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)

In murine keratinocytes, Ca(++)-induced terminal differentiation is accompanied by a rapid and sustained increase of inositol phosphates and diacylglycerol. Based on Western blotting analysis, basal keratinocytes cultured in 0.05 mM Ca++ medium express phospholipase C (PLC)-gamma 1 predominantly and no detectable PLC-beta 1. Differentiating keratinocytes cultured in 1.4 mM Ca++ express two- to threefold more PLC-gamma 1 protein and PLC-delta 1, but no detectable PLC-beta 1. Although the amount of PLC-gamma 1 and -delta 1 protein increased, PLC-gamma 1 and -delta 1 mRNA decreased in differentiating cells. Thus the sustained rise of PLC activity induced by Ca++ in differentiating keratinocytes may be associated with higher amounts of both PLC-gamma 1 and -delta 1 in maturing cells, determined by a posttranscriptional mechanism. Tyrosine phosphate content in PLC-gamma 1 was low in basal cells and did not change in cells exposed to 1.4 mM Ca++. However, genistein inhibited the increase in PLC activity induced by 1.4 mM Ca++. In contrast, transforming growth factor (TGF)alpha, which stimulates both PLC activity and growth in basal keratinocytes, increased tyrosine phosphorylation of PLC-gamma 1. These results suggest that tyrosine phosphorylation of PLC-gamma 1 by the epidermal growth factor (EGF) receptor is linked to stimulated proliferation, whereas stimulation of PLC activity by Ca++ is linked to keratinocyte differentiation and involves the action of a tyrosine kinase but not tyrosine phosphorylation of PLC-gamma 1. Based on studies using the intracellular free Ca++ chelator BAPTA, a rise in intracellular free Ca++ was not required for stimulation of PLC activity by raising extracellular Ca++. Phorbol esters inhibited PLC stimulation by 1.4 mM Ca++ medium and increased serine phosphorylation of PLC-gamma 1. Exogenous phosphatidylinositol-specific and phosphatidylcholine-specific bacterial PLC also inhibited endogenous inositol phosphate formation and increased endogenous diacylglycerol (DAG). Thus, direct serine phosphorylation of PLC-gamma 1 by protein kinase C is associated with the inhibition of Ca(++)-mediated PLC stimulation. These results show that keratinocytes have multiple mechanisms to regulate PLC activity in response to a specific signal.
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PMID:Keratinocyte differentiation is associated with changes in the expression and regulation of phospholipase C isoenzymes. 822 34

The major macromolecules on the surface of the parasitic protozoan Leishmania major appear to be down-regulated during transformation of the parasite from an insect-dwelling promastigote stage to an intracellular amastigote stage that invades mammalian macrophages. In contrast, the major parasite glycolipids, the glycoinositol phospholipids (GIPLs), are shown here to be expressed at near-constant levels in both developmental stages. The structures of the GIPLs from tissue-derived amastigotes have been determined by h.p.l.c. analysis of the deaminated and reduced glycan head groups, and by chemical and enzymic sequencing. The deduced structures appear to form a complete biosynthetic series, ranging from Man alpha 1-4GlcN-phosphatidylinositol (PI) to Gal alpha 1-3Galf beta 1-3Man alpha 1-3Man alpha 1-4GlcN-PI (GIPL-2). A small proportion of GIPL-2 was further extended by addition of a Gal residue in either alpha 1-6 or beta 1-3 linkage. From g.c.-m.s. analysis and mild base treatment, all the GIPLs were shown to contain either alkylacylglycerol or lyso-alkylglycerol lipid moieties, where the alkyl chains were predominantly C18:0, with lower levels of C20:0, C22:0 and C24:0. L. major amastigotes also contained at least two PI-specific phospholipase C-resistant glycolipids which are absent from promastigotes. These neutral glycolipids were resistant to both mild acid and mild base hydrolysis, contained terminal beta-Gal residues and were not lost during extensive purification of amastigotes from host cell membranes. It is likely that these glycolipids are glycosphingolipids acquired from the mammalian host. The GIPL profile of L. major amastigotes is compared with the profiles found in L. major promastigotes and L. donovani amastigotes.
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PMID:Characterization of glycoinositol phospholipids in the amastigote stage of the protozoan parasite Leishmania major. 824 Feb 57

