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)

Phosphatidylinositol-specific phospholipase C isozyme gamma (PLC-gamma, Mr 145,000) is an excellent substrate for the epidermal growth factor (EGF) receptor both in vivo and in vitro. PLC-beta-1, another PLC isozyme, is a poor substrate for the EGF receptor. We examined the relative phosphorylation of PLC-gamma and PLC-beta-1 by the 170-kDa native EGF receptor molecule, the 66-kDa cytoplasmic kinase domain of the EGF receptor (Arg647-Ala1186), the alpha 2 beta 2 native insulin receptor, and the 48-kDa cytoplasmic kinase domain of the insulin receptor beta subunit (Gly947-Ser1343). Similar to the intact EGF receptor, the cytoplasmic kinase domain of the EGF receptor preferentially phosphorylated PLC-gamma. High-performance liquid chromatographic comparison of tryptic phosphopeptides from PLC-gamma phosphorylated by both forms of the EGF receptor kinase indicated similar patterns of multiple tyrosine phosphorylations. These results imply that substrate selectivity, at least in terms of PLC isozymes, is independent of the extracellular ligand-binding and membrane anchor domains of the EGF receptor. In comparison, neither the intact insulin receptor nor the beta-chain kinase domain was able to phosphorylate PLC-gamma to a significant extent. Also, insulin failed to stimulate the phosphorylation of PLC-gamma in NIH 3T3/HIR cells, which overexpress the human insulin receptor. Thus PLC-gamma is not a phosphorylation substrate for the insulin receptor in vitro or in the intact cell.
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PMID:Selectivity of phospholipase C phosphorylation by the epidermal growth factor receptor, the insulin receptor, and their cytoplasmic domains. 215 2

Epidermal growth factor (EGF) treatment of NIH 3T3 cells transfected with wild-type EGF receptor induced tyrosine phosphorylation of phospholipase C-gamma (PLC-gamma). The EGF receptor and PLC-gamma were found to be physically associated such that antibodies directed against PLC-gamma or the EGF receptor coimmunoprecipitated both proteins. The association between PLC-gamma and wild-type EGF receptor was dependent on the concentration of EGF, but EGF did not enhance the association between PLC-gamma and a kinase-negative mutant of the EGF receptor. Oligomerization of the EGF receptor was not sufficient to induce association of the EGF receptor with PLC-gamma, since the kinase-negative mutant receptor underwent normal dimerization in response to EGF yet did not associate with PLC-gamma. The form of PLC-gamma associated with the EGF receptor appeared to be primarily the non-tyrosine-phosphorylated form. It is concluded that the kinase activity of the EGF receptor is essential for association of PLC-gamma with the EGF receptor, possibly by stimulating receptor autophosphorylation.
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PMID:Tyrosine kinase activity is essential for the association of phospholipase C-gamma with the epidermal growth factor receptor. 215 14

Studies were performed to examine a potential role for a guanine nucleotide-binding protein in epidermal growth factor (EGF)-stimulated phospholipase A2 (PLA2) activity. EGF increased prostaglandin E2 (PGE2) production in intact or saponin-permeabilized rat inner medullary collecting tubule (RIMCT) cells. Incubation of permeabilized cells with guanosine 5'-O-(thiotriphosphate) (GTP gamma S) enhanced and with guanosine 5'-O-(2-thiodiphosphate) (GDP beta S) inhibited the response to EGF. GDP beta S had no effect on ionomycin-stimulated PGE2 production. Exposure of intact cells to 25 mM NaF + 10 microM AlCl3 enhanced both basal and EGF-stimulated PGE2 production. Pertussis toxin ADP-ribosylated a 41-kDa protein in RIMCT cell membranes. Pretreatment of cells with pertussis toxin (100 ng/ml for 16 h) eliminated the response to EGF in intact cells and the response to EGF + GTP gamma S in permeabilized cells. Pertussis toxin had no effect on the response to ionomycin. The effect of pertussis toxin was not due to alterations in cAMP as cellular cAMP levels were unaffected by pertussis toxin both in the basal state and in the presence of EGF. PGE2 production in response to EGF was not transduced by a G protein coupled to phospholipase C (PLC) as neomycin, which inhibited PLC, did not decrease EGF-stimulated PGE2 production. Also, PGE2 production was not increased by inositol trisphosphate and did not require the presence of extracellular Ca2+. In contrast to EGF-stimulated PLC activity, stimulation of PLA2 by EGF was not susceptible to inhibition by phorbol 12-myristate 13-acetate. These results clearly demonstrate the existence of a PLA2-specific pertussis toxin-inhibitable guanine nucleotide-binding protein coupled to the EGF receptor in RIMCT cells.
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PMID:The epidermal growth factor receptor is coupled to a phospholipase A2-specific pertussis toxin-inhibitable guanine nucleotide-binding regulatory protein in cultured rat inner medullary collecting tubule cells. 215 14

