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)

A rat hepatocyte cell line was cultured in Higuchi's medium with fetal calf serum and insulin and labeled with 35SO2/4-. The cells were treated with a number of ligands to displace the heparan 35SO4 proteoglycan (HSPG) from the pericellular matrix. Maximum release was obtained with D-mannose-6-PO4 (50 mM), D-glucose-6-PO4 (50 mM), myo-inositol-2-PO4 (2-5 mM), myo-inositol hexaphosphate (2-5 mM), and DL-myo-inositol-1-PO4 (1-2 mM). D-myo-Inositol-1,3,4-(PO4)3 (1 mM) and L-myo-inositol-1-PO4 (2 mM) were intermediate in their ability to release the cell surface HSPG, whereas heparin (2 mg/ml), yeast phosphomannan (4 mg/ml), D-xylose-1-PO4 (50 mM), D-glucose-6-SO4 (50 mM), and myo-inositol hexasulfate (5 mM) were ineffective. When 35SO2/4- was added to cell cultures, the total cell surface HSPG increased linearly, but the percentage of the total cell surface [35SO4]HSPG that was released by myo-inositol-PO4 increased with time during the labeling period, reaching a maximum of 65% after 5 h. When cells were labeled for 12 h without insulin in the medium, the maximum amount of cell surface HSPG that was released by myo-inositol-PO4 was reduced to 30%. However, when cells labeled in the absence of insulin were treated with phosphatidylinositol-specific phospholipase C and then myo-inositol-PO4, the release of the cell surface [35SO4]HSPG was increased to 73%. When the [35SO4]HSPG that was released from the cell surface by treatment with myo-inositol-PO4 was added to cultures of unlabeled hepatocytes, it was taken up very rapidly and a portion of the internalized HSPG was converted to free heparan SO4 chains which appeared in the nucleus. Uptake was Ca2+- and Mg2+-independent. The amount of [35SO4]HSPG taken up was markedly reduced when the myo-inositol-PO4-releasable [35SO4]HSPG was pretreated with trypsin, thermolysin, alkaline borohydride, or alkaline phosphatase. When the cells were grown in inositol-deficient medium or in the presence of myo-inositol-PO4, the amount of heparan SO4 found in the nucleus was markedly reduced, and the cells no longer exhibited contact inhibition. These effects of myo-inositol deficiency on the growth and nuclear heparan SO4 were accentuated by addition of LiCl to the cultures to prevent phosphatidylinositol synthesis from the endogenous myo-inositol-PO4.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Involvement of phosphatidylinositol and insulin in the coordinate regulation of proteoheparan sulfate metabolism and hepatocyte growth. 295 71

The ADP-sensitive phosphoenzyme intermediate (E1P) of sarcoplasmic reticulum ATPase was formed using CaATP as a substrate and release of its bound calcium was investigated. Our previous study (Shigekawa, M., Wakabayashi, S., and Nakamura, H. (1983) J. Biol. Chem. 258, 14157-14161) indicated that 1 mol of E1P has 3 mol of high affinity binding sites for Ca2+, of which two are transport sites for Ca2+, whereas the remainder is presumably the nucleotide binding site. The calcium bound to the substrate site was readily replaced by other divalent cations or lanthanide ions. These ions induced a pronounced change in the rate of Ca2+ release from the transport sites on the same phosphoenzyme. In E1P, whose substrate site had been deprived of a metal, Ca2+ release from the transport sites was fast. It was similar to the Ca2+ release from the normal E1P, whose substrate site was occupied by either Ca2+ or Mg2+, in that it was accelerated by increasing pH or high ATP and inhibited markedly by the treatment of the enzyme with phospholipase C. Release of Ca2+ from E1P, whose substrate site had been rendered metal-free, increased the fluorescence intensity of the enzyme-bound 2'(3')-O-(2,4,6-trinitrophenyl)adenosine 5'-triphosphate, which decayed rapidly upon addition of Mg2+ plus K+. This result suggests that the active site of the resultant divalent cation-free phosphoenzyme has a conformation similar to that for the normal ADP-insensitive phosphoenzyme, E2P. These data suggest that the basic mechanism for Ca2+ release from the phosphoenzyme may be similar whether or not the phosphoenzyme has bound metal at its substrate site, although this metal affects the rate of Ca2+ release extensively.
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PMID:Effect of metal bound to the substrate site on calcium release from the phosphoenzyme intermediate of sarcoplasmic reticulum ATPase. 295 67

