Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.1.137 (phosphatidylinositol 3-kinase)
 11,064 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Substitution of phenylalanine for tyrosine-809 in the human colony-stimulating factor 1 receptor (CSF-1R) inhibited its ability to transduce ligand-dependent mitogenic signals in mouse NIH 3T3 cells. When combined with an "activating" mutation at codon 301 that induces constitutive CSF-1R tyrosine kinase activity, the codon 809 mutation suppressed ligand-independent cell transformation. Comparative mapping of tryptic phosphopeptides from mutant and wild-type CSF-1R indicated that tyrosine-809 is a site of ligand-dependent receptor phosphorylation in vivo. The mutant receptor was active as a tyrosine kinase in vitro and in vivo, underwent CSF-1-dependent association with a phosphatidylinositol 3-kinase, and induced expression of the protooncogenes c-fos and junB, underscoring its ability to trigger some of the known cellular responses to CSF-1. The mutant receptor is likely to be impaired in its ability to interact with critical cellular effectors whose activity is required for mitogenesis.
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PMID:A point mutation at tyrosine-809 in the human colony-stimulating factor 1 receptor impairs mitogenesis without abrogating tyrosine kinase activity, association with phosphatidylinositol 3-kinase, or induction of c-fos and junB genes. 216 57

Hepatocyte growth factor receptor is identified as a heterodimeric tyrosine kinase encoded by the c-met gene. This study was designed to determine how the c-met/hepatocyte growth factor receptor participates in the intracellular events involved in rat liver regeneration induced by administration of carbon tetrachloride. Expression of the rat c-met mRNA increased, peaking 24 h after carbon tetrachloride administration almost in parallel with MET protein expression. Histochemical studies demonstrated that expression of the rat c-met was enhanced in cells surrounding the damaged areas, and also that the distribution of cells expressing MET was almost in accordance with that of cells expressing proliferating cells nuclear antigen. The MET protein underwent intense tyrosine phosphorylation peaking at 12 h after carbon tetrachloride administration, and prior to DNA synthesis. Phospholipase C gamma and phosphatidylinositol 3-kinase, intracellular signal transducing molecules containing Src homology 2 domain, were associated with the MET protein following tyrosine phosphorylation in vivo. These observations suggest that expression and tyrosine phosphorylation of MET protein associated with signal transducing molecules may provide a mechanism whereby hepatocyte growth factor exerts its action on hepatocyte growth during rat liver regeneration induced by carbon tetrachloride administration.
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PMID:Expression and phosphorylation of rat c-met/hepatocyte growth factor receptor during rat liver regeneration. 749 89

The surface engagement of high affinity immunoglobulin E receptor (Fc epsilon RI) of rat basophilic leukemia 2H3 (RBL-2H3) cells induced histamine secretion and leukotriene release following activation of the tyrosine kinase Lyn together with phosphatidylinositol 3-kinase (PI3-kinase). Wortmannin inhibited the activity of partially purified PI3-kinase from calf thymus, as well as the PI3-kinase activity in anti-PI3-kinase p85 immunoprecipitates from RBL-2H3 cells, at a concentration as low as 1.0 nM and with IC50 values of 3.0 nM, but did not inhibit PI4-kinase activity. The inhibition of PI3-kinase by wortmannin was irreversible. Wortmannin inhibited both Fc epsilon RI-mediated histamine secretion and leukotriene release up to 80% with IC50 values of 2.0 and 3.0 nM, respectively. Wortmannin inhibited PI3-kinase activity in intact cells up to 80% with an IC50 value of 2.0 nM, which is almost equal to those for PI3-kinase in vitro and for histamine secretion and leukotriene release. With anti-wortmannin antibody, we have shown that wortmannin binds to the 110-kDa protein, but not to PI3-kinase 85-kDa regulatory subunit both in vitro and in whole cells. Furthermore, there was a positive correlation between the potencies of wortmannin derivatives as inhibitors of PI3-kinase and as inhibitors of histamine secretion. Wortmannin had no effect on the activation of the tyrosine kinase Lyn. These results suggest that PI3-kinase is involved in the signal transduction pathway responsible for histamine secretion following stimulation of Fc epsilon RI and that wortmannin blocks these responses through direct interaction with the catalytic subunit of this enzyme.
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PMID:Inhibition of histamine secretion by wortmannin through the blockade of phosphatidylinositol 3-kinase in RBL-2H3 cells. 750 89