Swiss 3T3 cells express receptors for both the polypeptide epidermal growth factor (EGF) and the tetradecapeptide bombesin and respond mitogenically to these substances. These cells thus provide a system to analyze potential signal transduction pathways involved in mitogenic stimulation. Here we have determined and compared the early ionic responses elicited by EGF and bombesin and their relation to diacylglycerol (DG) and inositolphosphate (InsPn) production. Whereas EGF fails to cause any significant change in intracellular Ca2+, bombesin effectively induces prompt and transient Ca2+ mobilization from intracellular stores. Further support of the idea that these receptors utilize distinct signalling pathways comes from the measurements of cytoplasmic pH (pHi). As in most target cells, EGF induces a delayed (1 min) but sustained intracellular alkalinization that reaches a new steady state after approximately 10 min. Bombesin, in contrast, elicits a biphasic response; within seconds, a rapid but transient rise in pHi is observed, followed by a further slower sustained alkalinization. Inhibition of the Na+/H+ exchanger prevents both EGF as well as bombesin-induced alkalinization. However, under these conditions, bombesin evokes a rapid and sustained acidification related to the Ca2+ response. Apparently, bombesin initiates a Ca2(+)-dependent acidifying process immediately after binding of the hormone to its receptor. Furthermore, we could demonstrate that the bombesin-induced alkalinization depends on protein kinase C activation whereas the EGF response does not. Determination of the total DG and InsPn accumulation revealed that EGF is ineffective in stimulating phospholipase C-mediated production of these second messengers. In contrast, bombesin causes a rapid DG and InsPn production coinciding with the Ca2+ response and the first phase of the rise in pHi followed by a slower DG accumulation coinciding with the second alkalinization phase. Our results show that in Swiss 3T3 cells the bombesin receptor activates the hydrolysis of inositol lipids as a mechanism of signal transduction, which consequently causes changes in Ca2+i and pHi. Clearly, the EGF receptor utilizes different pathways to evoke mitogenesis and stimulates Na+/H+ exchange independently of DG production and protein kinase C activation.
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PMID:Epidermal growth factor and bombesin differ strikingly in the induction of early responses in Swiss 3T3 cells. 215 9

We have previously reported that epidermal growth factor (EGF) activates phospholipase A2 (PLA2) independently of phospholipase C (PLC) in renal mesangial cells. In this study we use NIH 3T3 cell lines transfected with the normal EGF receptor (HER14 cells) or with EGF receptor defective in tyrosine kinase activity (K721A cells), to determine whether the intrinsic tyrosine kinase activity of the EGF receptor is required for the PLC-independent activation of PLA2. Intact cells were preincubated with EGF or other ligands, and then PLA2 activity was assayed in cell-free extracts with 1-stearoyl-2-[14C]arachidonyl phosphatidylcholine as the substrate. In HER14 cells, EGF increased PLA2 activity by 226 +/- 30%, and the tumour promoter phorbol myristate acetate (PMA) increased activity by 223 +/- 30%. The effect of EGF was not mediated through protein kinase C (PKC), whose activation by EGF requires tyrosine kinase activity, since raising intracellular Ca2+ alone with the Ca2+ ionophore A23187 did not mimic its effect, and the effect of EGF persisted in PKC-down-regulated cells. In K721A cells EGF was ineffective, whereas PMA was still active. Furthermore, in intact HER14 cells prelabelled with [14C]arachidonate, EGF-stimulated release of [14C]arachidonic acid was synergistic with A23187, but was unaccompanied by a rise in [14C]diacylglycerol. EGF had no effect on [14C]arachidonic acid release in intact K721A cells. We conclude that the tyrosine kinase activity of the EGF receptor is necessary for the PLC-independent stimulation of PLA2 by EGF.
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PMID:The tyrosine kinase activity of the epidermal-growth-factor receptor is necessary for phospholipase A2 activation. 215 83

Treatment of HER 14 cells with epidermal growth factor (EGF) or platelet-derived growth factor (PDGF) induced a translocation of phospholipase C-gamma (PLC-gamma) from cytosol to membrane. In such growth factor-treated cells, cytosolic PLC-gamma was found to contain more phosphotyrosine than membrane-associated enzyme. Because these growth factors have been shown to promote both the physical association of PLC-gamma with their receptors and the subsequent phosphorylation of the enzyme directly by the membrane-bound receptor tyrosine kinases, the membrane association of PLC-gamma may simply be due to the formation of transient enzyme (receptor)-substrate (PLC-gamma) complexes. If this is the case, membrane-associated PLC-gamma would be expected to be released from membrane after undergoing tyrosine phosphorylation. However, tyrosine phosphorylation of membrane-associated PLC-gamma by the EGF receptor in vitro did not result in the release of PLC-gamma from membrane. Thus, the association of PLC-gamma with membrane would appear to involve more than enzyme-substrate complex.
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PMID:Epidermal growth factor and platelet-derived growth factor promote translocation of phospholipase C-gamma from cytosol to membrane. 217 93

