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:2.7.10.2 (focal adhesion kinase)
44,029 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Previously, we characterized the prostaglandin (PG) F2 alpha receptor linked to phospholipase C activation and DNA synthesis in NIH-3T3 cells (Nakao, A., Watanabe, T., Taniguchi, S., Nakamura, M., Honda, Z-I., Shimizu, T., and Kurokawa, K. (1993) J. Cell. Physiol. 155, 257-264). To elucidate intracellular events evoked via this receptor, we examined changes caused by PGF2 alpha stimulation in the phosphotyrosine composition of cellular proteins. The addition of PGF2 alpha to cells in quiescent culture rapidly increased the levels of phosphotyrosine in cellular proteins with Mr values of 70,000 (pp70), 85,000 (pp85), 92,000 (pp92), 100,000 (pp100), and 125,000 (pp125); the latter was immunologically identified as p125 focal adhesion kinase. The PGF2 alpha-induced changes in the level of intracellular Ca2+ ([Ca2+]i) elevation, formation of inositol phosphates, and [3H]thymidine incorporation followed a similar dose dependence as PGF2 alpha-induced tyrosine phosphorylation. This tyrosine phosphorylation was independent of extracellular Ca2+, while a [Ca2+]i chelator, 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (50 microM), completely inhibited the PGF2 alpha-induced elevation of [Ca2+]i, tyrosine phosphorylation, and [3H]thymidine incorporation. Ionomycin (0.1 microM), which induced [Ca2+]i elevation without formation of inositol phosphates, mimicked the PGF2 alpha-induced tyrosine phosphorylation. 12-O-Tetradecanoylphorbol-13-acetate (TPA) also induced [3H]thymidine incorporation in a dose-dependent manner but had no significant effect on tyrosine phosphorylation. The PGF2 alpha-induced tyrosine phosphorylation could be observed even in the cells pretreated with TPA (5 microM, 24 h). PGF2 alpha exhibited an additive effect on TPA-induced [3H]thymidine incorporation but had no effect on the 32P-phosphorylation of a known 80-kDa protein kinase (PK) C substrate. Both staurosporine and H-7 inhibited the PGF2 alpha-induced increase in [3H]thymidine incorporation and tyrosine phosphorylation in a similar dose-dependent manner whether or not cells were pretreated with TPA (5 microM, 24 h). However, W-7 and KN-62 had no effect on these cellular responses even at the concentration for the almost complete inhibition of Ca2+/calmodulin-dependent PKs (20 microM). These results, taken together, indicate that PGF2 alpha receptor-mediated tyrosine phosphorylation is evoked by a [Ca2+]i-dependent mechanism that is sensitive to staurosporine and H-7 but which is independent of PKC or Ca2+/calmodulin PKs. Finally, the data suggest that this PGF2 alpha-induced signaling pathway is linked to the proliferation of cells.
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PMID:Prostaglandin F2 alpha enhances tyrosine phosphorylation and DNA synthesis through phospholipase C-coupled receptor via Ca(2+)-dependent intracellular pathway in NIH-3T3 cells. 802 Dec 71

The vav proto-oncogene product (Vav), which is specifically expressed in hematopoietic cells, contains multiple structural motifs commonly used by intracellular signaling molecules. Although a variety of stimuli including erythropoietin (Epo) have been shown to tyrosine phosphorylate Vav, little is known about the Vav signal transduction pathway. Here, we have investigated the role of Vav in the Epo signaling pathway by characterizing its interaction with other proteins, using the human Epo-responsive cell line, F-36P. Immunoprecipitation and immunoblot analyses have demonstrated that Vav was associated with the Epo receptor (EpoR) in an Epo-independent manner and was tyrosine-phosphorylated after Epo stimulation. Furthermore, two phosphotyrosine proteins (pp70 and pp100) co-immunoprecipitated with the regulatory subunit of phosphatidylinositol 3-kinase (PI3-kinase) (p85) were identified as EpoR and Vav, respectively. The interaction between Vav and p85 was shown to be mediated through the SH2 domains of p85 by an in vitro binding assay and confirmed by the presence of in vitro PI3-kinase activity associated with Vav. Treatment of the cells with antisense-vav and -p85 abrogated Epo-induced cell proliferation and PI3-kinase activity. Finally, we found that JAK2 was associated with Vav in vivo and that Vav could be tyrosine-phosphorylated by activated JAK2 in vitro. These results suggest the possible role of JAK2 for tyrosine phosphorylation of Vav and involvement of Vav and PI3-kinase in Epo-induced proliferative signals.
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PMID:Role of the vav proto-oncogene product (Vav) in erythropoietin-mediated cell proliferation and phosphatidylinositol 3-kinase activity. 916 69

