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)

The 145-kDa phospholipase C isozyme, PLC-gamma, is an excellent substrate for the epidermal growth factor (EGF) receptor both in vivo and in vitro. We now demonstrate that EGF treatment of HSC-1 cells, a human squamous cell carcinoma-derived cell line that expresses high levels of the EGF receptor, rapidly induces tyrosine phosphorylation of two-thirds of the total cellular PLC-gamma pool. A two-step immunoaffinity protocol was used for large-scale isolation of phosphorylated PLC-gamma from the cytosol of EGF-treated HSC-1 cells. Phosphorylated PLC-gamma was digested with trypsin, then phosphotyrosine-containing peptides were purified by phosphotyrosine affinity chromatography and reverse-phase high performance liquid chromatography. The two major phosphotyrosine-containing tryptic peptides were sequenced. Comparison of the sequence data with the bovine brain PLC-gamma amino acid sequence indicated that the major, EGF-sensitive tyrosine phosphorylation sites of human PLC-gamma correspond to the bovine brain PLC-gamma tyrosine residues 771 and 1254. The former residue is adjacent to regions of PLC-gamma that contain high homology to the non-catalytic, amino-terminal region of the src tyrosine kinase. The latter residue lies near the carboxyl terminus of the PLC-gamma molecule. The accompanying manuscript (Kim J.W., Sim, S.S., Kim, U-H., Nishibe, S., Wahl, M. I., Carpenter, G., and Rhe, S. G. (1990) J. Biol. Chem. 265, 3940-3943) identifies these same 2 residues plus 2 additional tyrosine phosphorylation sites through large-scale in vitro phosphorylation of purified bovine brain PLC-gamma by the EGF receptor.
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PMID:Identification of two epidermal growth factor-sensitive tyrosine phosphorylation sites of phospholipase C-gamma in intact HSC-1 cells. 168 11

Endothelin is a novel peptide secreted by endothelial cells, the vasoconstrictor effects of which appear dependent on the activation of phospholipase C. We examined in tissue culture its potential as a growth factor for vascular smooth muscle. In quiescent cultures of rat aortic smooth muscle cells, endothelin rapidly elevated levels of c-fos and c-myc mRNA. Peak effects on c-fos mRNA occurred between 15 and 30 min and were completely gone after 2 h. The elevation in c-fos mRNA was, in part, dependent on protein kinase C, since phorbol myristate acetate (PMA) also elevated c-fos mRNA and further increased c-fos mRNA expression by endothelin, but the effects were not additive. Furthermore, the endothelin-induced elevation in c-fos mRNA was attenuated but not abolished in protein kinase C-depleted cells. Maximum levels of c-myc mRNA occurred between 15 and 30 min after exposing the cells to endothelin and persisted for at least 6 h. The effects of simultaneous addition of endothelin and PMA on c-myc mRNA levels were essentially similar to those observed with c-fos mRNA. [3H]thymidine incorporation into DNA occurred 8 h after exposing the cells to endothelin. The mitogenic effect of endothelin was smaller than that observed with either fetal calf serum or epidermal growth factor and was dependent on both pertussis toxin-insensitive and -sensitive pathways. Sensitivity to the latter pathway did not appear dependent on attenuation of phospholipase C activity, since neither peak intracellular calcium concentrations nor c-fos mRNA levels were reduced in pertussis toxin-treated cells.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Growth factor activity of endothelin on vascular smooth muscle. 169 May 14

Forskolin-pretreatment of A431 cells reduced both intrinsic and epidermal growth factor (EGF)-induced EGF receptor phosphorylation, however, phosphorylation of phospholipase C-gamma (PLC-gamma) was stimulated under the same conditions. No significant difference was detected in the amount of phosphotyrosine of PLC-gamma between two cultures with or without forskolin treatment followed by EGF. On the other hand, phosphorylation of a 47 kDa protein (P47) which cross-reacted with an anti-PLC-gamma monoclonal antibody, was stimulated by both forskolin and EGF. Phosphorylation was exclusively on serine residues in this case. These results indicate that both PLC-gamma and P47 are phosphorylated by a cAMP-dependent protein kinase and the EGF-stimulated serine kinase, and suggest that serine phosphorylation of PLC-gamma has no effect on ligand-dependent coupling with the EGF receptor.
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PMID:Examination of the role of serine phosphorylation in phospholipase C-gamma and its related P47 in cAMP-mediated depression of epidermal growth factor signal transduction. 169 48