Xenopus laevis oocytes were injected with mRNA extracted from growth factor-responsive CCL39, Chinese hamster lung fibroblasts. The expression of functional growth factor receptors on the oocytes was demonstrated by growth factor-induced 45Ca2+ efflux. To determine the isozyme(s) of phospholipase C (PLC) coupled to growth factor receptors, growth factor-induced 45Ca2+ efflux were measured following coinjection of mRNA from CCL39 cells with PLC antibodies. PLC-gamma 1 antibody did not lead to loss of 45Ca2+ efflux induced by thrombin but resulted in loss of that induced by platelet-derived growth factor (PDGF). In contrast, PLC-delta 1 antibody did not block PDGF-induced 45Ca2+ efflux but led to inhibition of thrombin-induced 45Ca2+ efflux. PLC-beta 1 antibody did not affect Ca2+ efflux by the treatment of either thrombin or PDGF. These results suggest that these growth factor receptors are coupled to specific effectors, i.e. thrombin receptor to PLC-delta 1 and PDGF receptor to PLC-gamma 1.
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PMID:Selectivity of phospholipase C isozymes in growth factor signaling. 824 27

The hydrolysis of phosphatidylinositol 4,5-bisphosphate by specific phospholipase C (PLC) enzymes produces two second messengers, inositol 1,4,5-trisphosphate and diacylglycerol. Heterotrimeric guanine nucleotide-binding regulatory proteins (G proteins) of the Gq subfamily activate the PLC beta 1 isoform of PLC. We have purified three isozymes of PLC beta: PLC beta 1 and PLC beta 3 from rat brain and PLC beta 2 from HL-60 cells. Whereas the beta 1 and beta 2 isozymes appear restricted to a few cell types, beta 3 is broadly distributed. Gq alpha (the alpha subunit of the Gq subfamily) can activate all three isoforms but PLC beta 2 is much less sensitive. Thus all three enzymes are potential effectors for pertussis toxin-insensitive regulation by hormones. The three beta isozymes can also be activated by purified beta gamma subunits. The PLC beta 3 isoform gives the greatest activation with beta gamma; PLC beta 1 is least responsive. The results indicate that all the known isoforms of mammalian PLC beta can be regulated at unique sites by both Gq alpha and beta gamma subunits. The effect of beta gamma subunits may provide a pathway for the regulation of PLC beta isozymes by pertussis toxin-sensitive G proteins or may indicate that the alpha subunit of Gq and its associated beta gamma both participate in regulation of the same phospholipase molecule.
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PMID:G proteins in signal transduction: the regulation of phospholipase C. 829 29

In the present study an attempt was made to further elucidate the molecular mechanisms whereby protein kinase C (PKC) modulates the beta-cell stimulus-secretion coupling. Regulation of Ca2+ channel activity, [Ca2+]i, and insulin release were investigated in both normal pancreatic mouse beta-cells and in similar beta-cells deprived of PKC activity. [Ca2+]i was measured with the intracellular fluorescent Ca2+ indicator fura-2 and the Ca2+ channel activity was estimated by the whole cell configuration of the patch-clamp technique. To reveal the various isoenzymes of PKC present in the mouse beta-cell, proteins were separated by one-dimensional gel electrophoresis and Western blotting was performed. The production of inositol phosphates was measured by ion-exchange chromatography and insulin release was measured radioimmunologically. Acute stimulation with the phorbol ester 12-O-tetradecanoylphorbol-13-acetate resulted in suppression of both the carbamylcholine-induced increase in [Ca2+]i and production of inositol 1,4,5-trisphosphate. Under these conditions the increase in [Ca2+]i in response to glucose was similar to that found in control cells. When beta-cells were deprived of PKC, by exposure to 200 nM 12-O-tetradecanoylphorbol-13-acetate for 24-48 h, there was an enhanced response to carbamylcholine. This response constituted increases in both the [Ca2+]i signal and production of inositol 1,4,5-trisphosphate. Interestingly, cells with down-regulated PKC activity responded more slowly to glucose stimulation, when comparing the initial increase in [Ca2+]i, than control cells. On the other hand, the maximal increase in [Ca2+]i was similar whether or not PKC was present. Moreover, PKC down-regulated cells exhibited a significant reduction of maximal whole cell Ca2+ currents, a finding that may explain the altered kinetics with regard to the [Ca2+]i increase in response to the sugar. Both the alpha and beta 1 forms of the PKC isoenzymes were present in the mouse beta-cell and were also subjected to PKC down-regulation. Hence, either of these isoenzymes or both may be involved in the modulation of phospholipase C and Ca2+ channel activity. Since insulin release under physiological conditions is critically dependent on Ca(2+)-influx through the voltage-gated L-type Ca2+ channels, the kinetics of hormone release was expected to demonstrate a similar delay as that of the [Ca2+]i increase. Although not as pronounced, such a delay was indeed also observed in the onset of insulin release. There was, however, no effect on the total amounts of hormone released.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Protein kinase C modulates the insulin secretory process by maintaining a proper function of the beta-cell voltage-activated Ca2+ channels. 830 Jun 6

