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)

Phosphatidylinositol (PI) specific phospholipase C (PIase C) treatment of human platelets caused release of a surface glycoprotein in the medium. Human blood platelets were isolated by low speed centrifugation and surface glycoproteins were labelled with periodate/[3H]borohydride procedure. Intact surface-labelled platelets were treated with PIase C purified from culture filtrates of Staphylococcus aureus (SA) or Bacillus thuringiensis (BT). After PIase C treatments platelets were spun at low speed, pellet and supernatant were separated. The supernatant was further centrifuged at high speed (140,000 x g) for 30 min. The resulting supernatant and the pellet from low speed were subjected to SDS-PAGE analysis. Protein patterns were obtained by fluorography. Release of a specific glycoprotein of approx. 150 kDa in the medium was observed due to the PIase C treatment. Prolonged incubation of platelets in 0.25 M sucrose and depletion of NaCl concentrations also affected the release of this glycoprotein. BT-PIase C released more approx. 150 kDa protein than SA-PIase C. Western blot experiment with a monoclonal antibody (mAB), epitope SZ2, reactive to human platelet surface glycoprotein Ib (GPIb) complex, confirmed that released 150 kDa glycoprotein reacted with mAB of GPIb. The release of this protein by PIase C was not inhibited by proteinase inhibitors (EDTA, PMSF and leupeptin). Treatment of human platelet membranes with PIase C also caused release of this glycoprotein as evidenced by reactivity to GPIb-mAB. These studies demonstrate that PIase C treatment causes release of 150 kDa glycoprotein from human platelet membrane surface. It is suggested that 150 kDa glycoprotein is anchored to PI in human platelets and that this glycoprotein represents the GPIb complex.
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PMID:Release of a membrane surface glycoprotein from human platelets by phosphatidylinositol specific phospholipase(s) C. 184 99

Thirty and 60-min ischemic insults resulted in an increase in free fatty acid and 1,2- diacylglycerol contents of rat forebrain. No significant changes were detected in phospholipids except phosphatidylinositol 4-monophosphate and phosphatidylinositol 4,5-bisphosphate during ischemic insult. Phosphatidylinositol 4-monohosphate and phosphatidylinositol 4,5-bisphosphate contents decreased during ischemia. Although the increase in free fatty acid contents continued, 1,2-diacylglycerol did not show further increase after 30-min ischemia. These results suggest that there may be another pathway for the accumulation of free fatty acids in addition to phospholipase C coupled to di- and monoacylglycerol lipase. Free fatty acid and 1,2-diacylglycerol contents increased transiently and thereafter decreased to control levels within 90 min after postischemic recirculation. The decrease in arachidonic acid content preceded those of other FFA. Phosphatidylinositol 4-monophosphate and phosphatidylinositol 4,5-bisphosphate contents gradually increased after the initiation of recirculation in ischemic brains. Lysophosphatidylcholine decreased gradually after temporary increase during 15 and 5-min recirculations in 30 and 60-min ischemic groups. Phospholipase A, phospholipase C, and di- and monoacylglycerol lipase activities did not show significant changes during entire course of recirculation. Total activities of lysophospholipase and acylation enzymes of lysophospholipid demonstrated 1.5-and 2.2-fold increase during 30-min recirculation.
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PMID:Changes in lipid metabolites and enzymes in rat brain due to ischemia and recirculation. 191 Mar 56

Carbachol (CCh) a cholinergic agonist which hydrolyses phosphatidyl-inositol bisphosphate (PIP2) to produce the breakdown products inositol trisphosphate (IP3) and diacylglycerol (DAG) was tested for its ability to induce [3H]norepinephrine ([3H]NE) release and to accumulate [3H]inositol phosphate ([3H]IP) under normal and membrane depolarizing conditions. Our results suggest two major points: first, muscarinic acetylcholine receptor (mAChR) agonists and depolarizing agents (of which KCl is the most effective) act in concert to induce potentiation of PI turnover and potentiation of neurotransmitter release. The simultaneous presence of both a depolarizing agent and a receptor agonist is obligatory for eliciting potentiatory effect. Facilitation of release by muscarinic agonist and K+, added together, was 2 to 5-fold above additivity and the levels of [3H]IP accumulated were 3-5-fold above additivity by K+ and CCh. Enhancement of release and of [3H]IP formation is reversed by pirenzepine, a muscarinic (MI) specific antagonist, Kdiss = 0.4 and 0.8 microM, respectively. Second, synergy of IP accumulation in correlation with synergy of neurotransmitter release elicited by mAChR activation and membrane depolarization, suggests a possible role for phospholipase C (PLC) in the bifurcating control of neurotransmitter release and for the involvement of PLC and voltage sensitive channels in mediation of long-term potentiation (LTP).
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PMID:Potentiation of neurotransmitter release coincides with potentiation of phosphatidyl inositol turnover. A possible in vitro model for long term potentiation. 196 29

