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)

Signalling proteins such as phospholipase C-gamma (PLC-gamma) or GTPase-activating protein (GAP) of ras contain conserved regions of approximately 100 amino acids termed src homology 2 (SH2) domains. SH2 domains were shown to be responsible for mediating association between signalling proteins and tyrosine-phosphorylated proteins, including growth factor receptors. Nck is an ubiquitously expressed protein consisting exclusively of one SH2 and three SH3 domains. Here we show that epidermal growth factor or platelet-derived growth factor stimulation of intact human or murine cells leads to phosphorylation of Nck protein on tyrosine, serine, and threonine residues. Similar stimulation of Nck phosphorylation was detected upon activation of rat basophilic leukemia RBL-2H3 cells by cross-linking of the high-affinity immunoglobulin E receptors (Fc epsilon RI). Ligand-activated, tyrosine-autophosphorylated platelet-derived growth factor or epidermal growth factor receptors were coimmunoprecipitated with anti-Nck antibodies, and the association with either receptor molecule was mediated by the SH2 domain of Nck. Addition of phorbol ester was also able to stimulate Nck phosphorylation on serine residues. However, growth factor-induced serine/threonine phosphorylation of Nck was not mediated by protein kinase C. Interestingly, approximately fivefold overexpression of Nck in NIH 3T3 cells resulted in formation of oncogenic foci. These results show that Nck is an oncogenic protein and a common target for the action of different surface receptors. Nck probably functions as an adaptor protein which links surface receptors with tyrosine kinase activity to downstream signalling pathways involved in the control of cell proliferation.
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PMID:The SH2 and SH3 domain-containing Nck protein is oncogenic and a common target for phosphorylation by different surface receptors. 133 47

Treatment of Swiss 3T3 cells with a subsaturating concentration of recombinant Pasteurella multocida toxin (rPMT) markedly potentiated the production of inositol phosphates induced by bombesin, vasopressin, and endothelin but not by platelet-derived growth factor (PDGF) (AA and BB homodimers). Similarly, the neuropeptides but not PDGF caused a shift in the dose-dependent increase in inositol phosphates induced by rPMT. The rate of accumulation of inositol phosphates induced by bombesin was increased 2-fold by rPMT treatment while that of PDGF was unaffected. rPMT treatment also enhanced bombesin-induced inositol(1,4,5)trisphosphate, the direct product of phosphatidylinositol 4,5-bisphosphate hydrolysis. In contrast, treatment of cells with rPMT had no effect on the tyrosine phosphorylation of phospholipase C gamma. Depletion of protein kinase C increased rPMT-induced inositol phosphates in a manner similar to that observed for bombesin but not PDGF. Thus, rPMT selectively potentiates neuropeptide-mediated inositol phosphate production. The action of rPMT on phosphatidylinositol 4,5-bisphosphate hydrolysis persisted in streptolysin O-permeabilized cells. Addition of guanosine 5'-O-(beta-thiodiphosphate) to permeabilized cells markedly reduced rPMT-induced inositol phosphates in a time- and dose-dependent manner. rPMT also increased the sensitivity of phospholipase C for free calcium. Our results strongly suggest that the action of rPMT facilitates the coupling of G protein to phospholipase C.
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PMID:Pasteurella multocida toxin selectively facilitates phosphatidylinositol 4,5-bisphosphate hydrolysis by bombesin, vasopressin, and endothelin. Requirement for a functional G protein. 133 89

The HER2/neu gene encodes a receptor tyrosine kinase that is highly homologous to the epidermal growth factor receptor. Overexpression of the receptor in mammary and ovarian carcinoma correlates with poor patient prognosis. To determine how the overexpression of a normal receptor leads to the generation of an oncogenic signal, we compared the patterns of tyrosine phosphorylation in tumor-derived human cell lines expressing high levels of p185HER2/neu. In intact SKBR3 cells, basal phosphorylation of p185HER2/neu was not detected. However, pretreatment of cells with the tyrosine phosphatase inhibitor, sodium orthovanadate, led to the detection of phosphotyrosine on phospholipase C-gamma (PLC-gamma), GTPase-activating protein but not on the RAF-1 kinase. Strikingly, PLC-gamma was detected in a complex which contained multiple tyrosine-phosphorylated polypeptides. This complex was detected only in cytoplasmic fractions and had a distinct composition in different p185HER2/neu-overexpressing cell lines. Although GTPase-activating protein has been found previously in association with proteins of 190 and 62 kDa in fibroblasts, in SKBR3 cells it was found associated with multiple additional tyrosine-phosphorylated polypeptides. These experiments show that SKBR3 cells possess high levels of protein tyrosine phosphatase that can act upon p185HER2/neu. Moreover, they reveal, for the first time, the presence of PLC-gamma and GTPase-activating protein in cytosolic complexes containing a variety of other tyrosine-phosphorylated polypeptides. These observations suggest novel possibilities for the specific definition of receptor-generated signals in tumor cells.
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PMID:Tyrosine phosphatase inhibition permits analysis of signal transduction complexes in p185HER2/neu-overexpressing human tumor cells. 134 42

