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

---

Query: EC:3.1.4.3 (phospholipase C)
18,461 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Activation of phospholipase C (PLC), leading to a rise in cytosolic Ca2+, and of phospholipase A2 (PLA2) leading to a release of arachidonic acid, are among the early transmembrane signalling events that have been demonstrated in response to occupancy of the epidermal growth factor (EGF) receptor. The tyrosine kinase activity of the receptor has been shown to be necessary for both of these responses. This requirement for the tyrosine kinase activity could conceivably implicate a role for receptor autophosphorylation in the activation of PLA2. We now demonstrate that coupling of the EGF receptor to PLA2 was not impaired in a deletion mutant (CD126) devoid of the 126 amino acids from the C-terminus which include four major autophosphorylation sites. Functional coupling of the EGF receptor to PLA2 was demonstrated using three different experimental designs: (1) release of [14C]arachidonic acid from prelabelled intact cells. (2) release of [3H]arachidonic acid from prelabelled cells permeabilized with glass beads, and (3) direct measurement of PLA2 enzymic activity in cell-free extracts using an 'in vitro' assay employing exogenous phospholipid substrate. Functional coupling of the EGF receptor to PLA2 occurred despite the absence of a demonstrable Ca(2+)-signalling response and the detection of diminished but persistent PLC-gamma phosphorylation on tyrosine residues in the CD126 deletion mutants. These results point to a clear distinction in the biochemical mechanism and role for receptor autophosphorylation in functional coupling of the EGF receptor to PLA2 activation versus Ca2+ signalling.
...
PMID:Distinct structural specificities for functional coupling of the epidermal growth factor receptor to calcium-signalling versus phospholipase A2 responses. 190 21

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.
...
PMID:Selectivity of phospholipase C phosphorylation by the epidermal growth factor receptor, the insulin receptor, and their cytoplasmic domains. 215 2

The erbB-2 gene product, gp185erbB-2, displays a potent transforming effect when overexpressed in NIH 3T3 cells. In addition, it possesses constitutively high levels of tyrosine kinase activity in the absence of exogenously added ligand. In this study, we demonstrate that its carboxy-terminal domain exerts an enhancing effect on erbB-2 kinase and transforming activities. A premature termination mutant of the erbB-2 protein, lacking the entire carboxy-terminal domain (erbB-2 delta 1050), showed a 40-fold reduction in transforming ability and a lowered in vivo kinase activity for intracellular substrates. When the carboxy-terminal domain of erbB-2 was substituted for its analogous region in the epidermal growth factor receptor (EGFR) (EGFR/erbB-2COOH chimera), it conferred erbB-2-like properties to the EGFR, including transforming ability in the absence of epidermal growth factor, elevated constitutive autokinase activity in vivo and in vitro, and constitutive ability to phosphorylate phospholipase C-gamma. Conversely, a chimeric erbB-2 molecule bearing an EGFR carboxy-terminal domain (erbB-2/EGFRCOOH chimera) showed reduced transforming and kinase activity with respect to the wild-type erbB-2 and was only slightly more efficient than the erbB-2 delta 1050 mutant. Thus, we conclude that the carboxy-terminal domains of erbB-2 and EGFR exert different regulatory effects on receptor kinase function and biological activity. The up regulation of gp185erbB-2 enzymatic activity exerted by its carboxy-terminal domain can explain, at least in part, its constitutive level of kinase activity.
...
PMID:The carboxy-terminal domains of erbB-2 and epidermal growth factor receptor exert different regulatory effects on intrinsic receptor tyrosine kinase function and transforming activity. 218 97

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.
...
PMID:Src homology region 2 domains direct protein-protein interactions in signal transduction. 223 73