We studied a patient with severe insulin resistance and a remarkable decrease in the in vivo autophosphorylation of the insulin receptor. Using a polymerase chain reaction-single strand conformation polymorphism method and direct sequencing, we identified a heterozygous mutation substituting Gln for Arg1131 in the putative "catalytic loop" of the tyrosine kinase domain of the insulin receptor gene. The Gln1131 mutant receptor was expressed by transfection in Chinese hamster ovary cells and compared with cells expressing the wild-type insulin receptor. Both mutant and wild-type receptors were expressed on the cell surface and displayed similar insulin-binding affinity. The Gln1131 mutation impaired the activity of the receptor tyrosine kinase and inhibited the ability of insulin to phosphorylate the endogenous substrate insulin receptor substrate-I. In addition, the Gln1131 mutant receptor exhibited diminished tyrosine-phosphorylated phosphatidylinositol 3-kinase and myelin basic protein kinase activities compared with the wild-type cells. It also demonstrated a defective mediation of the insulin signal stimulating 2-deoxy-D-glucose transport and thymidine incorporation, resistance to endocytosis, and insulin-induced down-regulation. Unlike a previously described mutation in the putative catalytic loop of the receptor that substituted Glu for Ala1135, the Gln1131 mutation retained proteolytic cleavage of the proreceptor into separate subunits. Our results demonstrate that a naturally occurring mutation (R1131Q) in the putative catalytic loop of the insulin receptor results in severe impairment of the tyrosine kinase function in our patient. In addition, our results indicate that Arg1131 is important for receptor-mediated insulin action in vivo and suggest that the amino acids constituting the catalytic loop of protein kinases may possess different modes in order to retain kinase function.
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PMID:Substitution of glutamine for arginine 1131. A newly identified mutation in the catalytic loop of the tyrosine kinase domain of the human insulin receptor. 751 63

Signaling by tyrosine kinase receptors is mediated by selective interactions between individual Src homology 2 (SH2) domains of cytoplasmic effectors and specific phosphotyrosine residues in the activated receptor. Here, we report the existence in the hepatocyte growth factor/scatter factor (HGF/SF) receptor of a multifunctional docking site made of the tandemly arranged degenerate sequence YVH/NV. Phosphorylation of this site mediates intermediate- to high-affinity interactions with multiple SH2-containing signal transducers, including phosphatidylinositol 3-kinase, phospholipase C gamma, pp60c-src, and the GRB-2-Sos complex. Mutation of the two tyrosines results in loss of biological function, as shown by abrogation of the transforming activity in the oncogenic counterpart of the receptor. The same bidentate motif is conserved in the evolutionarily related receptors Sea and Ron, suggesting that in all members of the HGF/SF receptor family, signal transduction is channeled through a multifunctional binding site.
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PMID:A multifunctional docking site mediates signaling and transformation by the hepatocyte growth factor/scatter factor receptor family. 751 58

To identify novel proteins capable of associating with the Raf-1 serine/threonine kinase, we investigated whether Raf-1 could interact with the Src homology 2 (SH2) domains of various signal-transducing molecules. In this report, we demonstrate that Raf-1 associated with the SH2 domain of Fyn (a member of the Src tyrosine kinase family) but not with the SH2 domains of phospholipase C-gamma 1, the p85 alpha subunit of phosphatidylinositol 3-kinase, and SH2-containing protein tyrosine phosphatase 2. Unlike most SH2 domain interactions that require tyrosine-phosphorylated residues, the Raf-1/Fyn SH2 domain association was dependent on the serine phosphorylation of Raf-1. Our results also demonstrate that Raf-1 interacted with the SH2 domain of Src and that this interaction was destabilized by mutation of Arg175 found within the conserved SH2 domain FLVRES sequence. In addition, we show that inclusion of additional Src sequences containing the SH3 domain increased the association of Raf-1 with the Src SH2 domain. Finally, using the baculovirus/Sf9 cell system, we show that coexpression of Raf-1 with full-length Fyn/Src resulted in the coimmunoprecipitation of Raf-1 with Fyn/Src, the tyrosine phosphorylation of Raf-1, and the stimulation of Raf-1 kinase activity. These results suggest that Raf-1 may form a functional complex with Fyn/Src mediated in part by SH2 domains and the serine phosphorylation of Raf-1.
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PMID:Raf-1 interacts with Fyn and Src in a non-phosphotyrosine-dependent manner. 751 1