Ligand-induced dimerization of growth factor receptors is crucial for stimulation of their intrinsic protein tyrosine kinase activity promoting receptor autophosphorylation by an intermolecular mechanism. Moreover, the suppressive and negative dominant action of defective epidermal growth factor receptor (EGFR) was shown to be caused by formation of inactive heterodimers with normal EGFR leading to diminished biological signaling. In this report we explore the structural requirements and functional significance of heterodimerization between EGFR and HER2. HER2 (also called c-erbB-2 or neu) is a member of the EGFR family whose natural ligand is still unknown. We show that in response to EGF, wild type EGFR and various EGFR mutants were able to undergo heterodimerization with HER2. Addition of EGF to transfected cells co-expressing HER2 with a kinase negative point mutant of EGFR (K721A) stimulated heterodimer formation, tyrosine phosphorylation of K721A and HER2, and tyrosine phosphorylation of one of their known substrates, phospholipase C gamma. However, the binding of EGF to transfected cells co-expressing HER2 together with another EGFR mutant CD533 (a deletion mutant lacking most of the cytoplasmic domain of EGFR) caused heterodimerization and inhibition of tyrosine kinase activity. It appears therefore that EGF-induced heterodimerization of EGFR and HER2 can promote either stimulatory or inhibitory influences on kinase activity. We propose that the nature of receptor interactions on the cell surface can either activate or inhibit the initiation of growth factor-controlled cellular signaling.
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PMID:Heterodimerization of c-erbB2 with different epidermal growth factor receptor mutants elicits stimulatory or inhibitory responses. 134 15

The neu protooncogene encodes a tyrosine kinase receptor that is involved in the regulation of normal growth and malignant transformation. To circumvent the use of the incompletely characterized ligand of Neu, we constructed a chimeric protein composed of the ligand-binding domain of the epidermal growth factor receptor and the transmembrane and cytoplasmic portions of Neu. By expressing this Neu-epidermal growth factor receptor chimera (termed NEC), we found that following stimulation by the heterologous ligand, the tyrosine kinase of Neu became associated with a phosphatidylinositol (PI) kinase activity. The association was dependent on the concentration of the ligand and was almost maximal within 30 s after ligand binding. The lipid kinase was identified as a type I PI 3'-kinase on the basis of its inhibition by Nonidet P-40 and high pressure liquid chromatography of the phosphorylated product. To confirm the identification of PI 3'-kinase as an effector of Neu, we raised antibodies to the alpha-isoform of the regulatory subunit of PI 3'-kinase (p85). Using these antibodies, it was possible to directly demonstrate ligand-dependent formation of a tyrosine-phosphorylated complex of NEC and PI 3'-kinase. Apparently, both PI 3'-kinase and phospholipase C gamma, another substrate of the Neu kinase, simultaneously associated with the same activated NEC molecule. Nevertheless, immunofluorescence localization of PI 3'-kinase revealed no significant cellular redistribution of the enzyme after activation of the Neu kinase. Interestingly, PI 3'-kinase was localized primarily to the cell nucleus and to confined regions of the plasma membrane. Analysis of mutants of the Neu protein indicated that the oncogenic point-mutated Neu (Glu664) was permanently coupled to PI 3'-kinase; but two nontransforming versions of the oncoprotein, a kinase-defective protein and a carboxyl-terminally deleted Neu, were devoid of the constitutive association with PI 3'-kinase. Hence, we concluded that phosphatidylinositol 3'-kinase is a physiological substrate of the Neu receptor, but the regulation of this coupling is released upon oncogenic activation.
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PMID:Regulated coupling of the Neu receptor to phosphatidylinositol 3'-kinase and its release by oncogenic activation. 135 Oct 56

