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)

The hydrolytic activity of phosphatidylinositol (PI)-specific phospholipase C (PI-PLC) from Bacillus thuringiensis was studied in detail toward mixed liposomes consisting of PI and one of other phospholipids and cholesterol. Among PI-liposomes, small unilamellar vesicles (SUV) were the most sensitive to PI-PLC; the enzymatic hydrolysis of PI in SUV was not less than 10-fold that in large unilamellar vesicles (LUV) or in multilamellar vesicles (MLV). Thus, in a survey of the effects of coexisting lipids on PI-PLC activity, PI-SUV was used. Phosphatidylcholine (PC) was stimulative for the enzyme activity toward PI-SUV at any molar ratio of PC to PI. Also, the effects of the addition of sphingomyelin (SM), phosphatidylethanolamine (PE) and cholesterol on the enzymatic hydrolysis of PI were studied in detail on the basis of concentration of total lipids or PI.
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PMID:The effects of coexisting lipids on the action of Bacillus thuringiensis phosphatidylinositol-specific phospholipase C toward liposomal substrate. 179 44

Aggregation of the high affinity receptor for IgE (Fc epsilon RI) on mast cells by a polyvalent Ag leads to hydrolysis of phosphoinositides (PI) catalyzed by phospholipase C (PI-PLC). To understand this phenomenon in molecular terms, it is important to obtain active, cell-free preparations. In extensive preliminary studies, we could not demonstrate Fc epsilon RI-mediated activation of PI-PLC in plasma membranes prepared by conventional methods from rat basophilic leukemia cells. We now report a stepwise approach involving preparation of cytoplasts from such cells and then hypotonic lysis of the cytoplasts to obtain active membrane vesicles. These membranes, best described as "ghosts," appear to reseal after losing greater than 90% of their soluble, cytoplasmic components and contain receptors that when aggregated, activate PI-PLC to hydrolyze endogenous phospholipids. Per unit of plasma membrane, the ghosts retain approximately 25% of Fc epsilon RI-mediated stimulation of PI-PLC relative to the cells. This activity requires ATP, magnesium, phosphoenolpyruvate, and, to a limited degree, calcium. Although an adequate amount of phosphatidylinositol biphosphate is present, the predicted spike of (1,4,5)-inositol trisphosphate is not seen, and the predominant inositol phosphate isomer is (1,4)-inositol bisphosphate. This is the first report of Fc epsilon RI-mediated activation of PI-PLC in a cytoplasm-depleted system that demonstrates activation of endogenous enzyme acting on endogenous substrate. In addition, it is the first such report for any receptor of the Ig superfamily.
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PMID:Fc epsilon RI-mediated hydrolysis of phosphoinositides in ghosts derived from rat basophilic leukemia cells. 184 42

The molecular nature and cellular localization of Thy-1 antigen in human renal tissue were studied. Strong immunohistochemical staining was observed in frozen sections of human kidney using monoclonal anti-human Thy-1 antibody; this reaction was almost completely abolished by pretreating the kidney section with phosphatidyl inositol (PI)-specific phospholipase C (PI-PLC). Immunohistochemical analysis revealed that the Thy-1 antigen is localized on the proximal tubular epithelial cells and the Bowman's capsule of the glomerulus. Northern blot analysis of renal mRNA using a cloned human Thy-1 gene revealed the presence of human Thy-1 mRNA of a similar size to the one in human brain. When a human kidney cDNA library was screened with the same probe, a cDNA of human Thy-1 was isolated. Moreover, human Thy-1 protein with a molecular weight (MW) of 21,000 was detected in renal tissue by gel electrophoresis and Western blot analysis using monoclonal anti-human Thy-1 antibody. These data demonstrate for the first time the production of human Thy-1 as a PI-anchored protein with a unique cellular location in human renal tissue.
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PMID:Determination of the molecular nature and cellular localization of Thy-1 in human renal tissue. 196 87