Madin-Darby canine kidney cells (MDCK) were transfected with a cDNA encoding the glycosyl-phosphatidylinositol (GPI)-anchored protein mouse Thy-1 in order to study the steady-state surface distribution of exogenous and endogenous GPI-linked proteins. Immunofluorescence of transfected cells grown on collagen-coated coverslips showed that expression of Thy-1 was variable throughout the epithelium, with some cells expressing large amounts of Thy-1 adjacent to very faintly staining cells. Selective surface iodination of cells grown on collagen-coated or uncoated transwell filters followed by immunoprecipitation of Thy-1 demonstrated that all the Thy-1 was present exclusively in the apical plasma membrane. Although cells grown on uncoated filters had much smaller amounts of Thy-1, it was consistently localized on the apical surfaces. Immunofluorescent localization of Thy-1 on 1 micron frozen sections of filter-grown cells demonstrated that all the Thy-1 was on the apical surface and there was no detectable intracellular pool. Phosphatidylinositol-specific phospholipase C digestion of intact iodinated monolayers released Thy-1 only into the apical medium, indicating that Thy-1 was processed normally in transfected cells and was anchored by a GPI-tail. In agreement with previous findings, endogenous GPI-linked proteins were found only on the apical plasma membrane. These results suggest that there is a common mechanism for sorting and targeting of GPI-linked proteins in polarized epithelial cells.
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PMID:Polarity of endogenous and exogenous glycosyl-phosphatidylinositol-anchored membrane proteins in Madin-Darby canine kidney cells. 197 90

Two murine monoclonal antibodies have been produced which identify a novel surface antigen expressed on human leucocytes in a non-lineage-restricted distribution. Antibodies WM-63 and WM-68 were derived after immunization of mice with human T-CLL cells and the leukaemic cell line HSB-2. Both antibodies were shown to react with over 90 per cent of normal T and B lymphocytes from peripheral blood and tonsil, and also with monocytes from peripheral blood. A subset of bone marrow leucocytes, including granulocyte-macrophage progenitors, were also reactive. No activity with non-haemopoietic cells or tissues could be identified, however WM-63 and WM-68 showed binding to virtually all cases of chronic B cell malignancy, including chronic lymphatic leukaemia and non-Hodgkin's lymphoma, as well as a proportion of cases of acute leukaemia. Although the antigen recognized by these antibodies could not be immunoprecipitated from membrane extracts, it was removed from the surface of intact cells using the proteolytic enzymes protease and papain. Re-expression on cultured cells was inhibited by incubation with puromycin, cycloheximide, and tunicamycin, indicating that the epitopes detected by WM-63 and WM-68 are likely to be carbohydrate moieties on a protein backbone. Removal of the antigen from the cell surface by treatment with the enzyme phosphatidyl-inositol phospholipase C indicates that it is linked by a phosphatidyl-inositol bond. WM-63 and WM-68 were both recently clustered at the Fourth International Workshop on Human Leucocyte Differentiation Antigens into CD-48, together with four other monoclonal antibodies. Although no biological function has been ascribed to the molecule detected by these antibodies, its restriction to the haemopoietic lineage suggests a role in regulation of leucocyte function.
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PMID:A novel non-lineage antigen on human leucocytes: characterization with two CD-48 monoclonal antibodies. 208 34