Insulin rapidly stimulates tyrosine kinase activity of its receptor resulting in phosphorylation of its cytosolic substrate insulin receptor substrate 1 (IRS-1), which in turn associates with and activates the enzyme phosphatidylinositol 3-kinase (PI 3-kinase). In the present study we have examined these three initial steps in insulin action during the differentiation of 3T3-F442A adipocytes and after treatment with dexamethasone or insulin. The differentiation of 3T3-F442A cells was characterized by a 13-fold increase in insulin receptor protein, a 9-fold increase in IRS-1, and a 10- and 4.5-fold increase in their insulin-stimulated phosphorylation, respectively. The mRNA expression of these two proteins showed a similar 8-fold increase during differentiation. In addition there was a 3.5-fold increase in PI 3-kinase protein [85 kilodalton (kDa) subunit] and a 16-fold increase in IRS-1-associated PI 3-kinase activity between day 0 and day 8 of differentiation. Dexamethasone (1 microM) treatment of differentiated cells induced a further 48% (P < 0.05) increase in insulin receptor level, but the autophosphorylation of the receptor was decreased by 31 +/- 1% (P < 0.02). At the same time there was a decrease by 56 +/- 4% (P < 0.005) in IRS-1 protein and by 31 +/- 1% (P < 0.001) in IRS-1 phosphorylation. The expression of insulin receptor mRNA was unchanged, but the expression of IRS-1 mRNA was decreased by approximately 75% after dexamethasone. By contrast, dexamethasone induced a 69% increase in the level of PI 3-kinase as determined by immunoblotting. The combined effect of decreased IRS-1 phosphorylation and increased PI 3-kinase protein was a minimal change (15% decrease) in the association/activation between IRS-1 and PI 3-kinase. Chronic treatment with 100 nM insulin induced a time- and dose-dependent decrease in insulin receptor and IRS-1 protein levels reaching a nadir of 34 +/- 5% (P < 0.005) and 39 +/- 5% (P < 0.01) of control levels after 24 h, respectively. There was an even more marked decrease in the phosphorylation level of these proteins. Chronic insulin treatment also produced a 30% decrease in PI 3-kinase protein levels and a approximately 50% decrease in the association/activation between IRS-1/PI 3-kinase. The expression of insulin receptor and IRS-1 mRNA was unchanged during chronic insulin treatment.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Regulation of insulin receptor, insulin receptor substrate-1 and phosphatidylinositol 3-kinase in 3T3-F442A adipocytes. Effects of differentiation, insulin, and dexamethasone. 752 Jan 27

The Src family protein tyrosine kinases participate in signalling through cell surface receptors that lack intrinsic tyrosine kinase domains. All nine members of this family possess adjacent Src homology (SH2 and SH3) domains, both of which are essential for repression of the enzymatic activity. The repression is mediated by binding between the SH2 domain and a C-terminal phosphotyrosine, and the SH3 domain is required for this interaction. However, the biochemical basis of functional SH2-SH3 interaction is unclear. Here, we demonstrate that when the SH2 and SH3 domains of p59fyn (Fyn) were present as adjacent domains in a single protein, binding of phosphotyrosyl peptides and proteins to the SH2 domain was enhanced, whereas binding of a subset of cellular polypeptide ligands to the SH3 domain was decreased. An interdomain communication was further revealed by occupancy with domain-specific peptide ligands: occupancy of the SH3 domain with a proline-rich peptide enhanced phosphotyrosine binding to the linked SH2 domain, and occupancy of the SH2 domain with phosphotyrosyl peptides enhanced binding of certain SH3-specific cellular polypeptides. Second, we demonstrate a direct binding between purified SH2 and SH3 domains of Fyn and Lck Src family kinases. Heterologous binding between SH2 and SH3 domains of closely related members of the Src family, namely, Fyn, Lck, and Src, was also observed. In contrast, Grb2, Crk, Abl, p85 phosphatidylinositol 3-kinase, and GTPase-activating protein SH2 domains showed lower or no binding to Fyn or Lck SH3 domains. SH2-SH3 binding did not require an intact phosphotyrosine binding pocket on the SH2 domain; however, perturbations of the SH2 domain induced by specific high-affinity phosphotyrosyl peptide binding abrogated binding of the SH3 domain. SH3-SH2 binding was observed in the presence of proline-rich peptides or when a point mutation (W119K) was introduced in the putative ligand-binding pouch of the Fyn SH3 domain, although these treatments completely abolished the binding to p85 phosphatidylinositol 3-kinase and other SH3-specific polypeptides. These biochemical SH2-SH3 interactions suggest novel mechanisms of regulating the enzymatic activity of Src kinases and their interactions with other proteins.
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PMID:Physical and functional interactions between SH2 and SH3 domains of the Src family protein tyrosine kinase p59fyn. 752 May 28

