (P18, HRF20) was confirmed immunochemically and by N-terminal amino acid sequence analysis. A blocking antibody raised against the purified protein reacted with a single 18- to 20-kDa band on Western blots of human erythrocyte membranes. Prior incubation of human E with the F(ab) of this antibody increased subsequent lysis by the purified human C5b-9 proteins. Potentiation of C5b-9-mediated lysis was observed when erythrocytes were preincubated with this blocking antibody before C5b-9 assembly was initiated, or, when this antibody was added after 30 min, 0 degrees C incubation of C5b-8-treated E with C9. Chicken E incubated with purified CD59 were used to further characterize the mechanism of its C-inhibitory activity. Preincorporation of CD59 into these cells inhibited lysis by C5b-9, regardless of whether CD59 was added before or after assembly of the C5b-8 complex. When incorporated into the membrane, CD59 inhibited binding of 125I-C9 to membrane C5b-8 and reduced the extent of formation of SDS-resistant C9 polymer. The inhibitory effect of CD59 on 125I-C9 incorporation was most pronounced at near-saturating input of C9 (to C5b-8). By contrast, CD59 did not inhibit either C5b67 deposition onto the cell surface, or, binding of 125I-C8 to preassembled membrane C5b67. Taken together, these data suggest that CD59 exerts its C-inhibitory activity by limiting incorporation of multiple C9 into the membrane C5b-9 complex.
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PMID:The complement-inhibitory activity of CD59 resides in its capacity to block incorporation of C9 into membrane C5b-9. 169 60

Phosphatidylinositol-specific phospholipase C (PI-PLC) is an enzyme that has the capacity to release glycosyl-phosphatidyl inositol (G-PI)-anchored proteins from the cells surface. Pretreatment of the human T-cell leukemia cell line Molt-4 with PI-PLC resulted in a decrease in the susceptibility to lysis by natural killer (NK) cells. Treatment of the erythroleukemia cell line K562 with PI-PLC had no effect on its NK susceptibility. PI-PLC-treated and untreated Molt-4 bound equally well to lymphocytes in target-binding studies with effector cell preparations enriched for NK cells. Susceptibility to cytolytic granules isolated from rat LGL tumor cells remained the same after treatment of Molt-4 or K562 with PI-PLC. Combined treatment of Molt-4 with PI-PLC and rlFN-alpha or rlFN-gamma resulted in additive reductions of the NK susceptibility, suggesting that PI-PLC and interferons act on different mechanisms to protect cells from NK lysis. When expression of a number of antigens on Molt-4 and K562 was analyzed in flow cytometry, only the expression of CD58 was reduced after PI-PLC treatment. The susceptibility of Con A blasts to MLR derived cytotoxic T-cells was not altered by treatment with phospholipase. These data suggest that PI-PLC treatment reduces the capacity of some target cells to activate NK cells upon contract. The mechanism behind this phenomenon is presently unclear.
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PMID:Phospholipase C treatment of certain human target cells reduces their susceptibility to NK lysis without affecting binding or sensitivity to lytic granules. 170 25

Protection against the pore-forming activity of the human C5b-9 proteins was conferred on a nonprimate cell by transfection with cDNA encoding the human complement regulatory protein CD59. CD59 was stably expressed in Chinese hamster ovary cells using the pFRSV mammalian expression vector. After cloning and selection, the transfected cells were maintained in media containing various concentrations of methotrexate, which induced surface expression of up to 4.2 x 10(6) molecules of CD59/cell. Phosphatidylinositol-specific phospholipase C removed greater than 95% of surface-expressed CD59 antigen, confirming that recombinant CD59 was tethered to the Chinese hamster ovary plasma membrane by a lipid anchor. The recombinant protein exhibited an apparent molecular mass of 21-24 kDa (versus 18-21 kDa for human erythrocyte CD59). After N-glycanase digestion, recombinant and erythrocyte CD59 comigrated with apparent molecular masses of 12-14 kDa, suggesting altered structure of asparagine-linked carbohydrate in recombinant versus erythrocyte CD59. The function of the recombinant protein was evaluated by changes in the sensitivity of the CD59 transfectants to the pore-forming activity of human C5b-9. Induction of cell-surface expression of CD59 antigen inhibited C5b-9 pore formation in a dose-dependent fashion. CD59 transfectants expressing greater than or equal to 1.2 x 10(6) molecules of CD59/cell were completely resistant to human serum complement. By contrast, CD59 transfectants remained sensitive to the pore-forming activity of guinea pig C8 and C9 (bound to human C5b67). Functionally blocking antibody against erythrocyte CD59 abolished the human complement resistance observed for the CD59-transfected Chinese hamster ovary cells. These results confirm that the C5b-9 inhibitory function of the human erythrocyte membrane is provided by CD59 and suggest that the gene for this protein can be expressed in xenotypic cells to confer protection against human serum complement.
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PMID:Amplified gene expression in CD59-transfected Chinese hamster ovary cells confers protection against the membrane attack complex of human complement. 171 84

Phosphatidylinositol (PtdIns)-4- and -3-kinases, PtdIns(4)P-5-kinase, diacylglycerol (DAG) kinase, and PtdIns-phospholipase C were all detected in cytoskeletons of resting human platelets. The total cytoskeletal enzyme activities were greatly increased upon thrombin stimulation of the intact cells. Those reached a maximum after a 60-s stimulation for PtdIns(4)P-5-kinase and phospholipase C, while the other kinases appeared to be slightly delayed. Specific activities were stimulated from about 4-fold (PtdIns-3-kinase) to about 6-fold (PtdIns-4-kinase). Thrombin treatment also promoted a co-extraction of pp60c-src with the cytoskeletons and its disappearance from the Triton X-100 soluble fraction. These results suggest that stimulation of platelets by thrombin causes the association of enzymes responsible for lipid phosphorylation and hydrolysis with the cytoskeletons. This could occur at cytoskeleton anchoring points to the membranes.
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PMID:Interaction of pp60c-src, phospholipase C, inositol-lipid, and diacyglycerol kinases with the cytoskeletons of thrombin-stimulated platelets. 171 96

The antigen detected by monoclonal antibodies reacting with human osteosarcoma-associated antigen was shown to be a phosphatidyl-inositol (PI)-glycan-anchored protein, which can be released from the cell surface by PI-specific phospholipase C-treatment. The antigen detected by 2D3 and 2H10 antibodies exhibited alkaline phosphatase activity. Both antibodies strongly reacted with bone-type alkaline phosphatase. However, importantly, immunohistochemical analysis demonstrated that 2D3 and 2H10 did not react with alkaline phosphatase present in kidney or liver. In addition, neither placental nor intestinal alkaline phosphatase was recognized by 2D3 and 2H10 antibodies. These results indicated that two monoclonal antibodies, 2D3 and 2H10, are highly specific for bone-type alkaline phosphatase and can distinguish bone alkaline phosphatase from liver alkaline phosphatase in spite of the fact that liver and bone alkaline phosphatase are encoded by the same gene.
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PMID:Detection of bone-type alkaline phosphatase by monoclonal antibodies reacting with human osteosarcoma-associated antigen. 171 40


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