Phosphorylation of two newly identified epidermal growth factor (EGF) receptor substrates, eps8 and eps15, which do not possess Src homology (SH2) domains, was investigated using EGF receptor mutants of the autophosphorylation sites and deletion mutants of the carboxyl-terminal region. Two mutants, F5, in which all five tyrosine autophosphorylation sites substituted by phenylalanine, and Dc 123F, in which four tyrosines were removed by deletion and the fifth (Tyr-992) was mutated into phenylalanine, phosphorylated eps8 and eps15 as efficiently as the wild-type receptor. In contrast, SH2-containing substrates, phospholipase C gamma, the GTPase-activating protein of Ras, the p85 subunit of phosphatidylinositol 3 kinase, and the Src and collagen homology protein, are not phosphorylated by the F5 and Dc 123F mutants. A longer EGF receptor deletion mutant, Dc 214, lacking all five autophosphorylation sites, was unable to phosphorylate eps15 but phosphorylated eps8 13-fold more than the wild-type receptor. To determine the EGF receptor region important for phosphorylation of eps8 and eps15, progressive deletion mutants lacking the final 123, 165, 196, and 214 COOH-terminal residues were used. eps8 phosphorylation was progressively increased in Dc 165, Dc 196, and Dc 214 EGF receptor mutants, indicating that removal of the final 214 COOH-terminal residues increases the phosphorylation of this substrate by the EGF receptor. In contrast, eps15 was phosphorylated by Dc 123 and Dc 165 EGF receptor mutants but not by Dc 196 and Dc 214 mutants. This indicates that a region of 30 residues located between Dc 165 and Dc 196 is essential for eps15 phosphorylation. This is the first demonstration of structural requirements in the EGF receptor COOH terminus for efficient phosphorylation of non-SH2-containing substrates. In addition, enhanced eps8 phosphorylation correlates well with the increased transforming potential of EGF receptor deletion mutants Dc 196 and Dc 214, suggesting that this substrate may be involved in mitogenic signaling.
...
PMID:Structural requirements of the epidermal growth factor receptor for tyrosine phosphorylation of eps8 and eps15, substrates lacking Src SH2 homology domains. 760 94

A single point mutation, Glu627--> Val, equivalent to the activating mutation in the Neu oncogene, was inserted in the transmembrane domain of the human epidermal growth factor (EGF) receptor. Unlike the wild type, Glu627-EGF receptor, transfected in NIH3T3 cells, gave rise to focal transformation and growth in agar even in the absence EGF. Constitutive activity of mutant EGF receptor amounted to 20% of that of wild type receptor stimulated by EGF. In addition, the mutant receptor was more sensitive to EGF, reaching maximum transforming activity at 5 ng/ml EGF. NIH3T3 cells expressing Glu627-EGF receptor showed a transformed phenotype and were not arrested in G0 upon serum deprivation. The mutant receptor was constitutively autophosphorylated, and several other cellular proteins were phosphorylated on tyrosine in absence of the ligand. Among these, the SHC adaptor protein was phosphorylated in absence of EGF, the other adaptor, GRB-2 was constitutively associated with the Glu627-EGF receptor in vivo and in vitro, and mitogen-activated protein kinase was constitutively phosphorylated. In contrast, other EGF receptor substrates, like phospholipase C gamma, were not phosphorylated in absence of EGF. The mutant receptor showed a higher sensitivity to cleavage by calpain both in absence and presence of EGF, appeared as a 170- and 150-kDa doublet in cell extracts, and a specific calpain inhibitor blocked the appearance of the 150-kDa form. Since the calpain cleavage site is located in the receptor cytoplasmic tail, this finding suggests that the Glu627 mutation induces a slightly different conformation in the EGF receptor intracellular domain. In conclusion, our data show that a point mutation in the EGF receptor transmembrane domain was able to constitutively activate the receptor and to induce transformation via constitutive activation of the Ras pathway.
...
PMID:SHC and GRB-2 are constitutively by an epidermal growth factor receptor with a point mutation in the transmembrane domain. 764 41

Src homology regions 2 (SH2) and 3 (SH3) are noncatalytic domains that are conserved among several proteins implicated in the regulation of cell proliferation. Using bacterially expressed fusion proteins containing the SH2 domain of the abl tyrosine kinase, we have quantitated the binding of these domains to the activated epidermal growth factor (EGF) receptor (EGFR). A 35S-labeled abl SH2 fusion protein binds to the human EGFR immunoprecipitated from EGF-treated NIH3T3 cells that overexpress the receptor. This binding is totally dependent on the pretreatment of cells with EGF. The interaction is rapid, reaching 50% of maximum within 1 min, and attaining apparent equilibrium by 10 min. Dissociation of the complex is biphasic with a rapidly dissociating component (t1/2 of less than 1 min), as well as a slowly dissociable component. The 35S-labeled abl SH2 fusion protein specifically binds to the EGFR in a saturable manner and is differentially inhibited by unlabeled fusion proteins containing SH2 domains from phospholipase C, the p85 subunit of phosphatidylinositol-3 kinase, and the GTPase activation protein of ras. To identify residues critical for abl SH2-EGFR binding, six point mutants were constructed in the highly conserved FLVRES motif. Three mutants (V170L, E172Q, and E174Q) display binding affinities similar to that of wild type. However, three other mutants (R171K, S173C, and S175C) have greatly reduced affinity. Interestingly, the binding affinity to the EGFR determined by the in vitro assay directly correlates with the transforming ability of the corresponding v-abl constructs in vivo (Mayer, B. J., Jackson, P. K., Etten, R. A. V., and Baltimore, D. (1992) Mol. Cell. Biol. 12, 609-618). These data indicate that the Arg-171, Ser-173, and Ser-175 are critical for both transformation and abl SH2 domain binding to phosphotyrosine-containing proteins.
...
PMID:Direct analysis of the binding of the abl Src homology 2 domain to the activated epidermal growth factor receptor. 767 9