Receptor activation of phospholipase C (PLC) via G-proteins occurs by pertussis toxin-sensitive and toxin-insensitive signaling pathways. The alpha-subunits of the Gq family are presumed to mediate the toxin-insensitive pathway, but the nature of the G-proteins mediating the toxin-sensitive pathway is not established. Recently, PLC-beta has been shown to be activated by G-protein beta gamma-subunits of mixed or undefined composition. The relative activities of G-protein subunits that might activate PLC-beta were examined using defined recombinant alpha- and beta gamma-subunits obtained from the baculovirus expression system by reconstituting the purified subunits with purified bovine brain PLC-beta 1 or turkey erythrocyte PLC-beta in unilamellar phospholipid vesicles. Turkey erythrocyte G alpha 11 and recombinant G alpha 11 and G alpha q obtained after expression in Sf9 cells activated both bovine brain PLC-beta 1 and turkey erythrocyte PLC-beta. In contrast, under the same assay conditions, recombinant G alpha i1, G alpha i2, G alpha i3, and G alpha o were without effect on either type of PLC. All types of beta gamma-subunits tested (r beta 1 gamma 2, r beta 1 gamma 3, r beta 2 gamma 2, r beta 2 gamma 3, bovine brain beta gamma or turkey erythrocyte beta gamma) inhibited G alpha 11-mediated activation of PLC, presumably by promotion of formation of inactive heterotrimeric G-protein. All types of beta gamma-subunits also markedly stimulated the activity of turkey erythrocyte PLC-beta but did not activate bovine brain PLC-beta 1. Of the four different beta gamma complexes of defined composition, three stimulated PLC with similar activities whereas beta 2 gamma 3 was less effective. The data suggest that pertussis toxin-sensitive activation of PLC is mediated by the beta gamma-subunits of G-proteins acting on specific phospholipase C isoenzymes.
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PMID:Selective activation of phospholipase C by recombinant G-protein alpha- and beta gamma-subunits. 830 Jun 14

Recombinant wild-type beta 1 gamma 1 dimers of signal-transducing guanine nucleotide-binding proteins (G proteins) and beta 1 gamma 1 dimers carrying a mutation known to block gamma-subunit isoprenylation (beta 1 gamma 1 C71S) were expressed in baculovirus-infected insect cells. Both wild-type and mutant beta 1 gamma 1 dimers were found in soluble fractions of infected cells upon subcellular fractionation. Anion exchange chromatographic and metabolic-radiolabeling studies revealed that the soluble beta 1 gamma 1 preparation contained approximately equal amounts of non-isoprenylated and isoprenylated beta 1 gamma 1 dimers. Soluble wild-type and mutant beta 1 gamma 1 dimers and native beta 1 gamma 1 dimers purified from bovine retina were reconstituted with recombinant phospholipase C-beta 2. Only isoprenylated beta 1 gamma 1 dimers were capable of stimulating phospholipase C-beta 2. The results show that gamma-subunit isoprenylation and/or additional post-translational processing of the protein are required for beta gamma subunit stimulation of phospholipase C.
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PMID:Stimulation of phospholipase C-beta 2 by recombinant guanine-nucleotide-binding protein beta gamma dimers produced in a baculovirus/insect cell expression system. Requirement of gamma-subunit isoprenylation for stimulation of phospholipase C. 830 83