We have shown previously that exposure of a non-transformed continuous line of rat liver epithelial (WB) cells to epidermal growth factor (EGF), adrenaline, angiotensin II or [Arg8]vasopressin results in an accumulation of the inositol phosphates InsP1, InsP2 and InsP3 [Hepler, Earp & Harden (1988) J. Biol. Chem. 263, 7610-7619]. Studies were carried out with WB cells to determine whether the EGF receptor and other, non-tyrosine kinase, hormone receptors stimulate phosphoinositide hydrolysis by common, overlapping or separate pathways. The time courses for accumulation of inositol phosphates in response to angiotensin II and EGF were markedly different. Whereas angiotensin II stimulated a very rapid accumulation of inositol phosphates (maximal by 30 s), increases in the levels of inositol phosphates in response to EGF were measurable only following a 30 s lag period; maximal levels were attained by 7-8 min. Chelation of extracellular Ca2+ with EGTA did not modify this relative difference between angiotensin II and EGF in the time required to attain maximal phospholipase C activation. Under experimental conditions in which agonist-induced desensitization no longer occurred in these cells, the inositol phosphate responses to EGF and angiotensin II were additive, whereas those to angiotensin II and [Arg8]vasopressin were not additive. In crude WB lysates, angiotensin II, [Arg8]vasopressin and adrenaline each stimulated inositol phosphate formation in a guanine-nucleotide-dependent manner. In contrast, EGF failed to stimulate inositol phosphate formation in WB lysates in the presence or absence of guanosine 5'-[gamma-thio]triphosphate (GTP[S]), even though EGF retained the capacity to bind to and stimulate tyrosine phosphorylation of its own receptor. Pertussis toxin, at concentrations that fully ADP-ribosylate and functionally inactivate the inhibitory guanine-nucleotide regulatory protein of adenylate cyclase (Gi), had no effect on the capacity of EGF or hormones to stimulate inositol phosphate accumulation. In intact WB cells, the capacity of EGF, but not angiotensin II, to stimulate inositol phosphate accumulation was correlated with its capacity to stimulate tyrosine phosphorylation of the 148 kDa isoenzyme of phospholipase C. Taken together, these findings suggest that, whereas angiotensin II, [Arg8]vasopressin and alpha 1-adrenergic receptors are linked to activation of one or more phospholipase(s) C by an unidentified G-protein(s), the EGF receptor stimulates phosphoinositide hydrolysis by a different pathway, perhaps as a result of its capacity to stimulate tyrosine phosphorylation of phospholipase C-gamma.
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PMID:Evidence that the epidermal growth factor receptor and non-tyrosine kinase hormone receptors stimulate phosphoinositide hydrolysis by independent pathways. 169 55

Recent studies have shown that the receptor for epidermal growth factor (EGF) can associate with and tyrosine-phosphorylate the gamma-isozyme of phosphoinositide (PtdIns)-specific phospholipase C (PLC gamma), suggesting a possible mechanism for activation of PtdIns hydrolysis by EGF. In the present study, the coupling between PtdIns hydrolysis and PLC gamma tyrosine phosphorylation in WB liver epithelial cells was examined. Peak levels of [P-Tyr]PLC gamma, measured by anti-P-Tyr immunoblotting, occurred at 0.5-2 min of EGF treatment and coincided with the onset of [3H]inositol phosphate production. The termination of PtdIns hydrolysis after EGF stimulation was accompanied by return of [P-Tyr]PLC gamma to near-basal levels. Activation of protein kinase C (PKC) with a phorbol ester inhibited (IC50 = 3-10 nM) both EGF-dependent PtdIns hydrolysis and PLC gamma phosphorylation by more than 90%. Both EGF-stimulated responses were potentiated in cells depleted of PKC by prolonged phorbol ester treatment. At physiological ionic strength, monoclonal antibodies to PLC gamma specifically precipitated (in addition to PLC gamma) the EGF receptor and at least six other [P-Tyr]proteins from extracts of EGF-treated cells. PKC activation had differential effects on the tyrosine phosphorylation of these coprecipitating proteins, i.e. the relative abundance of certain [P-Tyr] proteins decreased, whereas that of another protein increased. In conclusion, EGF-stimulated tyrosine phosphorylation of PLC gamma is broadly correlated with stimulation of PtdIns hydrolysis, consistent with a role for tyrosine phosphorylation in PLC activation. The attendant diacylglycerol release and activation of PKC may terminate PLC gamma activation, in part by inhibiting PLC gamma phosphorylation by the EGF receptor. Our results suggest further that PKC may exert regulatory effects by altering the relationship of PLC gamma to its associated [P-Tyr]proteins.
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PMID:Protein kinase C inhibits epidermal growth factor-dependent tyrosine phosphorylation of phospholipase C gamma and activation of phosphoinositide hydrolysis. 169 45

