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)

Signal transduction pathways may have important regulatory roles in cellular events in the human parasite Schistosoma mansoni. The presence of the phosphoinositide response in S. mansoni was examined by radiolabeling intact worms with 20 muCi of [3H]myoinositol for 24 hr and stimulating parasites with 25 mM NaF and 10 microM AlCl3 in the presence of 10 mM LiCl. Total inositol phosphates were increased within 2 min and maximal accumulation was achieved after 30 min. Similar results were seen with the non-hydrolyzable GTP analogues GTP gamma S and GppNHp while only minimal changes were detected with GMP. Neomycin inhibited NaF-induced inositol phosphate production. NaF stimulated a significant 3.6-fold increase of inositol phosphates in females compared to males. These data suggest that stimulation of guanine nucleotide-binding regulatory proteins activates phospholipase C resulting in production of inositol phosphates in S. mansoni.
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PMID:Schistosoma mansoni: characterization of phosphoinositide response. 130 1

Low density lipoproteins (LDL) bound to the surface of Schistosoma mansoni may protect the parasite from assault by the immune system and provide essential lipids for the parasite in human schistosomiasis. Here we have characterized the LDL binding sites on the surface of schistosomula by comparing the binding of fluorescently labeled LDL to the parasite with LDL binding proteins as seen by ligand blotting before and after enzymatic treatment of viable parasites. Ligand blotting revealed two LDL binding bands, 17.8 +/- 0.8 and 15.7 +/- 0.6 kDa, in intact schistosomula. Trypsinization eliminated all of the specific and approximately two-thirds of the total LDL binding capacity of schistosomula in a time and concentration-dependent manner. LDL did not bind to any bands on blots of trypsinized, viable worms. Specific LDL binding was also eliminated by phosphatidylinositol-specific phospholipase C (PIPLC). PIPLC treatment removed both LDL binding bands from the worms and caused the appearance of an LDL binding band, 16.6 +/- 0.3 kDa, in the culture medium. LDL binding to the parasite recovered within 24 to 48 h after trypsinization but the recovery was inhibited by either monensin or puromycin. Both LDL binding bands reappeared in ligand blots of cultured worms within 24 h; the reappearance was blocked by puromycin but not by monensin. These studies suggest that the specific binding of human LDL to schistosomula is mediated by GPI-linked low molecular weight proteins that are continually synthesized and transported to the parasite surface.
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PMID:Characterization of human low density lipoprotein binding proteins on the surface of schistosomula of Schistosoma mansoni. 132 38

We have recently demonstrated that a 200-kDa antigen that serves as a target of antibodies acting in synergy with praziquantel is linked to the surface membrane of Schistosoma mansoni by a glycosylphosphatidylinositol (GPI) anchor. In the present study we have examined the potential role of this GPI anchor in the therapeutic action of praziquantel by monitoring the release of surface antigens from living adult schistosomes cultured in the presence or absence of praziquantel and exogenous phospholipases. Phosphatidylinositol-specific phospholipase C (PIPLC) selectively released the 200-kDa antigen from the surface of adult schistosomes, as determined by immunoprecipitation experiments; none of the other GPI-anchored proteins, including alkaline phosphatase and a 22-kDa protein, were released by this enzyme. Anti-cross-reacting determinant antiserum (anti-CRD), which recognizes an epitope on GPI-anchored proteins only after the anchor has been removed by PIPLC, specifically precipitated the 200-kDa antigen, confirming the cleavage of its anchor. When the worms were exposed to both praziquantel and PIPLC, the amount of 200-kDa cleaved from the worms was increased five-fold. The selective release of this antigen was also detected by indirect immunofluorescent labeling of praziquantel-exposed adult worms cultured in the presence of phospholipases. Taken together these observations suggest that modulation of the phospholipase-mediated release of GPI-anchored antigens by praziquantel may contribute to the therapeutic action of the drug.
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PMID:Selective release of a glycosylphosphatidylinositol-anchored antigen from the surface of Schistosoma mansoni. 164 1

Schistosoma mansoni parasites recovered from the blood stream were found to be nonactivators of the alternative complement pathway (ACP) when exposed to sera of homologous but not heterologous host species. Schistosomes could be converted into activators of the homologous ACP by treatment with phospholipase C. Antibodies to either human or guinea pig decay accelerating factor (DAF), a 70-kDa glycosylphosphatidylinositol anchored membrane glycoprotein which controls ACP activation on the mammalian cell plasma membrane, bound to the surface of immature schistosomes and immunoprecipitated a molecule of similar molecular mass from detergent extracts of surface iodinated parasites. Phospholipase C treatment drastically reduced the reactivity of the worms with the anti-DAF antibodies. These data suggest that schistosomes evade the ACP by inserting functional host DAF into their surfaces, possibly through adsorption of the molecule's lipophilic diacyglycerol membrane anchor moiety into the outer lipid bilayer of the parasite.
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PMID:Host-specific evasion of the alternative complement pathway by schistosomes correlates with the presence of a phospholipase C-sensitive surface molecule resembling human decay accelerating factor. 169 Jul 76

