Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.4.3 (phospholipase C)
 18,461 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The human CSF-1 receptor (c-fms protooncogene product) was introduced into CSF-1-unresponsive Chinese hamster lung fibroblasts (CCL39 cell line) in order to study its coupling to biochemical signal-transducing systems and to compare the growth-regulating properties of CSF-1 to those of other growth factors. Independent clones expressing different levels of CSF-1 receptors were isolated and characterized. CSF-1 increased [3H]thymidine incorporation in serum-starved cells and potentiated the mitogenic effects of FGF and thrombin. As already observed for other growth factors activating receptor tyrosine kinases (EGF, FGF, IGF-I), CSF-1 alone did not trigger inositol phosphate formation, but slightly enhanced the activity of phospholipase C agonists (thrombin, A1F4- complex). Activation of the CSF-1 receptor by its ligand was evidenced by the rapid activation of the Na+/H+ exchanger resulting in amiloride-sensitive cytoplasmic alkalinization (0.1-0.2 pH units) within minutes after stimulation. Whereas pertussis toxin does not affect the action of EGF, FGF, or IGF-I in CCL39 cells, it partially inhibited both DNA synthesis reinitiation and activation of Na+/H+ exchange by CSF-1, indicating that the CSF-1 receptor can communicate with a signal-transducing GTP binding protein. A point-mutated form of the c-fms gene product, in which Tyr 969, a residue negatively modulating signal transduction, had been replaced with Phe [fms (F969)], did not generate responses significantly different from those obtained with the wild-type c-fms gene product. In the absence of CSF-1, cells expressing either wild-type or fms (F969) showed a considerably higher basal level of thymidine incorporation and decreased anchorage dependence compared with parental CCL39 cells. Monoclonal antibodies that interfere with signal transduction by the human CSF-1 receptor inhibited both basal [3H]thymidine incorporation and soft agar colony formation, indicating that relaxation of growth control was dependent on CSF-1 receptor expression.
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PMID:Functional expression of the human receptor for colony-stimulating factor 1 (CSF-1) in hamster fibroblasts: CSF-1 stimulates Na+/H+ exchange and DNA-synthesis in the absence of phosphoinositide breakdown. 215 62

The receptors involved in the regulation of phospholipase C by hormones, neurotransmitters and other ligands have seven transmembrane-spanning hydrophobic regions (seven-helix motif) and no known enzymatic activity. Furthermore these receptors can be isolated as complexes with guanine nucleotide binding (G) proteins. Guanine nucleotides affect the binding of hormones that stimulate phospholipase C and it has been possible to see activation of GTPase activity in membranes upon addition of these ligands. Further indirect evidence for a Gp (p stands for phospholipase C activation) protein is the finding that in membranes agonist activation of phospholipase C requires the presence of GTP gamma S a non-hydrolyzable analog of GTP. Furthermore, fluoride is able to activate phospholipase C but its inhibition of phosphatidylinositol-4' kinase (PI-4' kinase) can interfere with efforts to demonstrate this in intact cells. There are four major isozymes of phospholipase C that have been cloned and sequenced. Recently it was found that phospholipase C-gamma as well as PI-3'-kinase are substrates for phosphorylation on tyrosine residues by the EGF and PDGF receptors. The PI-3' kinase is able to convert phosphatidylinositol 4,5-bisphosphate (PIP2) to phosphatidylinositol 3,4,5-trisphosphate (PIP3) but the function of this lipid is unknown since it is not a substrate for any known phospholipase C. While much has been learned about the structure and regulation of the phosphoinositide specific kinases and phosphodiesterase enzymes this is a relatively new field in which we can expect many advances during the next few years.
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PMID:Regulation of phosphoinositide-specific phospholipase C. 216 88