The deacylated forms of the phosphoinositides were used to determine whether the guinea pig uterus phosphoinositide-specific phospholipase C (PI-PLC I, Mr 60,000) required fatty acids at the sn-1 and sn-2 positions for the hydrolysis of the sn-3 phosphodiester bond. L-alpha-Glycerophospho-D-myo-inositol 4-phosphate (Gro-PIP), but not glycerol 3-phosphate (Gro-3-P), L-alpha-glycerophospho-D-myo-inositol (Gro-PI), or L-alpha-glycerophospho-D-myo-inositol 4,5-bisphosphate (Gro-PIP2), inhibited PI-PLC I in a concentration-dependent manner. Assays performed with 10 microM [3H]phosphatidylinositol ([3H]PI), 10 microM [3H]phosphatidylinositol 4-phosphate ([3H]PIP) or 10 microM [3H]phosphatidylinositol 4,5-bisphosphate ([3H]PIP2) as substrates, with increasing [Gro-PIP] revealed an IC50 = 380 microM. Kinetic studies with increasing [3H]PI substrate concentrations in the presence of 100 microM and 300 microM Gro-PIP demonstrated that Gro-PIP exhibited competitive inhibition; Kis = 40 microM. Ca2+ concentrations over the range 1.1 microM to 1 mM did not effect inhibition, suggesting that Gro-PIP inhibition of [3H]PI hydrolysis was calcium-independent. To determine whether Gro-PIP was a substrate, 20 microM and 500 microM [3H]Gro-PIP were incubated with PI-PLC I. Anion-exchange HPLC analysis revealed no [3H]IP2 product formation, indicating that [3H]Gro-PIP was not hydrolyzed. Assays performed with [3H]PI and [3H]PIP substrates in the presence of 500 microM [3H]Gro-PIP revealed approx. 75% less [3H]inositol 1-phosphate ([3H]IP1) and [3H]inositol 1,4-bisphosphate ([3H]IP2) product formation, respectively, indicating that [3H]Gro-PIP inhibited the hydrolysis of the substrates by PI-PLC I. These data suggest that Gro-PIP does not serve as a substrate, and that it inhibits PI-PLC I by competitive inhibition in a Ca2(+)-independent fashion.
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PMID:Glycerol-3-phospho-D-myo-inositol 4-phosphate (Gro-PIP) is an inhibitor of phosphoinositide-specific phospholipase C. 215 9

The effect of electroconvulsive treatment (ECT) on activities of phospholipase C hydrolyzing phosphatidylinositol (PI-PLC) and phosphatidylinositol 4,5-bisphosphate (PIP2-PLC) and guanosine-5'-(3-O-thio)triphosphate (GTP gamma S) binding activity were examined in membrane and cytosol fractions from four discrete areas (prefrontal cortex, hippocampus, striatum, and amygdala) of the rat brain. A single ECT resulted in an increase in cytosolic activities of PI-PLC in the prefrontal cortex and of PIP2PLC in all 4 brain regions examined. There were no significant changes in either PI-PLC or PIP2-PLC activity in membrane fractions after a single ECT. Repeated ECT caused regionally specific changes in PLC activities as follows: in the prefrontal cortex, both cytosolic PI-PLC and PIP2-PLC and membranous PI-PLC activities were decreased; in the hippocampus, no changes in any PLC activities were seen; in the striatum, only membranous PI-PLC activities were increased; and, in the amygdala, cytosolic and membranous PI-PLC and cytosolic PIP2-PLC activities were increased. The pattern of changes in GTP gamma S binding activity following repeated ECT resembled those found in PLC activity as follows: in the prefrontal cortex, GTP gamma S binding activities were significantly reduced in both membrane and cytosol; in the hippocampus, the activity was decreased in membrane; in the striatum, no changes in GTP gamma S binding activity were seen in any fraction; and, in the amygdala, the activity was increased in cytosol. These findings suggest that ECT has complex effects on the G protein-phospholipase C system, possibly affecting neuronal signal transduction.
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PMID:Electroconvulsive treatment: effects on phospholipase C activity and GTP binding activity in rat brain. 216 50

About 50-70 mg in total of carcinoembryonic antigen (CEA) (or CEA-related antigens) was detected in normal adult feces evacuated during one day (200-250 g). Ten percent or less of the antigen was found to be in soluble form in fresh feces (naturally solubilized antigen), while 90% or more was still in membrane-bound form which was releasable with phosphatidylinositol-specific phospholipase C (PI-PLC-solubilized antigen). The naturally solubilized and PI-PLC-solubilized antigens are antigenically different from each other and similar to normal fecal antigen (NFA)-2 and CEA, respectively, suggesting that "CEA-distinctive" antigenicity detected so far in CEA from cancerous tissues is not due to the difference between antigens in normal and malignant tissues but is probably due to the presence of the glycosylinositolphosphate moiety at the carboxyl-terminus of the antigen molecule. Thus, "CEA-distinctive" antigenicity is by no means cancer-specific, but this antigenicity seems to be critical for the clinical significance of CEA as a tumor marker, because an assay system (Kit II) which is able to distinguish CEA from NFA-2 revealed much improved features in cancer diagnosis as reported recently.
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PMID:Characterization of carcinoembryonic antigen-related antigens in normal adult feces. 216 22