The beta and gamma subunits of heterotrimeric guanine nucleotide-binding regulatory proteins (G proteins) form tightly associated complexes. To examine functional differences among the large number of possible combinations of unique beta and gamma subunits, we have synthesized and characterized beta gamma complexes containing gamma 5 and gamma 7, two widely distributed gamma subunits. When either gamma 5 or gamma 7 is expressed concurrently with beta 1 or beta 2 subunits in a baculovirus/Sf9 cell system, all four subunit complexes support pertussis toxin-catalyzed ADP-ribosylation of rGi alpha 1 (where "r" indicates recombinant), indicating formation of functional complexes. Each of the complexes was purified by subunit exchange chromatography, using the G203A mutant of rGi alpha 1 as the immobilized ligand. The purified preparations were compared with other recombinant beta gamma subunits, including beta 1 gamma 1 and beta 1 gamma 2, for their ability to modulate type I and II adenylyl cyclase activities; stimulate phosphoinositide-specific phospholipase C beta; support pertussis toxin-catalyzed ADP-ribosylation of rGi alpha 1 and Go alpha; and inhibit steady-state GTP hydrolysis catalyzed by Gs alpha, Go alpha, and myristoylated rGi alpha 2. The results emphasize the unique properties of beta 1 gamma 1. The properties of the complexes containing gamma 5 or gamma 7 were similar to each other and to those of beta 1 gamma 2.
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PMID:G protein beta gamma subunits. Simplified purification and properties of novel isoforms. 830 9

Members of the Gq alpha subfamily of heterotrimeric guanine nucleotide-binding proteins (G proteins) activate phospholipase C (PLC). The complementary DNAs (cDNAs) for the G protein alpha subunits Gq alpha and G11 alpha were expressed in insect (Sf9) cells using recombinant baculovirus. Active, nonaggregated, and membrane-associated protein was generated only when the alpha subunit cDNA was expressed together with cDNAs encoding G protein beta and gamma subunits. Recombinant alpha subunits (rGq alpha and rG11 alpha) were purified by three-step procedures, as was a PLC-activating alpha subunit(s) endogenous to Sf9 cells. Guanosine 5'-3-(thio)triphosphate (GTP gamma S) activated rGq alpha and rG11 alpha with an apparent K0.5 of 30 microM; similarly high concentrations of the nucleotide were required to observe [35S]GTP gamma S binding to rGq alpha. Activated rGq alpha and rG11 alpha each stimulated all three isoforms of purified PLC-beta with the rank order of potency PLC-beta 1 = PLC-beta 3 > or = PLC-beta 2; both alpha subunits also stimulated PLC-beta 1 and PLC-beta 3 to a much greater extent (10-fold) than they did PLC-beta 2. In contrast, activated rGq alpha and rG11 alpha failed to stimulate either PLC-delta 1 or PLC-gamma 1. Recombinant Gi alpha 1, Gi alpha 2, Gi alpha 3, Go alpha (A), Gs alpha, and Gz alpha all failed to stimulate any of the isoforms of PLC. The apparent affinities of rGq alpha and rG11 alpha for PLC-beta 1 and their capacities to activate the enzyme were similar to values observed for purified brain Gq alpha/11 alpha. Purified brain beta gamma subunits also stimulated the three isoforms of PLC-beta. The capacities of rGq alpha and rG11 alpha to activate PLC-beta 1 and PLC-beta 3 greatly exceeded those of beta gamma, whereas Gq alpha, G11 alpha and beta gamma were roughly equiefficacious with PLC-beta 2; the alpha subunits were more potent than beta gamma in all cases. The effects of alpha and beta gamma together were nonadditive for both PLC-beta 1 and PLC-beta 2. These results demonstrate that Gq alpha and G11 alpha specifically and selectively stimulate beta isoforms of PLC and confirm the idea that these members of the Gq alpha subfamily of G proteins are physiological regulators of this signaling pathway.
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PMID:Purification from Sf9 cells and characterization of recombinant Gq alpha and G11 alpha. Activation of purified phospholipase C isozymes by G alpha subunits. 831 96

The heat shock response in mammals consists of a complex array of intracellular reactions initiated by stress, although its regulation is poorly understood. We have investigated the role of transmembrane signal transduction in the response, examining mechanisms involved in the activation of phospholipase C (PLC) by heat shock. In rodent fibroblasts permeabilized with digitonin, heat shock and receptor-mediated PLC activity exhibited a strict GTP analog dependency. This indicates that heat shock-mediated phospholipase activation, in common with receptor mediated stimulation, does not involve direct effects on the phospholipases and suggests the participation of GTP binding (G) proteins in the activation process. When cells were treated with the inhibitor pertussis toxin (PTX), the phospholipases retained their inducibility by heat shock, but became refractory to thrombin treatment, indicating that heat shock may influence PLC activity through a distinct population of G proteins compared to thrombin. The data seem to exclude a role for PTX sensitive G proteins in the production of IP3 after heating and suggest a pathway involving the direct thermal activation of the Gq class of G proteins, which are coupled to the PLC beta 1 isoform.
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PMID:Activation of phospholipase C by heat shock requires GTP analogs and is resistant to pertussis toxin. 831 54


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