Ligand stimulation of the platelet-derived growth factor receptor (PDGF-R) results in rapid activation of the receptor tyrosine kinase, stimulation of phosphoinositide hydrolysis, an increase in intracellular free Ca2+ concentration ([Ca2+]i), and, ultimately, cellular proliferation. In a previous study, we demonstrated that staurosporine, a known inhibitor of protein kinase C, blocked PDGF-induced [Ca2+]i increases in Swiss mouse 3T3 fibroblasts by a mechanism that appeared unrelated to inhibition of protein kinase activity (Olsen, R., Melder, D., Seewald, M., Abraham, R., and Powis, G. (1990) Biochem. Pharmacol. 39, 968-972). In the present study, we report that staurosporine inhibits ligand-dependent PDGF-R tyrosine kinase activation in cell-free receptor preparations and in intact Swiss 3T3 cells. At the same concentrations (10(-8)-10(-6) M), staurosporine suppressed both the tyrosine phosphorylation of phospholipase C activity and the hydrolysis of phosphoinositides induced by PDGF stimulation of intact cells. In contrast, guanine nucleotide-binding protein-dependent phospholipase C activation induced by bradykinin or fluoroaluminate anion was relatively insensitive to staurosporine. A preferential inhibitory effect of staurosporine on signal generation by the PDGF-R was indicated by findings that epidermal growth factor receptor (EGF-R) tyrosine kinase activity and EGF-dependent phospholipase C in A-431 carcinoma cells were approximately 100-fold less sensitive to this drug. These data indicate that submicromolar concentrations of staurosporine inhibit PDGF-dependent phosphoinositide hydrolysis and Ca2+ mobilization through a proximal inhibitory effect on ligand-induced activation of the PDGF-R tyrosine kinase.
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PMID:Preferential inhibition of the platelet-derived growth factor receptor tyrosine kinase by staurosporine. 217 5

Binding of EGF to cells expressing human EGF receptor stimulated rapid tyrosine phosphorylation of phospholipase C-II (PLC-II), as revealed by immunoblotting analysis with phosphotyrosine-specific antibodies. Tyrosine phosphorylation of PLC-II was stimulated by low physiological concentrations of EGF (1 nM), was quantitative, and was already maximal after a 30 sec incubation with 50 nM EGF at 37 degrees C. Interestingly, antibodies specific for PLC-II were able to coimmunoprecipitate the EGF receptor and antibodies against EGF receptor also coimmunoprecipitated PLC-II. According to this analysis, approximately 1% of EGF receptor molecules were associated with PLC-II molecules. The protein tyrosine kinase inhibitor tyrphostin RG50864, which blocks EGF-dependent cell proliferation, blocked EGF-induced tyrosine phosphorylation of PLC-II, its association with EGF receptor, and EGF-induced Ca2+ release. Hence, EGF-induced tyrosine phosphorylation of PLC-II may be a regulatory event linking the tyrosine kinase activity of EGF receptor to the PIP2 hydrolysis signaling pathway.
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PMID:EGF induces tyrosine phosphorylation of phospholipase C-II: a potential mechanism for EGF receptor signaling. 247 18

This paper has reviewed, in a broad sense, the potential involvement of the oncogenes and their progenitors, the protooncogenes, in signal transduction pathways. The membrane-associated oncogene products appear to be connected with the generation and/or regulation of secondary messengers, particularly those associated with Ca2+/phospholipid-dependent activation of the serine/threonine kinase protein kinase C. Activation of transmembrane receptors, either through binding their native ligand or through point mutations that lead to constitutive expression, results in the expression of their intrinsic tyrosine-specific protein kinases. In PDGF-stimulated cells, this results in the increased turnover of phosphatidylinositols and the subsequent release of IP3 (Habenicht et al., 1981; Berridge et al., 1984). This coincides with activation of a PI kinase activity (Kaplan et al., 1987). Likewise, the fms product, which is the receptor for CSF-1, induces a guanine nucleotide-dependent activation of phospholipase C (Jackowski et al., 1986). Receptor functions are potentially regulated through differential binding of ligands (as proposed with PDGF), through interactions with other receptors, and through the "feedback" regulation mediated by protein kinase C. PDGF stimulation leads to modulation of the EGF receptor through protein kinase C (Bowen-Pope et al., 1983; Collins et al., 1983; Davis and Czech, 1985). Similarly, the neu product becomes phosphorylated on tyrosine residues following treatment of cells with EGF, although the neu protein does not bind EGF itself (King et al., 1988; Stern and Kamps, 1988). The tyrosine kinases of the src family are not receptors themselves, although they may mediate specific receptor-generated signals. The clck product is physically and functionally associated with the T-cell receptors CD4 and CD8, and becomes active upon specific stimulation of cells expressing those markers (Veillette et al., 1988a,b). The precise physiological role of the src family products has not been established, but their kinase activity is intrinsic to that function. The v- and c-src products are hyperphosphorylated during mitosis (Chackalaparampil and Shalloway, 1988), which correlates with periods of reduced cell-to-cell adhesion and communication (Warren and Nelson, 1987; Azarnia et al., 1988). Furthermore, pp60c-src is associated with a PI kinase activity when complexed with MTAg of polyoma virus, suggesting a function in stimulating increased turnover of the phosphatidylinositols (Heber and Courtneidge, 1987; Kaplan et al., 1987).(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Oncogenes, protooncogenes, and signal transduction: toward a unified theory? 269 May 95