Autophosphorylation of gp185erbB-2 in vivo is confined to its carboxy terminus and is required for optimal erbB-2 transforming activity under conditions of receptor overexpression. It remains unresolved, however, to what extent autophosphorylation regulates erbB-2 mitogenic signaling in normal cells, nor is the biochemical basis for such a regulatory function known. To address these issues, we utilized a chimeric molecule encompassing the extracellular domain of the epidermal growth factor (EGF) receptor (EGFR) fused to the transmembrane and intracellular domains of the erbB-2 product. In this EGFR/erbB-2 chimera, erbB-2 kinase activity is regulated by EGF binding. An EGFR/erbB-2 mutant bearing multiple Tyr----Phe substitutions at erbB-2 autophosphorylation sites (EGFR/erbB-2 5P) displayed markedly reduced phosphotyrosine content following EGF stimulation in comparison with the non-mutated chimera. When expressed in NR6 cells, the EGFR/erbB-2 5P mutant was unable to deliver a sizeable mitogenic signal when activated by EGF at physiological levels. In intact cells, the 5P mutant was still able to stimulate phosphorylation of the gamma isozyme of phospholipase C (PLC-gamma), a prototype erbB-2 substrate, although with a delayed time course, indicating that the 5P mutation decreased the affinity of the erbB-2 kinase for this substrate. This conclusion was further supported by the inability of the 5P mutant to associate with PLC-gamma in co-immunoprecipitation experiments. We infer that a major role of autophosphorylation is to increase the affinity of the erbB-2 kinase for its cellular substrates, so that, under physiological conditions, autophosphorylation is absolutely required for erbB-2 mitogenic signaling.
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PMID:erbB-2 autophosphorylation is required for mitogenic action and high-affinity substrate coupling. 135 97

The chicken CT10 virus oncogene product, P47gag-crk, contains SH2/SH3 domains that have been identified as conserved domains among proteins involved in signal transduction. We studied the functional similarity of the SH2/SH3 domains by replacing those of v-Crk with those of phosphatidylinositol-specific phospholipase C-gamma, v-Src, or c-Src. The transforming activity of v-Crk was partially retained in a mutant with a v-Src SH3 domain but not in the other mutants with heterologous SH2/SH3 domains. Mutant viruses with Crk-SH2/SH2' domains induced tyrosine phosphorylation of cellular proteins, but mutants with phosphatidylinositol-specific phospholipase C-gamma or Src SH2/SH2' domains did not. However, the mutant proteins with heterologous SH2/SH2' regions were able to weakly associate with some phosphotyrosine-containing proteins in vitro. These results indicate that in the context of the P47gag-crk structure, the requirement of Crk-SH2/SH3 is more stringent for its activity to induce cell transformation than to cause phosphorylation of cellular proteins. The substitution with heterologous sequences least perturbs the capacity to bind phosphotyrosine-containing proteins. In each case, the SH3 domain is more flexible to substitution than is the SH2 domain.
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PMID:Biological and biochemical activity of v-Crk chimeras containing the SH2/SH3 regions of phosphatidylinositol-specific phospholipase C-gamma and Src. 137 84

The mechanisms by which phorbol 12-myristate 13-acetate (PMA) and cAMP attenuate the hydrolysis of phosphatidylinositol 4,5-bisphosphate (PtdIns 4,5-P2) induced by ligation of the T-cell antigen receptor complex (TCR) was studied in the human Jurkat T-cell line. It has previously been shown that stimulation of Jurkat cells with antibodies to CD3, components of the TCR, elicits a rapid and transient phosphorylation of phospholipase C (PLC)-gamma 1, the predominant PLC isozyme in Jurkat cells, at multiple tyrosine residues and that such tyrosine phosphorylation leads to activation of PLC-gamma 1. Prior incubation of Jurkat cells with PMA or forskolin, which increases intracellular cAMP concentrations, prevented tyrosine phosphorylation of PLC-gamma 1 as well as the hydrolysis of PtdIns 4,5-P2 induced by ligation of CD3. Dose-response curves of PMA and of forskolin for the inhibition of PLC-gamma 1 tyrosine phosphorylation and of PtdIns 4,5-P2 hydrolysis were similar. These results suggest that the inhibition of PtdIns 4,5-P2 hydrolysis by PMA and cAMP is attributable to reduced tyrosine phosphorylation of PLC-gamma 1. Treatment of Jurkat cells with PMA or forskolin stimulated the phosphorylation of PLC-gamma 1 at serine 1248. PMA treatment also elicited the phosphorylation of PLC-gamma 1 at an unidentified serine site. Phosphopeptide map analysis indicated that the sites of PLC-gamma 1 phosphorylated in Jurkat cells treated with PMA and forskolin are the same as those phosphorylated in vitro by protein kinase C (PKC) and cAMP-dependent protein kinase (PKA), respectively. Stimulation of Jurkat cells with antibodies to CD3 also elicited phosphorylation of PLC-gamma 1 at serine 1248 and at the unidentified serine site phosphorylated in PLC-gamma 1 from PMA-treated cells. Thus, phosphorylation of PLC-gamma 1 by PKC or PKA at serine 1248 may modulate the interaction of PLC-gamma 1 with the protein tyrosine kinase or the protein tyrosine phosphatase; this altered interaction may, at least in part, be responsible for the decreased tyrosine phosphorylation of PLC-gamma 1 seen in PMA- and forskolin-treated Jurkat cells. Furthermore, in the absence of PMA, activation of PKC by diacylglycerol provides a negative feedback signal responsible for reducing the phosphotyrosine contents of PLC-gamma 1.
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PMID:Inhibition of CD3-linked phospholipase C by phorbol ester and by cAMP is associated with decreased phosphotyrosine and increased phosphoserine contents of PLC-gamma 1. 137 Apr 76