Elevated eicosanoid biosynthesis characterizes certain forms of human and experimental glomerular proliferative disease. Thromboxane A2 (TxA2) and other prostaglandins (PG) act through specific receptors and mechanisms of intracellular signal transduction in human mesangial cells. We studied the actions of U-46619, a TxA2 mimetic which stimulates mesangial phospholipase C, and of the PGI2 analogue, Iloprost, a potent activator of adenylate cyclase, on proliferation of cultured human mesangial cells. When applied alone to quiescent cells, U-46619 had only weak mitogenic activity, as assessed by [3H]thymidine [( 3H]-TdR) incorporation and cell counts. On the other hand, addition of U-46619 10 minutes prior to stimulation of the cells with 1 to 17% fetal bovine serum (FBS) for 24 hours, potently and dose-dependently inhibited FBS-stimulated [3H]-TdR incorporation. Similarly, U-46619 inhibited the effects of 10 ng/ml platelet-derived growth factor (PDGF), epidermal growth factor or basic fibroblast growth factor on [3H]-TdR incorporation, by 55, 79 and 88%, respectively. The effects of U-46619 were not mimicked by another stimulus of phospholipase C, angiotensin II. Iloprost also inhibited FBS-activated proliferation. Neither eicosanoid inhibited the rise of cytosolic Ca2+ induced by FBS or PDGF. The actions of TxA2 and Iloprost in cultured cells point to multiple functional interactions between eicosanoids and growth factors in the control of mesangial cell proliferation.
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PMID:Regulation of human mesangial cell growth in culture by thromboxane A2 and prostacyclin. 169 33

Phospholipase C-gamma 1 (PLC-gamma 1), an isozyme of the phosphoinositide-specific phospholipase C family, which occupies a central role in hormonal signal transduction pathways, is an excellent substrate for the epidermal growth factor (EGF) receptor tyrosine kinase. Epidermal growth factor elicits tyrosine phosphorylation of PLC-gamma 1 and phosphatidylinositol 4,5-bisphosphate hydrolysis in various cell lines. The ability of tyrosine phosphorylation to activate the catalytic activity of PLC-gamma 1 was tested. Tyrosine phosphorylation in intact cells or in vitro increased the catalytic activity of PLC-gamma 1. Also, treatment of EGF-activated PLC-gamma 1 with a tyrosine-specific phosphatase substantially decreased the catalytic activity of PLC-gamma 1. These results suggest that the EGF-stimulated formation of inositol 1,4,5-trisphosphate and diacylglycerol in intact cells results, at least in part, from catalytic activation of PLC-gamma 1 through tyrosine phosphorylation.
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PMID:Increase of the catalytic activity of phospholipase C-gamma 1 by tyrosine phosphorylation. 170 Aug 66

Previous studies have demonstrated enhanced phosphorylation of phospholipase C-tau (PLC-tau), a key regulatory enzyme in phosphoinositide metabolism, in cells treated with platelet-derived growth factor (PDGF) and epidermal growth factor, both of which act via specific receptor tyrosine kinases. Our studies on BALB/c-3T3 cells show that agents that promote cellular cyclic AMP accumulation also increase the phosphorylation, specifically the serine phosphorylation, of this enzyme. Increased phosphorylation of PLC-t (2-3-fold) was evident within 5-10 min of addition of isobutylmethylxanthine (IBMX) and either cholera toxin or forskolin to cells, and persisted for at least 3 h. Treatment of cells with cyclic AMP agonists also enhanced, with similar kinetics, the phosphorylation of a 76 kDa protein co-precipitated by anti-PLC-tau monoclonal antibodies. Brief exposure of cells to cholera toxin/IBMX or forskolin/IBMX decreased inositol phosphate formation induced by the GTP-binding protein (G-protein) activator aluminium fluoride by approx. 50%, but was without effect on PDGF-stimulated inositol phosphate formation. These findings suggest that PLC-tau, and perhaps the 76 kDa co-precipitated protein, are substrates of cyclic AMP-dependent protein kinase in BALB/c-3T3 cells: however, the lack of effect of cyclic AMP elevation on PDGF-stimulated inositol phosphate formation indicates that the intrinsic activity of PLC-tau is unaltered by cyclic AMP-mediated phosphorylation.
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PMID:Cyclic AMP agonists induce the phosphorylation of phospholipase C-tau and of a 76 kDa protein co-precipitated by anti-(phospholipase C-tau) monoclonal antibodies in BALB/c-3T3 cells. Relationship to inositol phosphate formation. 170 22