Stable expression of high levels of activated forms of Haras (T24) or v-Ki-ras by transfection of Chinese hamster lung fibroblasts (CCL39) yielded cells highly tumorigenic in nude mice. Two classes of transformed cells were distinguished, one with moderate p21 expression (10-fold increased) had retained growth factor dependency, the second with higher level of p21 (greater than 50-fold) appeared autonomous for growth. Neither class of transformants expressing Ki-ras or Ha-ras displayed a significant basal activity of polyphosphoinositide-specific phospholipase C, measured either in serum-starved cells or during exponential growth in the presence of growth factors of the tyrosine kinase family (EGF, FGF, insulin). In the growth-factor-dependent class of T24-Ha-ras-transfected cells (clone 39THaB), phospholipase C could be stimulated normally by serum, thrombin and AlF-4. In the more growth autonomous class (clones 39THaC and 39Ki9), release of inositol phosphates after stimulation with thrombin or serum was drastically reduced. This desensitization, apparently at the receptor level since the response to AlF-4 persisted, is, however, not specific to ras expression. We observed it to the same degree in polyoma virus-transformed CCL39 cells. Finally, expression of mutated forms of p21 ras did not abrogate the sensitivity of phospholipase C activation to pertussis toxin. We conclude that the transforming potential of activated forms of p21ras does not result from persistent activation of phospholipase C and that ras GTP-binding proteins cannot substitute for Gp.
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PMID:Deregulation of hamster fibroblast proliferation by mutated ras oncogenes is not mediated by constitutive activation of phosphoinositide-specific phospholipase C. 283

Interactions between the different signaling roles of myo-inositol 1,4,5-trisphosphate and 1,2-diacylglycerol, the products of agonist-stimulated phosphatidylinositol 4,5-bisphosphate breakdown, are assessed in isolated rat hepatocytes. Measurements of the kinetics of accumulation of individual [3H]inositol phosphates after the addition of different Ca2+-mobilizing agonists in general support the role of inositol 1,4,5-trisphosphate as the second messenger responsible for release of sequestered intracellular Ca2+. Various agonists, when added at maximal concentrations, however, produce qualitatively and quantitatively different responses, which reflect varying abilities of the agonists to activate phospholipase C. Qualitative differences are revealed by a pronounced biphasic pattern to the Ins(1,4,5)P3 accumulation after vasopressin and phenylephrine stimulation, which is indicative of negative feedback. It is suggested that this effect is mediated by a partial diacylglycerol activation of protein kinase C, which in vitro causes an activation of inositol phosphate 5-phosphatase and hence promotes removal of Ins(1,4,5)P3 to Ins(1,4)P2. An alternative mechanism proposed by Biden and Wollheim (1986) of a secondary Ca2+ activation of Ins(1,4,5)P3 3-kinase is considered less likely as a general mechanism, since highly purified kinase prepared from rat brain shows only an inhibition by Ca2+. Glucagon, 8-Br-cAMP, and EGF induce small increases of Ins(1,4,5)P3 in hepatocytes, together with slower and smaller increases of cytosolic free Ca2+ than those produced by vasopressin or phenylephrine, with Ca2+ being mobilized from the same intracellular pools with each of the agonists. The Ca2+-mobilizing effect of glucagon, therefore, may be entirely due to a cAMP-dependent process, although a direct receptor-mediated activation of phospholipase C, as suggested by Wakelam et al. (1986), remains a possibility. The EGF receptor appears to be coupled to phospholipase C, presumably via a G-protein. It is speculated that the mechanism by which cAMP increases Ins(1,4,5)P3 levels in hepatocytes could either be by phosphorylation and inhibition of inositol phosphate 5-phosphatase or by phosphorylation and facilitation of the coupling between the G-protein and phospholipase C. When protein kinase C is maximally activated by pretreatment of hepatocytes with PMA, the stimulatory effects of phenylephrine, glucagon, 8-Br-cAMP, and EGF on the accumulation of inositol phosphates and increase of cytosolic free Ca2+ are largely inhibited.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Mechanisms involved in receptor-mediated changes of intracellular Ca2+ in liver. 285 Jun 13