Sm25, a major antigen in the surface tegument of the parasitic helminth Schistosoma mansoni, is a 25 kDa N-glycosylated glycoprotein which co-purifies with isolated surface membranes and behaves as an integral membrane protein in Triton X-114 (TX-114). The deduced amino acid sequence of Sm25 shows a short C-terminal hydrophobic domain between residues 163 and 180, containing six uncharged polar amino acids and followed by a Lys181-Ser192 dipeptide. We were interested in whether or not this marginal C-terminal amphiphilic domain is responsible for the association of Sm25 with the membrane or whether a post-translational modification such as the addition of glycosyl phosphatidyl inositol (GPI) represents the membrane anchor for this molecule. We find that treatment with phosphatidyl inositol-specific phospholipase C, which cleaves many GPI anchors, does not reveal Cross Reacting Determinant (CRD) on Sm25, nor affect the association of this protein with membranes, providing no support for the addition of GPI. However, Sm25 is palmitoylated via a thioester bond to the single Cys residue, at position 168, which lies within the C-terminal hydrophobic domain. Removal of palmitate by reduction results in a marked decrease in the hydrophobicity of Sm25, as demonstrated by its partitioning into the aqueous rather than detergent phase of TX-114 and its quantitative release from membrane preparations. The hydrophobicity of several membrane proteins in addition to Sm25 is also decreased by reduction, raising the possibility that fatty acylation by thioester linkage is an important mechanism used by schistosomes to stabilize protein-membrane interactions.
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PMID:Sm25, a major schistosome tegumental glycoprotein, is dependent on palmitic acid for membrane attachment. 183 82

We have previously shown that two ectoenzymes, acetylcholinesterase (AChE) and alkaline phosphatase, are released from the surface and from particulate fractions of the parasite Schistosoma mansoni, by a phosphatidylinositol-specific phospholipase C (PtdIns-PLC) of bacterial origin. Exposure to PtdIns-PLC not only removes large amounts of AChE from the surface of intact, viable Schistosoma in culture, but is accompanied by a concomitant increase in overall levels of AChE in the parasite. The same phenomenon is observed with PtdIns-PLC from two different bacterial sources; Staphylococcus aureus and Bacillus thuringiensis. The increase in AChE levels may be ascribed to de novo synthesis since exposure to PtdIns-PLC, in the presence of the protein-synthesis inhibitor cycloheximide, totally blocked the increase in AChE activity. Furthermore, PtdIns-PLC induced an increased incorporation of [35S]methionine into the AChE immunoprecipitated by a specific anti-AChE serum. This increase is selective for AChE, since total protein synthesis remained almost unchanged after PtdIns-PLC addition, and little or no effect was observed on the enzymatic activity of alkaline phosphatase, which is also glycophosphatidylinositol anchored. Since cleavage of the phosphatidylinositol anchor by PtdIns-PLC should liberate diacylglycerol, which may act as second messenger, we investigated the effect of exogenous diacylglycerols on the synthesis of AChE in S. mansoni. Three different diacylglycerols were tested as possible inducers of AChE activity in the parasite. Both 1-oleoyl-2-acetyl-sn-glycerol and 1,2-dimyristoyl-sn-glycerol were able to increase AChE activity by 35-40% at concentrations of 25 micrograms/ml. A higher concentration of 1,2-dioctanoyl-sn-glycerol (70 micrograms/ml) was needed to produce an equivalent effect. Moreover, addition of phorbol-12-myristate-13-acetate, together with the calcium ionophore A23187, produced a similar increase in AChE activity. Finally, polymixin B, a specific inhibitor of protein kinase C, partially blocked the increase in AChE activity induced by PtdIns-PLC. Our results suggest the involvement of glycophosphatidyl membrane-anchor breakdown products as putative second messengers in the parasite S. mansoni.
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PMID:Phosphatidylinositol-specific phospholipase C induces biosynthesis of acetylcholinesterase via diacylglycerol in Schistosoma mansoni. 184 73

Metabolic radiolabeling of adult worms of Schistosoma mansoni with [3H]myristic acid has revealed that the fatty acid is incorporated into more than 15 proteins. We have shown that two of these proteins, a 200-kDa glycoprotein known to be exposed on the surface of the adult worm following praziquantel treatment and a 22-kDa glycoprotein that shows an enhanced immune reactivity with sera of vaccinated mice, are anchored to the adult worm membrane via a glycosylphosphatidylinositol (GPI) linkage. Both antigens partitioned preferentially into the detergent phase of Triton X-114 and were susceptible, following immunoaffinity purification, to hydrolysis by phosphatidylinositol-specific phospholipase C (PIPLC) from Bacillus thuringiensis and phospholipase C from Bacillus cereus. Diacylglycerol (DAG) was released following hydrolysis by bacterial PIPLC; however, Trypanosoma brucei GPIPLC failed to release the diacylglycerol from either protein. Treatment with nitrous acid generated phosphatidylinositol (PI) from both proteins, and phospholipase D from rat serum cleaved phosphatidic acid from the 200-kDa protein. Although the functional significance of these GPI-anchored proteins is unknown, their release from the surface of the schistosome may contribute to immune evasion.
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PMID:Identification and characterization of glycosylphosphatidylinositol-linked Schistosoma mansoni adult worm immunogens. 213 72