The cellular mechanism(s) by which GH produces insulin resistance in peripheral tissues is poorly understood. Recent evidence suggests that insulin exerts certain of its intracellular actions by rapidly activating phosphatidylinositol-specific phospholipase C(s) (PI-PLC) in the plasma membranes of target cells. Therefore, the present study was conducted to determine whether insulin can activate PI-PLC in adipose tissue of the genetically obese (ob/ob) mouse, an animal that responds markedly to GH with enhanced peripheral insulin resistance. Also, experiments were performed to determine whether the activation of PI-PLC by insulin could be blocked by S-carboxymethylated human GH (RCM-hGH), a GH derivative possessing mainly diabetogenic activity. Isolated adipose segments were incubated for various periods with insulin (10 mU/ml), homogenized and centrifuged to obtain a 150,000 x g pellet, and the latter was assayed for the ability to produce [3H]inositol phosphate from phosphatidyl[3H]inositol. PI-PLC activity was significantly stimulated 5 min after exposure of the segments to insulin. By 10 min, the insulin effect was no longer apparent, and after 30 min, insulin reduced the activity of the enzyme. One hour after exposure to insulin, PI-PLC activity returned to the control level. When adipose segments of RCM-hGH-treated mice (200 micrograms/day for 3 days sc) were incubated for 5 min with insulin, the ability of insulin to activate PI-PLC was abolished. However, RCM-hGH did not alter basal PI-PLC, indicating that its action involves the mechanism by which the enzyme is activated by insulin. Also, studies utilizing acute RCM-hGH treatment showed that its inhibitory effect on insulin activation of PI-PLC occurs within the same time frame as the onset of enhanced insulin resistance in the adipose tissue. Thus, the ability of GH to inhibit the activation of PI-PLC by insulin in adipocytes may account, at least in part, for its ability to induce insulin resistance in these cells.
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PMID:Growth hormone inhibits activation of phosphatidylinositol phospholipase C by insulin in ob/ob mouse adipose tissue. 240 23

Protein sequence analysis of a bovine brain phosphoinositide-specific phospholipase C (PI-PLC; PLC-154) has permitted the isolation of a cDNA that appears to code for this protein. Transient expression of this cDNA in COS-1 cells demonstrates that the cDNA encodes a functional phospholipase C that migrates at approximately 150,000 daltons. A transcript of approximately 7 kb is observed in RNA derived from bovine brain and a related transcript of the same size is present in certain human cell lines. Southern blot analysis indicates that one or possibly two genes hybridize with a PLC-154 probe. Regions of homology between PLC-154 and the previously described PLC-148 allow the assignment of a putative catalytic domain to the central region of PLC-154.
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PMID:Determination of the primary structure of PLC-154 demonstrates diversity of phosphoinositide-specific phospholipase C activities. 245 1

The phosphatidylinositol (PI)-specific phospholipase C (PLC) of Bacillus cereus was cloned into Escherichia coli by using monoclonal antibody probes raised against the purified protein. The enzyme is specific for hydrolysis of the membrane lipid PI and PI-glycan-containing membrane anchors, which are important structural components of one class of membrane proteins. The protein expressed in E. coli comigrated with B. cereus PI-PLC in sodium dodecyl sulfate-polyacrylamide gel electrophoresis, as detected by immunoblotting, and conferred PI-PLC activity on the host. This enzyme activity was inhibited by PI-PLC-specific monoclonal antibodies. The nucleotide sequence of the PI-PLC gene suggests that this secreted bacterial protein is synthesized as a larger precursor with a 31-amino-acid N-terminal extension to the mature enzyme of 298 amino acids. From analysis of coding and flanking sequences of the gene, we conclude that the PI-PLC gene does not reside next to the gene cluster of the other two secreted phospholipases C on the bacterial chromosome. The deduced amino acid sequence of the B. cereus PI-PLC contains a stretch of significant similarity to the glycosylphosphatidylinositol-specific PLC of Trypanosoma brucei. The conserved peptide is proposed to play a role in the function of these enzymes.
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PMID:Phosphatidylinositol-specific phospholipase C of Bacillus cereus: cloning, sequencing, and relationship to other phospholipases. 250 27

Phospholipase C (PLC)-mediated degradation of polyphosphoinositides (phosphatidylinositol 4,5-bisphosphate (PIP2) and phosphatidylinositol 4-phosphate (PIP] was found to be present in rat heart ventricular soluble and total membrane fractions (100,000g supernatant and pellet). Distribution of polyphosphoinositide-specific phospholipase C activity between the membrane and soluble fraction was approximately 63 and 33% of total activity, respectively, whereas, phosphatidylinositol (PI) degradation could be detected only in the soluble fraction. Optimal PIP2-PLC activity occurred at a pCa2+ of 4.5. A similar peak in PIP-PLC activity could be demonstrated in soluble and membrane preparations; however, the rate of PIP degradation in the soluble fraction continued to increase at the highest calcium level tested (pCa2+ 3). With the exception of Sr2+, other noncalcium polycations did not support homogenate PIP2-PLC activity. In the presence of Ca2+, addition of Mg2+, La3+, or Sr2+ (10(-3) M) inhibited PIP2-PLC while Mn2+ and Gd3+ stimulated activity. In both the total membrane and soluble fractions, maximal polyphosphoinositide degradation occurs at pH 5.5 and 6.8. The detergents deoxycholate, cholate, and saponin exert a biphasic effect on PIP2-PLC activity (stimulating at lower concentrations and inhibiting at higher concentrations). The deoxycholate effect is observed in both the cytosolic and membrane fractions. Neutral and cationic detergents inhibit PIP2-PLC activity in a concentration-dependent manner. Similar to cytosolic PI-PLC activity, PIP2-PLC appears to depend on intact sulfhydryl groups. In the presence of a mixture of all three inositol phospholipids or the three phosphoinositides plus noninositol phospholipids, polyphosphoinositides are preferentially degraded.
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PMID:Characterization of phospholipase C-mediated polyphosphoinositide hydrolysis in rat heart ventricles. 253 55


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