Some of the enzyme choline-O-acetyltransferase (ChAT) associated with central cholinergic nerve terminals appears to be non-ionically associated with membranes. In the present study, we tested the possibility that some membrane-bound ChAT might be anchored to membranes by a phosphatidylinositol linkage by incubating rat hippocampal tissue with phospholipase C (PLC) from Bacillus cereus. The PLC selectively augmented the release of ChAT; also, the glycosylphosphatidylinositol-PLC inhibitor, zinc, blocked this increase in release. When control and PLC-treated hippocampal tissues were subjected to Triton X-114 phase separation, a procedure that separates amphiphilic from hydrophilic proteins, the detergent-soluble, membrane-bound fraction of tissue ChAT appeared to be the source of the ChAT released by PLC into the incubation medium. Zinc also blocked the temperature-dependent release of ChAT, but not lactic dehydrogenase, from hippocampal tissue. Extracellular membrane-bound ChAT appeared to be the source of the ChAT released by a low exogenous concentration of PLC, as well as that released by a temperature-dependent process during tissue incubation. Phosphatidylinositol-specific PLC from Bacillus thuringiensis released ChAT, but not lactic dehydrogenase, from a crude synaptosomal fraction prepared from rat hippocampal tissue. These results suggest that some of the membrane-bound ChAT in rat hippocampal tissue may be extracellular and anchored to the membrane by phosphatidylinositol, and also that an endogenous factor in hippocampal tissue may function to remove this extracellular ChAT from the membrane.
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PMID:Effect of phospholipase C from Bacillus cereus on the release of membrane-bound choline-O-acetyltransferase from rat hippocampal tissue. 210 7

Here we present evidence that a fibroblast heparan sulphate proteoglycan of approx. 300 kDa and with a core protein of apparent molecular mass 70 kDa is covalently linked to the plasma membrane via a linkage structure involving phosphatidylinositol. Phosphatidylinositol-specific phospholipase C releases such a heparan sulphate proteoglycan only from cells labelled with [35S]sulphate in the absence of serum. Cell cultures labelled with [3H]myo-inositol in the absence or presence of serum produce a radiolabelled heparan sulphate proteoglycan which was purified by gel-permeation chromatography and ion-exchange chromatography on MonoQ. Digestion with heparan sulphate lyase and analysis by gel-permeation chromatography and sodium dodecylsulphate-polyacrylamide gel-electrophoresis revealed that the 3H-label is associated with a core protein of apparent mass 70 kDa.
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PMID:A fibroblast heparan sulphate proteoglycan with a 70 kDa core protein is linked to membrane phosphatidylinositol. 213 53

Neutrophils express two distinct types of receptor for the Fc region of IgG, FcRII and FcRIII, in amounts of 10,000 to 20,000 FcRII (40 Kd) and 100,000 to 200,000 FcRIII (50 to 80 Kd) per neutrophil. We showed that the FcRIII exhibits genetically determined heterogeneity, detectable by differences in electrophoretic mobility with sodium dodecyl sulfate (SDS) as well as by reaction with antibodies against the biallelic neutrophil-specific antigen system NA. FcRIII was precipitated with an FcRIII-specific monoclonal antibody (MoAb) from the neutrophils of 35 donors. NA1NA1 donors expressed an FcRIII with a molecular weight (mol wt) of 50 to 65 Kd, NA1NA2 donors expressed an FcRIII with a mol wt of 50 to 80 Kd, and NA2NA2 donors expressed an FcRIII with a mol wt of 65 to 80 Kd. Statistical analysis showed that the electrophoretic heterogeneity corresponds with the NA polymorphism (k = 1). Sequential immunoprecipitation with a MoAb against NA1 and a MoAb against anti-FcRIII, followed by SDS-polyacrylamide gel electrophoresis (PAGE), showed that NA1-FcRIII is distinct from NA2-FcRIII. Moreover, immunoprecipitation with a MoAb against NA1 yielded a protein of 50 to 65 Kd, and immunoprecipitation with human anti-NA2 sera or an MoAb against NA2 yielded a protein of 65 to 80 Kd. Preincubation of NA1NA2 neutrophils with F(ab')2 fragments of an MoAb against anti-NA1 reduced binding of IgG dimers to these cells with about 50%, whereas it completely prevented binding of the dimers to NA1NA1 neutrophils. Inhibition experiments with the MoAb against NA2 yielded the same results for NA1NA2 cells, whereas binding of IgG dimers to NA2NA2 cells was completely prevented. Thus, the products of both NA alleles bind IgG. Immunoprecipitation from the medium of neutrophils either stimulated with formyl- methionyl-leucyl-phenylalanine (FMLP) or treated with glycosyl-phosphatidyl-inositol-specific phospholipase C (GPI- PLC) showed that both the NA1-FcRIII and the NA2-FcRIII are released from the cell surface, indicating that both forms of FcRIII have some structural features in common. Deglycosylation of FcRIII from homozygous donors yielded material that showed several bands on SDS-PAGE. GPI-PLC treatment of neutrophils indicated that all of this material is phosphatidyl-inositol linked.
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PMID:Biallelic neutrophil Na-antigen system is associated with a polymorphism on the phospho-inositol-linked Fc gamma receptor III (CD16). 213 3