Interleukin (IL)-2, a critical cytokine with indispensable functions in regulating lymphoid homeostasis, induces the activation of several biochemical pathways. Precisely how these pathways are linked and how they relate to the biological action of IL-2 is incompletely understood. We previously identified SHP-2 (Src homology 2 domain containing phosphatase 2) as an important intermediate in IL-2-dependent MAPK activation and showed its association with a 98-kDa phosphoprotein in response to IL-2. Here, we demonstrate that Gab2, a recently identified adapter molecule, is the major SHP-2 and phosphatidylinositol 3'-kinase-associated 98-kDa protein in normal, IL-2-activated lymphocytes. We further demonstrate that phosphorylation of both Gab2 and SHP-2 is largely dependent upon tyrosine 338 of the IL-2 receptor beta chain. Gab2 can be a substrate of all the three major classes of non-receptor tyrosine kinases associated with the IL-2R, but in terms of IL-2 signaling, JAK3 but not Lck or Syk is essential for Gab2 phosphorylation. We also demonstrate that only IL-2 and IL-15, but not other gammac cytokines induce Gab2 phosphorylation; the ability to phosphorylate Gab2 correlates with Shc phosphorylation and ERK1/ERK2 activation. Finally, we also show that Gab2 levels are regulated by T cell activation, and resting T cells express little Gab2. Therefore, up-regulation and activation of Gab2 may be important in linking the IL-2 receptor to activation of MAPK and may be an important means of achieving specificity in cytokine signaling.
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PMID:The docking molecule gab2 is induced by lymphocyte activation and is involved in signaling by interleukin-2 and interleukin-15 but not other common gamma chain-using cytokines. 1084 28

A subset of chromosomal translocations that participate in leukemia involve activated tyrosine kinases. The ets transcription factor, TEL, undergoes translocations with several distinct tyrosine kinases including JAK2. TEL-JAK2 transforms cell lines to factor independence, and constitutive tyrosine kinase activity results in the phosphorylation of several substrates including STAT1, STAT3, and STAT5. In this study we have shown that TEL-JAK2 can constitutively activate the phosphatidylinositol 3'-kinase (PI 3'-kinase) signaling pathway. The regulatory subunit of PI 3'-kinase, p85, associates with TEL-JAK2 in immunoprecipitations, and this was shown to be mediated by the amino-terminal SH2 domain of p85 but independent of a putative p85-binding motif within TEL-JAK2. The scaffolding protein Gab2 can also mediate the association of p85. TEL-JAK2 constitutively phosphorylates the downstream substrate protein kinase B/AKT. Importantly, the pharmacologic PI 3'-kinase inhibitor, LY294002, blocked TEL-JAK2 factor-independent growth and phosphorylation of protein kinase B. However, LY294002 did not alter STAT5 tyrosine phosphorylation, indicating that STAT5 and protein kinase B activation mediated by TEL-JAK2 are independent signaling pathways. Therefore, activation of the PI 3'-kinase signaling pathway is an important event mediated by TEL-JAK2 chromosomal translocations.
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PMID:TEL-JAK2 mediates constitutive activation of the phosphatidylinositol 3'-kinase/protein kinase B signaling pathway. 1143 25

Previous studies using cytochalasins and latrunculin B, inhibitors of actin polymerization, showed that filamentous (F)-actin had a negative regulatory role in Fc epsilon receptor I (Fc epsilon RI) signaling. How F-actin is involved in regulating the activation of mast cells is unknown. In this study we investigated the role of F-actin in mast cell activation induced by aggregation of the glycosylphosphatidylinositol (GPI)-anchored proteins Thy-1 and TEC-21, and compared it to activation via Fc epsilon RI. Pretreatment of rat basophilic leukemia cells with latrunculin B inhibited the Thy-1-induced actin polymerization and elevated the Thy-1-mediated secretory and calcium responses. Inhibition of actin polymerization followed by Thy-1 aggregation resulted in an increased tyrosine phosphorylation of Syk, phospholipase C gamma (PLC gamma), Gab2 and linker for activation of T cells (LAT) adapters, and some other signaling molecules. Enzymatic activities of phosphatidylinositol 3-kinase, PLC gamma, and phosphatase SHP-2 were also up-regulated, but tyrosine phosphorylation of ezrin was inhibited. Similar changes were observed in Fc epsilon RI-activated cells. Significant changes in intracellular distribution, tyrosine phosphorylation, and/or enzymatic activities of signaling molecules occurred in latrunculin-pretreated cells before cell triggering. The combined data suggest that actin polymerization is critical for setting the thresholds for mast cell signaling via aggregation of both Fc epsilon RI and GPI-anchored proteins.
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PMID:Involvement of filamentous actin in setting the threshold for degranulation in mast cells. 1516 32