The tyrosine kinase inhibitors ST271, ST638 and erbstatin inhibited phospholipase D (PLD) activity in human neutrophils stimulated by fMet-Leu-Phe, platelet-activating factor and leukotriene B4. These compounds did not inhibit phorbol ester-stimulated PLD, indicating that they do not inhibit PLD per se, but probably act at a site between the receptor and the phospholipase. In contrast, the protein kinase C inhibitor Ro-31-8220 inhibited phorbol 12,13-dibutyrate- but not fMet-Leu-Phe-stimulated PLD activity, arguing against the involvement of protein kinase C in the receptor-mediated activation of PLD. ST271 did not inhibit Ins(1,4,5)P3 generation, but did inhibit protein tyrosine phosphorylation stimulated by fMet-Leu-Phe. The phosphotyrosine phosphatase inhibitor pervanadate increased tyrosine phosphorylation and stimulated PLD. These results suggest that tyrosine kinase activity is involved in receptor coupling to PLD but not to PtdIns(4,5)P2-specific phospholipase C in the human neutrophil.
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PMID:Tyrosine phosphorylation is involved in receptor coupling to phospholipase D but not phospholipase C in the human neutrophil. 137 83

Stimulation of thymocytes or mature T cells via the T cell receptor (TcR)/CD3 complex activates a cascade of processes inducing cells to enter the cell cycle. A key step is the activation of phosphatidylinositol-specific phospholipase C (PI-PLC) within seconds following TcR/CD3 stimulation, an event which is strongly enhanced by co-ligation of the CD4 (or CD8) accessory molecule with TcR/CD3. In contrast, co-ligation of CD45 inhibits the same TcR/CD3 responses. The machinery which couples the TcR/CD3 complex, CD4, and CD45 to PI-PLC appears to involve regulation of tyrosine phosphorylation, as the TcR/CD3 and CD4 receptors are associated with the tyrosine kinases p59fyn and p56lck, respectively, and CD45 has intrinsic tyrosine phosphatase activity. Here, we have examined the ability of CD45 to regulate signal transduction via TcR/CD3 in human thymocytes. Co-cross-linking CD45 to the TcR/CD3 complex strongly suppressed the tyrosine phosphorylation of several intracellular substrates normally seen following TcR/CD3 stimulation. This effect of CD45 was associated with inhibition of a rise in intracellular calcium following TcR/CD3 ligation. Since TcR/CD3 stimulation of mature T cells induces tyrosine phosphorylation of PLC gamma 1, we investigated this phenomenon in thymocytes, and asked whether ligation of CD45 might regulate this process. By immunoprecipitation we found that TcR/CD3 stimulation induced tyrosine phosphorylation of PLC gamma 1, an effect which was enhanced by co-cross-linking CD4 to TcR/CD3. In contrast, co-ligation of CD45 strongly blocked PLC gamma 1 phosphorylation induced by either stimulus. Consistent with previous findings in mature T cells, CD45 cross-linking was able to partially inhibit TcR/CD3-induced thymocyte proliferation when interleukin 2 was used as a second signal, but almost completely (80%-90%) blocked proliferation when anti-CD28 mAb was used as the second signal, suggesting that CD45 cross-linking may be able to block interleukin 2 production via the CD28 pathway. These effects of CD45 on TcR/CD3 signaling and proliferation in thymocytes point towards a potential role for this pathway in thymic selection.
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PMID:CD45 modulates T cell receptor/CD3-induced activation of human thymocytes via regulation of tyrosine phosphorylation. 137 71


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