c-Mpl is a member of the cytokine receptor superfamily, expressed primarily on hematopoietic cells. Recently, the c-Mpl ligand was cloned and found to have thrombopoietic activity. In this paper we report that ligand binding induced tyrosine phosphorylation in BaF3 cells engineered to express the murine Mpl receptor (BaF3/mMpl). Phosphorylation occurred within 1 min at cytokine concentrations sufficient for proliferation of receptor-bearing cells. Using specific antibodies for immunoprecipitation and Western blotting, several of these phosphorylated proteins were identified. Shc and Jak2, known cytokine signaling molecules, and the c-Mpl receptor were shown to be major substrates for tyrosine phosphorylation. In contrast, phospholipase C-gamma and phosphatidylinositol 3-kinase displayed little and no tyrosine phosphorylation, respectively, after thrombopoietin stimulation. Co-immunoprecipitation studies demonstrated that Jak2 became physically associated with c-Mpl relatively late in the observed time course (20-60 min), significantly later than tyrosine phosphorylation of Jak2 (1-5 min). These results suggest that c-Mpl induces signal transduction pathways similar to those of other known cytokines. Additionally, in light of its late physical association with c-Mpl following ligand binding, Jak2 may not be the initiating tyrosine kinase in the thrombopoietin-induced signaling cascade.
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PMID:The c-Mpl ligand (thrombopoietin) stimulates tyrosine phosphorylation of Jak2, Shc, and c-Mpl. 753 85

Activation of the tyrosine kinase activity of the insulin receptor by autophosphorylation leads to phosphorylation of cellular substrates on tyrosine. Thus far, the best characterized is the insulin receptor substrate (IRS) 1, which has been proposed to serve as a docking protein for other molecules involved in signal transduction. A number of other proteins that become phosphorylated in response to insulin have been identified, some of which are reported to be tissue-specific. A 60 kDa phosphoprotein has been detected in adipocytes after insulin stimulation [Lavan and Lienhard (1993) J. Biol. Chem. 268, 5921-5928]. We have identified a protein of similar molecular mass in rat hepatoma cells transfected with the human insulin receptor. The 60 kDa protein in hepatoma cells is tyrosine-phosphorylated in response to insulin in a dose-dependent manner, with maximal phosphorylation occurring at 50 nM insulin. Although the dose-response of p60 phosphorylation mirrors that of IRS-1, the time course is slightly slower, with maximal phosphorylation observed 5 min after addition of insulin. Like the adipocyte protein, the 60 kDa protein detected in liver cells binds to the SH2 domain of the p85 regulatory subunit of phosphatidylinositol 3-kinase, but not to other SH2 domains. Binding of p60 to p85 is similar to the interaction between p85 and IRS-1 in that a tyrosine-phosphorylated peptide containing the YVXM motif can inhibit the association. The presence of this 60 kDa tyrosine-phosphorylated protein in adipocytes and hepatoma cells suggests that it represents another important intermediate in the insulin-receptor signal-transduction pathway.
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PMID:Detection of a 60 kDa tyrosine-phosphorylated protein in insulin-stimulated hepatoma cells that associates with the SH2 domain of phosphatidylinositol 3-kinase. 753 11


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