The active forms of STAT5A (signal transducer and activator of transcription 5A) and STAT5B are able to relieve the cytokine dependence of haematopoietic cells and to induce leukaemia in mice. We have demonstrated previously that activation of the PI3K (phosphoinositide 3-kinase) signalling cascade plays a major role in cell growth and survival induced by these proteins. Interaction between STAT5 and p85, the regulatory subunit of the PI3K, has been suggested to be required for this activation. We show in the present study that the scaffolding protein Gab2 [Grb2 (growth-factor-receptor-bound protein 2)-associated binder-2] is an essential component of this interaction. Gab2 is persistently tyrosine-phosphorylated in Ba/F3 cells expressing caSTAT5 (constitutively activated STAT5), independent of JAK2 (Janus kinase 2) activation where it interacts with STAT5, p85 and Grb2, but not with Shp2 [SH2 (Src homology 2)-domain-containing tyrosine phosphatase] proteins. Interaction of STAT5 with Gab2 was also observed in Ba/F3 cells stimulated with interleukin-3 or expressing the oncogenic fusion protein Tel-JAK2. The MAPKs (mitogen-activated protein kinases) ERK1 (extracellular-signal-regulated kinase 1) and ERK2 were constitutively activated in the caSTAT5-expressing cells and were found to be required for caSTAT5-induced cell proliferation. Overexpression of Gab2-3YF, a mutant of Gab2 incapable of binding PI3K, inhibited the proliferation and survival of caSTAT5-expressing cells as well as ERK1/2 and Akt/protein kinase B phosphorylation. Taken together, our results indicate that Gab2 is required for caSTAT5-induced cell proliferation by regulating both the PI3K/Akt and the Ras/MAPK pathways.
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PMID:Activated STAT5 proteins induce activation of the PI 3-kinase/Akt and Ras/MAPK pathways via the Gab2 scaffolding adapter. 1583 84

We analyzed the effects of the Janus kinase 3 (Jak3)-specific inhibitor WHI-P131 (4-(4'-hydroxyphenyl)-amino-6,7-dimethoxyquinazoline) and the Jak3/Syk inhibitor WHI-P154 (4-(3'-bromo-4'-hydroxyphenyl)-amino-6,7-dimethoxyquinazoline) on the antigen-induced activation of mast cells. In the rat mast cell line RBL-2H3, both WHI-P131 and WHI-P154 inhibited the antigen-induced degranulation and phosphorylation of p44/42 mitogen-activated protein kinase (MAPK), p38 MAPK and c-Jun N-terminal kinase (JNK). The phosphorylation of Gab2, Akt and Vav was also inhibited by WHI-P131 and WHI-P154, indicating that these inhibitors suppress the activation of phosphatidylinositol 3-kinase (PI3K). In bone marrow-derived mast cells (BMMCs) from Jak3-deficient (Jak3-/-) mice, degranulation and activation of MAPKs were induced by the antigen in almost the same extent as in BMMCs from wild-type mice. In addition, the antigen-induced degranulation and activation of MAPKs were inhibited by WHI-P131 and WHI-P154 in both groups of BMMCs, indicating that these compounds inhibit a certain step except for Jak3. The antigen-induced increase in the activity of Fyn, a probable tyrosine kinase of Gab2, was also inhibited by WHI-P131 and WHI-P154 in RBL-2H3 cells. In BMMCs from Jak3-/- mice, the antigen stimulation induced tyrosine phosphorylation of Fyn, which was inhibited by WHI-P131, as well as in BMMCs from wild-type mice and in RBL-2H3 cells. These findings suggest that Jak3 does not play a significant role in the antigen-induced degranulation and phosphorylation of MAPKs, and that WHI-P131 and WHI-P154 inhibit the PI3K pathway by preventing the antigen-induced activation of Fyn, thus inhibiting the antigen-induced degranulation and phosphorylation of MAPKs in mast cells.
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PMID:Inhibition of the antigen-induced activation of rodent mast cells by putative Janus kinase 3 inhibitors WHI-P131 and WHI-P154 in a Janus kinase 3-independent manner. 1585 29