Many hormones, neurotransmitters and growth factors, on binding to G protein-coupled receptors or receptors possessing tyrosine kinase activity, increase intracellular levels of the second messengers inositol 1,4,5-trisphosphate and 1,2-diacylglycerol. This is due to activation of phosphoinositide-specific phospholipase(s) C (PLC), the isozymes of which are classified into groups, alpha, beta, gamma and delta. The beta, gamma and delta groups themselves contain PLC isozymes which have both common and unique structural domains. Only the gamma 1 isozyme has been implicated in a signal transduction mechanism. This involves association with, and tyrosine phosphorylation by, the ligand-bound epidermal growth factor and platelet-derived growth factor receptors, probably by means of the PLC-gamma 1-specific src homology (SH2) domain. Because EGF receptor-mediated tyrosine phosphorylation of PLC-gamma 1 stimulates catalytic activity in vitro and G proteins have been implicated in the activation of PLC, we investigated which PLC isozymes are subject to G protein regulation. We have purified an activated G protein alpha subunit that stimulates partially purified phospholipase C and now report that this G protein specifically activates the beta 1 isozyme, but not the gamma 1 and delta 1 isozymes of phospholipase C. We also show that this protein is related to the Gq class of G protein alpha subunits.
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PMID:Activation of the beta 1 isozyme of phospholipase C by alpha subunits of the Gq class of G proteins. 170 1

Mutant epidermal growth factor (EGF) receptors (obtained by substitution of one, two or three C-terminal autophosphorylable tyrosine residues with phenylalanine residues or by deletion of the C-terminal 19 amino acids, including the distal tyrosine) were expressed in mouse NIH-3T3 fibroblast clones at densities comparable (less than 25% difference) with those in control clones expressing the wild-type receptor. Total EGF-induced phosphorylation of the mutated receptors was not appreciably changed with respect to controls, whereas autophosphorylation at tyrosine residues was decreased, especially in the double and the triple mutants. In the latter mutant, expression of the EGF-receptor-activated lipolytic enzyme phospholipase C gamma was unchanged, whereas its tyrosine phosphorylation induced by the growth factor was lowered to approx. 25% of that in the controls. In all of the cell clones employed, the accumulation of inositol phosphates induced by treatment with fetal calf serum varied only slightly, whereas the same effect induced by EGF was consistently lowered in those lines expressing mutated receptors. This decrease was moderate for those receptors missing only the distal tyrosine (point and deletion mutants), intermediate in the dual mutants and almost complete in the triple mutants. Likewise, increases in intracellular Ca2+ concentrations [( Ca2+]i) induced by fibroblast growth factor were approximately the same in all of the clones, whereas those induced by EGF were decreased in the mutants, again in proportion to the loss of the phosphorylable C-terminal tyrosine residues. The same trend occurred with membrane hyperpolarization, an effect secondary to the increase in [Ca2+]i via the activation of Ca2(+)-dependent K+ channels. We conclude that C-terminal autophosphorylable tyrosine residues play a positive role in the regulation of transmembrane signalling at the EGF receptor. The stepwise decrease in signal generation observed in single, double and triple point mutants suggest that the role of phosphotyrosine residues is not in the participation in specific amino acid sequences, but rather in the introduction of strong negative charges at strategic sites of the receptor tail. As a consequence of autophosphorylation, the receptor could become competent for specific association with phospholipase C gamma, with ensuing activation by tyrosine phosphorylation followed by the chains of intracellular responses ultimately leading to DNA synthesis and cell duplication.
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PMID:Transmembrane signalling at the epidermal growth factor receptor. Positive regulation by the C-terminal phosphotyrosine residues. 171 44


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