FcRIII is not present on peripheral blood monocytes, but becomes expressed upon culturing and can be demonstrated on tissue macrophages. We studied the expression of FcRIII of cultured monocytes in detail and compared its structure with FcRIII of neutrophils and NK cells. The cell density of FcRIII reached a plateau within 3 days of culturing. During that time, the expression of FcRI and FcRIIa, also present on monocytes, did not change significantly. FcRIII on cultured monocytes lacked, as did NK cell FcRIII, the NA1-allotypic variant of the NA system present on the neutrophil FcRIII. Studies with glycosyl-phosphatidyl-inositol-specific phospholipase C and analysis of cells of patients with paroxysmal nocturnal hemoglobinuria revealed that FcRIII on cultured monocytes is not anchored by phosphatidyl-inositol-glycan in the cell membrane. Similarly, FcRIII on NK cells was resistant to glycosyl-phosphatidyl-inositol-specific phospholipase C treatment, suggesting that NK cell FcRIII is also not anchored by a phosphatidyl-inositol-glycan moiety, in contrast to neutrophil FcRIII. Analysis by SDS-PAGE showed that the FcRIII of cultured monocytes had a similar mobility as the FcRIII on NK cells, but was clearly distinct from neutrophil FcRIII. Treatment with N-glycanase showed that the protein backbone of deglycosylated FcRIII of cultured monocytes was similar to that of FcRIII of NK cells, but deglycosylated neutrophil FcRIII was different. Specific blocking of FcRIII of cultured monocytes with an anti-FcRIII mAb did not reduced the lytic action of the cultured monocytes towards sensitized erythrocytes. However, FcRIII was modulated from the cell surface by incubation with sensitized E, whereas non-FcR Ag were not. These findings indicate that FcRIII is involved in binding of immune complexes, but does not act as a trigger molecule for extracellular lysis of sensitized E.
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PMID:The Fc-receptor III of cultured human monocytes. Structural similarity with FcRIII of natural killer cells and role in the extracellular lysis of sensitized erythrocytes. 213 96

Phosphatidylinositol-specific phospholipase C isozyme gamma (PLC-gamma, Mr 145,000) is an excellent substrate for the epidermal growth factor (EGF) receptor both in vivo and in vitro. PLC-beta-1, another PLC isozyme, is a poor substrate for the EGF receptor. We examined the relative phosphorylation of PLC-gamma and PLC-beta-1 by the 170-kDa native EGF receptor molecule, the 66-kDa cytoplasmic kinase domain of the EGF receptor (Arg647-Ala1186), the alpha 2 beta 2 native insulin receptor, and the 48-kDa cytoplasmic kinase domain of the insulin receptor beta subunit (Gly947-Ser1343). Similar to the intact EGF receptor, the cytoplasmic kinase domain of the EGF receptor preferentially phosphorylated PLC-gamma. High-performance liquid chromatographic comparison of tryptic phosphopeptides from PLC-gamma phosphorylated by both forms of the EGF receptor kinase indicated similar patterns of multiple tyrosine phosphorylations. These results imply that substrate selectivity, at least in terms of PLC isozymes, is independent of the extracellular ligand-binding and membrane anchor domains of the EGF receptor. In comparison, neither the intact insulin receptor nor the beta-chain kinase domain was able to phosphorylate PLC-gamma to a significant extent. Also, insulin failed to stimulate the phosphorylation of PLC-gamma in NIH 3T3/HIR cells, which overexpress the human insulin receptor. Thus PLC-gamma is not a phosphorylation substrate for the insulin receptor in vitro or in the intact cell.
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PMID:Selectivity of phospholipase C phosphorylation by the epidermal growth factor receptor, the insulin receptor, and their cytoplasmic domains. 215 2


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