Human platelets contain phospholipase C (PLC)-gamma 2, a distinct isoform closely related to PLC-gamma 1. Both inositol phospholipid-specific phospholipases C contain the src-related SH2 regions. Stimulation of platelets with the potent agonist, thrombin, led to a rapid and transient phosphorylation of PLC-gamma 2 on tyrosine residues. Activated platelets lysed in the absence of sodium orthovanadate had levels of tyrosine-phosphorylated PLC-gamma 2 paralleling those seen in unstimulated platelets. Previously, it had been shown that PLC-gamma 1 was phosphorylated on tyrosine residues by the agonist-occupied platelet-derived growth factor (PDGF) receptor and epidermal growth factor (EGF) receptor in cells other than platelets. In addition, more recent data have indicated that PLC-gamma 2 is also capable of being tyrosine-phosphorylated in cells of hematopoietic origin, such as B cells and natural killer (NK) cells. Here we report that PLC-gamma 2 expressed in a terminally-differentiated hematopoietic cell is also tyrosine-phosphorylated in response to an agonist.
...
PMID:Thrombin activation of human platelets causes tyrosine phosphorylation of PLC-gamma 2. 768 59

We have investigated coupling between the epidermal growth factor (EGF) receptor and the phospholipase C (PLC)/protein kinase C (PKC) signal-transduction system in normal skin fibroblasts and keratinocytes, for which EGF and transforming growth factor alpha (TGF-alpha) are mitogenic. EGF and TGF-alpha induced a rapid increase in tyrosine phosphorylation of the EGF receptor, in both fibroblasts and keratinocytes, but failed to induce tyrosine phosphorylation of PLC-gamma 1 or detectable phosphoinositide hydrolysis, as measured by two sensitive assays. In fibroblasts, EGF induced phosphatidylcholine (PC) hydrolysis, resulting in increased diacylglycerol (DAG). In contrast, in keratinocytes, there was no detectable PC hydrolysis or elevation of DAG in response to EGF or TGF-alpha. EGF and TGF-alpha activated PKC in fibroblasts, as evidenced by increased phosphorylation of a specific cellular PKC substrate (myristoylated alanine-rich C-kinase substrate, 'MARCKS'). In keratinocytes, TGF-alpha and EGF induced only a modest increase in MARCKS protein phosphorylation. This apparent modest activation of PKC, in the absence of detectable DAG formation, may have been mediated by arachidonic acid, which was released from keratinocytes in response to TGF-alpha, and has been shown to stimulate PKC activity in vitro. These data demonstrate that (1) in dermal fibroblasts and keratinocytes, which express normal levels of EGF receptors, EGF receptor activation is not coupled to tyrosine phosphorylation of PLC-gamma 1 or PtdIns hydrolysis, suggesting that these events are not required for the mitogenic activity of EGF or TGF-alpha in these cells, (2) coupling of EGF receptor to PC hydrolysis is cell-type specific, and (3) in skin fibroblasts, DAG, formed through EGF-induced PC hydrolysis, is capable of activating PKC.
...
PMID:Differential induction of phosphatidylcholine hydrolysis, diacylglycerol formation and protein kinase C activation by epidermal growth factor and transforming growth factor-alpha in normal human skin fibroblasts and keratinocytes. 769 May 46

NIH 3T3 cells expressing high levels of the human epidermal growth factor (EGF) receptor were used to examine the effects of the lectin concanavalin A (Con A) on EGF-mediated signaling events. Proliferation of NIH 3T3 cells expressing high levels of the human EGF receptor was inhibited in a dose-dependent manner by Con A. At the same time, Con A also inhibited both dimerization and tyrosine phosphorylation of the EGF receptor. Tyrosine phosphorylation of the enzyme phospholipase C-gamma, a substrate of the phosphorylated EGF receptor kinase, was also inhibited. In contrast, EGF-stimulated changes in pH, calcium, and levels of inositol phosphates were unaffected by the presence of Con A. These results indicate that certain signals (changes in the levels of intracellular calcium, pH, and inositol phosphates) mediated by EGF binding to its receptor still occur when receptor dimerization and phosphorylation are dramatically decreased, suggesting that multiple independent signals are transmitted by the binding of EGF to its receptor.
...
PMID:EGF receptor-mediated signals are differentially modulated by concanavalin A. 781 52


<< Previous 1 2 3 4 5 Next >>

---