Schistosomula of Schistosoma mansoni were examined for the presence of glycosylphosphatidylinositol (GPI) anchored surface membrane Ag. Parasites were surface iodinated and cultured in the presence or absence of a crude phospholipase C (PLC) preparation or phosphatidylinositol-specific PLC (PIPLC). Culture supernatants were then analyzed: 1) by centrifugation to ascertain which molecules released from the surface were soluble or contained in membrane vesicles; 2) by immunoprecipitation with antibodies specific for the "cross-reacting determinant," an epitope revealed on some GPI-anchored proteins only after cleavage of the diacylglycerol from the protein by PIPLC, and 3) by immunoprecipitation with immune mouse sera to establish co-identity with previously described, immunologically relevant surface Ag. By using these techniques, schistosomula were shown to possess three GPI-anchored surface Ag of m.w. 38,000, 32,000 and 18,000 which are spontaneously released from the surface of schistosomula in association with membrane, but remain insoluble until cleaved by PIPLC. All three molecules were recognized by antibodies from mice vaccinated with irradiated cercariae and/or chronically infected mice. Moreover, the m.w. 38,000 component was recognized by a previously described protective mAb (E.1). A major developmental modification appears to occur in the expression of these molecules because, by the same techniques, no GPI-anchored surface Ag were detectable on 7-day-old lung stage parasites. The finding that these important parasite immunogens are GPI-anchored and released from the surface of the parasite in membrane vesicles may, in part, explain why they elicit strong immune responses capable of damaging the schistosomulum tegument.
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PMID:Three major surface antigens of Schistosoma mansoni are linked to the membrane by glycosylphosphatidylinositol. 253 17

Two enzymes, alkaline phosphatase and acetylcholinesterase (AChE), have been shown previously to be components of the surface of the trematode parasite Schistosoma mansoni. In this study we report that both these enzymes and other serine hydrolases are susceptible to release from the S. mansoni tegumental membrane by a phosphatidylinositol-specific phospholipase C (PIPLC) of bacterial origin. These data suggest that AChE and alkaline phosphatase of S. mansoni, as in higher organisms, are anchored to the membrane via covalently attached phosphatidylinositol. The release of AChE from the vesicular fraction of the parasite with PIPLC occurs in a concentration-dependent manner. Sucrose gradient centrifugation of the PIPLC-released AChE showed a single 8.3 S molecular form, similar to that observed for AChE solubilized by Triton X-100. PIPLC removed large amounts of AChE from the surface of intact schistosomula in culture, with no impairment of the viability of the parasite. In this case, an increase in the overall levels of AChE in the intact parasite was observed after addition of PIPLC.
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PMID:Acetylcholinesterase in Schistosoma mansoni is anchored to the membrane via covalently attached phosphatidylinositol. 313 66

Biosynthetic labelling experiments with cercariae and schistosomula of the multicellular parasitic trematode Schistosoma mansoni were performed to determine whether [3H]palmitate or [3H]ethanolamine was incorporated into proteins. Parasites incorporated [3H]palmitate into numerous proteins, as judged by SDS/polyacrylamide-gel electrophoresis and fluorography. The radiolabel was resistant to extraction with chloroform, but sensitive to alkaline hydrolysis, indicating the presence of an ester bond. Further investigation of the major 22 kDa [3H]palmitate-labelled species showed that the label could be recovered in a Pronase fragment which bound detergent and had an apparent molecular mass of 1200 Da as determined by gel filtration on Sephadex LH-20. Schistosomula incubated with [3H]ethanolamine for up to 24 h incorporated this precursor into several proteins; labelled Pronase fragments recovered from the three most intensely labelled proteins were hydrophilic and had a molecular mass of approx. 200 Da. Furthermore, reductive methylation of such fragments showed that the [3H]ethanolamine bears a free amino group, indicating the lack of an amide linkage. We also evaluated the effect of phosphatidylinositol-specific phospholipase C from Staphylococcus aureus: [3H]palmitate-labelled proteins of schistosomula and surface-iodinated proteins were resistant to hydrolysis with this enzyme. In conclusion, [3H]palmitate and [3H]ethanolamine are incorporated into distinct proteins of cercariae and schistosomula which do not bear glycophospholipid anchors. The [3H]ethanolamine-labelled proteins represent a novel variety of protein modification.
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PMID:Characterization of [3H]palmitate- and [3H]ethanolamine-labelled proteins in the multicellular parasitic trematode Schistosoma mansoni. 317 67


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