Cytoplasmic proteins that regulate signal transduction or induce cellular transformation, including cytoplasmic protein-tyrosine kinases, p21ras GTPase-activating protein (GAP), phospholipase C gamma, and the v-crk oncoprotein, possess one or two copies of a conserved noncatalytic domain, Src homology region 2 (SH2). Here we provide direct evidence that SH2 domains can mediate the interactions of these diverse signaling proteins with a related set of phosphotyrosine ligands, including the epidermal growth factor (EGF) receptor. In src-transformed cells GAP forms heteromeric complexes, notably with a highly tyrosine phosphorylated 62-kDa protein (p62). The stable association between GAP and p62 can be specifically reconstituted in vitro by using a bacterial polypeptide containing only the N-terminal GAP SH2 domain. The efficient phosphorylation of p62 by the v-Src or v-Fps tyrosine kinases depends, in turn, on their SH2 domains and correlates with their transforming activity. In lysates of EGF-stimulated cells, the N-terminal GAP SH2 domain binds to both the EGF receptor and p62. Fusion proteins containing GAP or v-Crk SH2 domains complex with similar phosphotyrosine proteins from src-transformed or EGF-stimulated cells but with different efficiencies. SH2 sequences, therefore, form autonomous domains that direct signaling proteins, such as GAP, to bind specific phosphotyrosine-containing polypeptides. By promoting the formation of these complexes, SH2 domains are ideally suited to regulate the activation of intracellular signaling pathways by growth factors.
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PMID:Src homology region 2 domains direct protein-protein interactions in signal transduction. 223 73

The biological responses of epidermal growth factor (EGF) are mediated by a surface receptor denoted as the EGF receptor. The EGF receptor possesses intrinsic protein tyrosine kinase activity which is essential for signal transduction. Recent evidence shows that EGF receptor phosphorylates several substances including: phospholipase C-gamma and the GTPase-activating protein (GAP). Moreover, these proteins become associated with the activated receptor in an immunocomplex. Autophosphorylation of the EGF receptor appears to be required for the association with phospholipase C-gamma. Mutational analysis indicates that the intrinsic autophosphorylation sites compete with exogenous substrates for the substrate-binding site in the kinase domain. The ligand-binding site for EGF was analysed using a chimeric receptor approach. Subdomains of the extracellular ligand-binding region of the chicken EGF receptor, which binds EGF with low affinity, were replaced by corresponding regions of the human EGF receptor, which binds EGF with high affinity. On the basis of this analysis, it is concluded that subdomain III of the extracellular domain of the EGF receptor is a major ligand-binding domain. Together, domain I and domain III are able to reconstitute nearly all interactions which bring about high-affinity binding. Growth factor receptors with protein tyrosine kinase (PTK) activity could be envisioned as membrane-associated allosteric enzymes. Unlike water-soluble allosteric enzymes, the configuration of the growth factor receptors dictates that the ligand-binding domain and PTK activity of the receptor molecules are separated by the plasma membrane. Therefore, ligand-induced signal must cross the membrane barrier to activate the PTK function.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Mutational analysis of the epidermal growth factor-receptor kinase. 225 59

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

Phosphatidylinositol (PI) kinase is activated by growth factors, such as epidermal growth factor (EGF), and is thought to be involved in cellular proliferation. Psoriasis is a hyperproliferative epidermal disease in which EGF receptor expression is altered and phospholipase C activity is increased. Considering the potential importance of growth factor stimulated phosphoinositide metabolism in the genesis of abnormal growth, we measured PI kinase activity in epidermal keratome biopsies from normal skin and the lesional and nonlesional skin of psoriatic patients. The PI kinase activity in 10 psoriatic involved plaques was increased 6.7-fold (Vmax = 67.1 +/- 23.9 pmol formed/min/mg protein +/- SE) when compared with 11 normal epidermal biopsies (Vmax = 10.0 +/- 1.3 pmol/min/mg protein, p less than 0.025). Similar results were noted when enzyme activity was standardized using DNA content. The apparent Km of PI kinase for ATP in involved psoriatic biopsies (0.45 +/- 0.14 mM) was also significantly (p less than 0.025) increased compared with normals (0.11 +/- 0.02 mM). The PI kinase activity in 11 biopsies of nonlesional psoriatic epidermis was not statistically different from normal epidermis. Both psoriatic and normal PI kinases required Mg++ and were inhibited by Ca++. The polyamine, spermine, a known activator of PI kinase in other tissues, stimulated normal but not psoriatic epidermal PI kinase. Both normal and psoriatic PI kinase activities had an apparent mol wt of 85,000. Increased synthesis of phosphoinositides by PI kinase in psoriatic tissue may provide more substrate for phospholipase C; a key enzyme in growth factor-mediated signal transduction.
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PMID:Increased phosphatidylinositol kinase activity in psoriatic epidermis. 254 14


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