Various regulators of protein kinase activities were tested for their effects on the in vitro transfer of phosphate from [gamma-32P]ATP to four proteins of rat brain synaptic particulate preparations. One protein, of apparent molecular weight 44,000, accepted 32P in the presence of 8 mM EDTA and no added Mg2+. It was the major phosphoprotein of brain mitochondria. Its phosphorylation was inhibited by pyruvate and stimulated by K+, and it comigrated in electrophoretic gels with authentic alpha-subunit of pyruvate: lipoamide oxidoreductase (decarboxylating) (EC 1.2.4.1) from bovine heart. The major kinase acting on three proteins of apparent molecular weights 24,000, 21,000, and 19,000 was stimulated by Ca2+, by preincubation with phospholipase C, and by 12-tetradecanoyl 4-beta-phorbol 13-acetate. Phosphorylation of these lower-molecular-weight proteins was inhibited by ACTH1-24, by cyclic 3',5'-adenosine monophosphate, and by 50 microM trifluoperazine. The stimulatory effect of Ca2+ was antagonized by calmodulin. The kinase in question appears to be B-50 protein kinase or protein kinase C.
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PMID:Regulation of phosphate incorporation into four brain phosphoproteins that are affected by experience. 298 Dec 89

Enzyme, electrolyte and fluid secretion from exocrine glands is stimulated by neurotransmitters and peptide hormones. Whereas for some of these secretagogues calcium is an important intracellular messenger, for others it is cyclic AMP. Regulation of steady state free Ca2+ concentration at rest and at stimulation have been studied in isolated permeabilized acinar cells from pancreas, parotid and lacrimal glands by measuring the free Ca2+ concentration of the surrounding incubation medium with a Ca2+-specific macroelectrode. Ca2+ transport mechanisms have been further characterized in subcellular membrane fractions by measuring 45Ca2+ uptake into membrane vesicles from rough endoplasmic reticulum (RER) and plasma membranes (PM). The data show that the intracellular messenger for secretagogue-induced Ca2+ release from RER is inositol-1,4,5-trisphosphate (IP3) which is produced during stimulation by phospholipase C mediated hydrolysis of phosphatidylinositol-bisphosphate. At rest both Ca2+ uptake into RER and Ca2+ extrusion from the cell is promoted by (Ca2+ + Mg2+)-ATPases with different characteristics in both types of membranes and by a coupled Na+/Ca2+ countertransport in the PM which keep cytosolic free Ca2+ concentration at a low level of approximately 2 - 4 X 10(-7) mol/l. During stimulation the Ca2+ permeability of endoplasmic reticulum membrane increases via IP3 and that of the PM by a yet unknown "receptor-operated" mechanism. These events lead to increase in cytosolic free Ca2+ concentration that is a trigger for enzyme, electrolyte and fluid secretion.
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PMID:Stimulus-secretion coupling in exocrine glands: the role of inositol-1,4,5-trisphosphate, calcium and cAMP. 299 Aug 22

The activation of platelet V1-receptors by vasopressin (0.01-1 microM) induces the rapid formation of inositol phosphates, 1,2-diacylglycerol and phosphatidic acid, indicating inositol phospholipid hydrolysis by phospholipase C. Vasopressin immediately induces the formation of inositol bisphosphate and inositol trisphosphate. Accumulation of inositol 1-monophosphate and inositol 4-monophosphate occurs later after a time lag of 15 s. Low concentrations (10-100 nM) of vasopressin only activate phospholipase C, whereas high concentrations (1 microM) induce activation of phospholipase C and subsequently the production of arachidonate metabolites. Cyclo-oxygenase metabolites are associated with further activation of phospholipase C, release reaction and irreversible platelet aggregation. Vasopressin requires for its action extracellular Mg2+, but not Ca2+. The described platelet changes are not induced by 1-desamino-[8-D-arginine]vasopressin, a V2-receptor agonist, and are blocked by a specific V1-receptor antagonist. The results indicate that platelets possess a V1-receptor that is coupled to polyphosphoinositide hydrolysis by phospholipase C, leading to the formation of 1,2-diacylglycerol and inositol trisphosphate. Those compounds may act as second messengers for platelet responses induced by vasopressin, whereas endoperoxides and thromboxane A2 stimulated by vasopressin may serve as amplifiers for platelet activation.
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PMID:Activation of V1-receptors by vasopressin stimulates inositol phospholipid hydrolysis and arachidonate metabolism in human platelets. 300 64