The p85alpha subunit of phosphatidylinositol 3-kinase (PI-3k) forms a complex with a protein network associated with oncogenic fusion tyrosine kinases (FTKs) such as BCR/ABL, TEL/ABL, TEL/JAK2, TEL/PDGFbetaR, and NPM/ALK, resulting in constitutive activation of the p110 catalytic subunit of PI-3k. Introduction of point mutations in the N-terminal and C-terminal SH2 domain and SH3 domain of p85alpha, which disrupt their ability to bind phosphotyrosine and proline-rich motifs, respectively, abrogated their interaction with the BCR/ABL protein network. The p85alpha mutant protein (p85mut) bearing these mutations was unable to interact with BCR/ABL and other FTKs, while its binding to the p110alpha catalytic subunit of PI-3k was intact. In addition, binding of Shc, c-Cbl, and Gab2, but not Crk-L, to p85mut was abrogated. p85mut diminished BCR/ABL-dependent activation of PI-3k and Akt kinase, the downstream effector of PI-3k. This effect was associated with the inhibition of BCR/ABL-dependent growth of the hematopoietic cell line and murine bone marrow cells. Interestingly, the addition of interleukin-3 (IL-3) rescued BCR/ABL-transformed cells from the inhibitory effect of p85mut. SCID mice injected with BCR/ABL-positive hematopoietic cells expressing p85mut survived longer than the animals inoculated with BCR/ABL-transformed counterparts. In conclusion, we have identified the domains of p85alpha responsible for the interaction with the FTK protein network and transduction of leukemogenic signaling.
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PMID:Phosphatidylinositol 3-kinase p85{alpha} subunit-dependent interaction with BCR/ABL-related fusion tyrosine kinases: molecular mechanisms and biological consequences. 1613 92

Although targeting the BCR-ABL tyrosine kinase activity by imatinib mesylate has rapidly become first-line therapy in chronic myeloid leukemia (CML), drug resistance suggests that combination therapy directed to a complementing target may significantly improve treatment results. To identify such potential targets, we used lentivirus-mediated RNA interference (RNAi) as a tool for functional genomics in cell lines as well as primary normal and CML CD34+ cells. In a conditional cell culture model, we demonstrate that RNAi-mediated reduction of SHP2, STAT5, and Gab2 protein expression inhibits BCR-ABL-dependent but not cytokine-dependent proliferation in a dose-dependent manner. Similarly, colony formation of purified primary CML but not of normal CD34+ colony-forming cells is specifically reduced by inhibition of SHP2, STAT5, and Gab2 expression, respectively. In addition, coexpression of both anti-BCR-ABL and anti-SHP2 shRNAs from a single lentiviral vector induces stronger inhibition of colony formation as compared to either shRNA alone. The data indicate that BCR-ABL expression may affect the function of normal signaling molecules. Targeting these molecules may harbor significant therapeutic potential for the treatment of patients with CML.
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PMID:Enhanced sensitivity to inhibition of SHP2, STAT5, and Gab2 expression in chronic myeloid leukemia (CML). 1627 4

Aggregation of the type 1 Fc-epsilon receptors (Fc-epsilon-RI) on mast cells initiates a network of biochemical processes culminating in secretion of both granule-stored and de novo-synthesized inflammatory mediators. A strict control of this response is obviously a necessity; nevertheless, this regulation is hardly characterized. Here we report that a prototype inhibitory receptor, the mast cell function-associated antigen (MAFA), selectively regulates the Fc-epsilon-RI stimulus-response coupling network and the subsequent de novo production and secretion of inflammatory mediators. Specifically, MAFA suppresses the PLC-gamma2-[Ca2+]i, Raf-1-Erk1/2, and PKC-p38 coupling pathways, while the Fyn-Gab2-mediated activation of PKB and Jnk is essentially unaffected. Hence, the activities of several transcription/nuclear factors for inflammatory mediators (NF-kappaB, NFAT) are markedly reduced, while those of others (Jun, Fos, Fra, p90rsk) are unaltered. This results in a selective inhibition of gene transcription of cytokines including IL-1beta, IL-4, IL-8, and IL-10, while that of TNF-alpha, MCP-1, IL-3, IL-5, or IL-13 remains unaffected. Taken together, these results illustrate the capacity of an immunoreceptor tyrosine-based inhibitory motif-containing receptor to cause tight and specific control of the production and secretion of inflammatory mediators by mast cells.
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PMID:Selective inhibition of the Fc epsilon RI-induced de novo synthesis of mediators by an inhibitory receptor. 3070 14


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