The mechanisms of growth factor action were studied in a fibroblastic cell line capable of reversible growth arrest in G0-G1. This cell line, derived from Chinese hamster lung, can be stimulated to divide by a limited set of purified growth factors, including EGF, FGF, PDGF, alpha-thrombin (THR), serotonin (5-HT) and insulin. THR and 5-HT stimulate, via a G-protein (Gp), a polyphosphoinositide-specific phospholipase C (PtdIns(4,5)P2-PLC). In contrast, the mitogens EGF, FGF, PDGF, and insulin do not stimulate PtdIns(4,5)P2-PLC unless this pathway has been preactivated by THR or AlF-4. Finally, from the specific inhibitory action of pertussis toxin on THR- and 5-HT-induced DNA synthesis, and from the exploitation of the 5-HT pharmacological tools, we conclude that: (i) there are at least two distinct G-proteins involved in signalling growth: Gp, coupling receptors to PtdIns(4,5)P2-PLC, and Gi, coupling receptors negatively to adenylyl cyclase and probably to other unknown effector(s); (ii) activation of receptor-tyrosine kinases provides an alternate growth factor signalling pathway, independent of Gp- and Gi-mediated actions; and (iii) tyrosine kinases positively 'cross-communicate' with the inositol-lipid pathway (phosphorylation of Gp, PLC, PtdIns kinases...?).
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PMID:Transmembrane signalling pathways initiating cell growth in fibroblasts. 290 48

Lipocortin I, a 35-kDa protein, has been detected in terminally differentiated monocytes and neutrophils. This calcium-phospholipid binding protein appears to be identical to a 35-kDa protein that can serve as a substrate for the EGF-receptor/tyrosine kinase. We have used the human myelocytic cell line HL-60 to explore whether differentiation of hematopoietic cells is associated with changes in the level of lipocortin I. We find that differentiation of HL-60 cells toward the macrophage lineage by the addition of phorbol esters or vitamin D3 or toward neutrophils with dibutyryl cyclic AMP or dimethyl sulfoxide is accompanied by an increase in the cellular content of lipocortin I. In comparison, treatment of HL-60 cells with bryostatin 1, a compound that activates protein kinase C but does not differentiate HL-60 cells, did not effect the level of 35 kDa protein. We have developed a radioimmunoassay to quantitate this protein by using a polyclonal antibody to a synthetic amino terminal peptide of the 35-kDa protein. This antibody recognizes purified pig lung 35-kDa protein as well as a single 35-kDa protein in HL-60 and A-431 cells as determined by Western blotting and immune precipitation. Differentiated HL-60 cells contain 2.6-fold the amount of 35-kDa protein found in undifferentiated HL-60 cells. Our findings that the addition of phorbol esters to HL-60 cells results in an increase in the mRNA for the 35-kDa protein and in an increase in the incorporation of 35S-methionine into the protein suggest that transcriptional activation or increased stability of the mRNA is responsible for the increased rate of synthesis and accumulation of lipocortin I during differentiation of these cells. In the absence of added divalent cations, we have determined that in differentiated HL-60 cells 79% of lipocortin I protein is located in the cytosol while 21% of the total cellular protein is bound to the particulate fraction. The 35-kDa protein can be removed from the particulate fraction by incubation with chelators or treatment with phospholipase A2 or phospholipase C. Addition of the calcium ionophore A23187 to intact differentiated HL-60 cells causes the 35-kDa protein to associate with the particulate fraction of the cell, suggesting that modulation of intracellular calcium levels may play a role in changing the intracellular location of this protein.
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PMID:Differentiation of HL-60 cells is associated with an increase in the 35-kDa protein lipocortin I. 297 67

Prior treatment with pertussis toxin of G0-arrested hamster fibroblasts (CCL39) results in a dose-dependent inhibition of two early events of the mitogenic response elicited by alpha-thrombin: accumulation of inositol phosphates in Li+-treated cells, and activation of the Na+/H+ antiport, measured either by the amiloride-sensitive 22Na+ influx or by the increase in intracellular pH. At 10(-1) U/ml of alpha-thrombin, the maximal inhibition was approximately 50% for these two early cellular responses, but the pertussis toxin effect was more pronounced at lower thrombin concentrations. In contrast, pertussis toxin does not affect the Na+/H+ antiport activation induced by phorbol esters or EGF, the action of which is not mediated by the phosphoinositide-metabolizing pathway in CCL39 cells. Therefore, our data suggest the following. A GTP-binding regulatory protein is probably involved in signal transduction between thrombin receptors and the phosphatidylinositol 4,5-bisphosphate-specific phospholipase C. This regulation does not seem to be exerted via modulations of cyclic AMP levels. The thrombin-induced activation of Na+/H+ antiport is, at least in part, mediated by the protein kinase C, as a consequence of stimulation of phosphatidylinositol turnover.
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PMID:Pertussis toxin inhibits thrombin-induced activation of phosphoinositide hydrolysis and Na+/H+ exchange in hamster fibroblasts. 300 18


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