From various rat organs, alkaline phosphodiesterase I was liberated by the action of phosphatidylinositol-specific phospholipase C obtained from Bacillus thuringiensis. Especially, a large amount of alkaline phosphodiesterase I was released from slices of small intestine, testis, lung, and kidney, but not from pancreas and liver. The release of the enzyme induced by phospholipase C was dependent on, or proportional to, the reaction time and the concentrations of the phospholipase C and the weight of the slices of small intestine or testis. Furthermore, little enzyme was released from the homogenate of pancreas. These results suggest an important role of phosphatidylinositol in the binding of alkaline phosphodiesterase I to the plasma membranes of rat small intestine and pancreas. The alkaline phosphodiesterase I released from slices of rat small intestine and testis had a molecular weight of about 240,000, and was activated by Mg2+ and Ca2+ but inhibited by EDTA. The enzyme hydrolyzed the phosphodiester linkage of p-nitrophenyl-thymidine 5'-monophosphate at pH 8.9, having the Km values of 0.36 mM (small intestine) and 0.25 mM (testis). The intestinal enzyme differed from the testis enzyme in pI values, thermostability, and Arrhenius plot having a single breakpoint.
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PMID:Alkaline phosphodiesterase I release from eucaryotic plasma membranes by phosphatidylinositol-specific phospholipase C. I. The release from rat organs. 301 61

Phosphatidylinositol-4-phosphate (PtdIns-P) kinase was purified approximately 30-fold from rat brain cytosol. No contaminating activity of PtdIns kinase or of phosphomonoesterase and phospholipase C using PtdIns-P or PtdIns-P2 as substrate could be detected in the enzyme preparation. The PtdIns-P kinase activity was severalfold higher when PtdIns-P/PtdEtn vesicles rather than PtdIns-P alone were used as substrate. This might be due to increased accessibility of the enzyme for the vesicular substrate, further indicated by the lower activity obtained when PtdCho or PtdIns, phospholipids with bulky head groups, was also present in the vesicles. The product PtdIns-P2 was a competitive inhibitor with respect to PtdIns-P and 50% inhibition of enzyme activity was observed at the same product concentration regardless of whether the substrate-product mixture was presented in vesicular or micellar form, or the substrate and product were added in separate vesicles. The polyamines spermine and spermidine enhanced PtdIns-P kinase activity severalfold. Spermine also caused a shift in the MgCl2 saturation curve from sigmoidal to hyperbolic, lowering the Mg2+ concentration required for optimum kinase activity to the physiological range. Myelin basic protein enhanced the enzyme activity when PtdIns-P/PtdEtn vesicles were used as substrate, whereas it was inhibitory when PtdIns-P was added alone. The possible role of polyamines and the product PtdIns-P2 in the regulation of PtdIns-P kinase activity is discussed.
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PMID:Phosphatidylinositol-4-phosphate kinase from rat brain. Activation by polyamines and inhibition by phosphatidylinositol 4,5-bisphosphate. 302 90

Guanine nucleotide-binding regulatory proteins (G proteins) transduce a remarkably diverse group of extracellular signals to a relatively limited number of intracellular target enzymes. In the neutrophil, transduction of the signal following fMet-Leu-Phe receptor-ligand interaction is mediated by a pertussis toxin substrate (Gi) that activates inositol-specific phospholipase C. We have utilized a plasma membrane-containing fraction from unstimulated human neutrophils as the target enzyme to explore the role of G proteins in arachidonate and cytosolic cofactor-dependent activation of the NADPH-dependent O-2-generating oxidase. When certain guanine nucleotides or their nonhydrolyzable analogues were present during arachidonate and cytosolic cofactor-dependent activation, they exerted substantial dose-dependent effects. The GTP analogue, GTP gamma S, caused a 2-fold increase in NADPH oxidase activation (half-maximal stimulation, 1.1 microM). Either GDP or its nonhydrolyzable analogue, GDP beta S, inhibited up to 80% of the basal NADPH oxidase activation (Ki GDP = 0.12 mM, GDP beta S = 0.23 mM). GTP caused only slight and variable stimulation, whereas F-, an agent known to promote the active conformation of G proteins, caused a 1.6-fold stimulation of NADPH oxidase activation. NADPH oxidase activation in the cell-free system was absolutely and specifically dependent on Mg2+. Although O2- production in response to fMet-Leu-Phe was inhibited greater than 90% in neutrophils pretreated with pertussis toxin, cytosolic cofactor and target oxidase membranes from neutrophils treated with pertussis toxin showed no change in basal- or GTP gamma S-stimulated NADPH oxidase activation. Cholera toxin treatment of neutrophils also had no effect on the cell-free activation system. Our results suggest a role for a G protein that is distinct from Gs or Gi in the arachidonate and cytosolic cofactor-dependent NADPH oxidase cell-free activation system.
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PMID:Regulation of neutrophil NADPH oxidase activation in a cell-free system by guanine nucleotides and fluoride. Evidence for participation of a pertussis and cholera toxin-insensitive G protein. 302 97

Ectoenzyme release from porcine intestinal brush border membranes by phosphatidylinositol-specific phospholipase C of Bacillus thuringiensis was studied. Alkaline phosphodiesterase I, alkaline phosphatase and 5'-nucleotidase were released from both slices and brush border membranes. The pattern of alkaline phosphodiesterase I release was the same as that of alkaline phosphatase. The release of alkaline phosphodiesterase I induced by phospholipase C was dependent on, or proportional to, the reaction time and the concentration of phospholipase C. The Arrhenius plot for phosphodiesterase I release showed a single break at 30 degrees C for brush border membranes. Only 40% of alkaline phosphodiesterase I present in the brush border membranes were solubilized by phosphatidylinositol-specific phospholipase C treatment. The data indicate the presence of two forms of phosphodiesterase I, which are different in their sensitivity to phospholipase C. The released alkaline phosphodiesterase I had a molecular weight of 240,000 and was activated by Mg2+ and Ca2+, but strongly inhibited by EDTA.
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PMID:Alkaline phosphodiesterase I release from eucaryotic plasma membranes by phosphatidylinositol-specific phospholipase C. II. The release from brush border membranes of porcine intestine. 302

The leukotriene D4 (LTD4) receptor on rat basophilic leukemia (RBL-1) cell membranes was characterized using a radioligand binding assay. [3H]LTD4 binding to RBL-1 membrane receptors was stereoselective, specific, and saturable. The binding affinity and maximum binding density of [3H]LTD4 to RBL-1 membrane receptors were 0.9 +/- 0.2 nM and 800 +/- 125 fmol/mg protein, respectively. Binding of [3H]LTD4 to the receptors was enhanced by divalent cations (Ca2+, Mg2+, and Mn2+) and inhibited by guanine nucleotides and sodium ions, specifically, indicating that a guanine nucleotide-binding protein may regulate the agonist-receptor interaction. LTD4, LTE4 agonist and antagonist analogs competed with the radioligand in binding to the RBL-1 LTD4 receptors. The binding affinities of these analogs correlated with (a) those determined from the guinea pig lung LTD4 receptors and (b) the pharmacological activities in smooth muscle contraction. LTD4 and related agonists also induced time- and concentration-dependent phosphatidylinositol hydrolysis in RBL-1 cells. The LTD4 induction of inositol 1-phosphate was potent, stereoselective, specific, and was blocked by LTD4 receptor antagonists. The rank order potency of agonist-induced inositol 1-phosphate formation in RBL-1 cells was equivalent to the receptor binding affinity determined using either RBL-1 cell or guinea pig lung membranes. These studies have demonstrated the G protein coupled LTD4 receptors on RBL-1 cell membranes. Binding of agonists to the receptor may activate the G protein-regulated phospholipase C to induce hydrolysis of phosphatidylinositol. The hydrolytic products of phosphatidylinositol, possibly inositol trisphosphate and diacylglycerol, may be the intracellular messengers for LTD4 receptors in RBL-1 cells.
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PMID:Identification and characterization of leukotriene D4 receptors and signal transduction processes in rat basophilic leukemia cells. 303 Oct 59


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