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)

Shear stress, the tangential component of hemodynamic forces, activates the extracellular signal-regulated kinase (ERK) and c-Jun NH2-terminal kinase (JNK) signal transduction pathways in cultured vascular endothelial cells to induce the transcriptional activation of many immediate early genes. It appears that integrins, protein-tyrosine kinases, and the structural integrity of actin are important factors involved in these shear stress-induced responses. The underlying molecular events were investigated by the application of a shear stress of 12 dyn/cm2 on bovine aortic endothelial cells (BAEC). We found that such a shear stress increased the tyrosine phosphorylation and the kinase activity of focal adhesion kinase (FAK) and its association with growth factor receptor binding protein 2 (Grb2) in a rapid and transient manner, suggesting that FAK may be linked to these mitogen-activated protein kinase signaling pathways through a Grb2. Son of sevenless (Sos) complex. FAK(F397Y), which encodes a dominant negative mutant of FAK, attenuated the shear stress-induced kinase activity of Myc epitope-tagged ERK2 and hemagglutinin epitope-tagged JNK1. DeltamSos1, encoding a dominant negative mutant of Sos in which the guanine nucleotide exchange domain has been deleted, also attenuated shear stress activation of Myc-ERK2 and hemagglutinin-JNK1. Pretreating the confluent BAEC monolayers with a blocking type anti-vitronectin receptor monoclonal antibody had similar inhibitory effects in these shear stress-activated ERKs and JNKs. Confocal microscopic observation further demonstrated that FAK tended to cluster with vitronectin receptor near the abluminal side of the sheared BAEC. These results demonstrate that FAK signaling is critical in the shear stress-induced dual activation of ERK and JNK.
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PMID:Fluid shear stress activation of focal adhesion kinase. Linking to mitogen-activated protein kinases. 937 37

Leptin receptors include a long form (OBRl) with 302 cytoplasmic residues that is presumed to mediate most or all of leptins signaling, and several short forms, including one (OBRs) that has 34 cytoplasmic residues, is widely expressed, and is presumed not to signal but to mediate transport or clearance of leptin. We studied the abilities of these two receptor isoforms to mediate signaling in transfected cells. In response to leptin, OBRl, but not OBRs, underwent tyrosine phosphorylation that was enhanced by co-expression with JAK2. In cells expressing receptors and JAK2, both OBRs and OBRl mediated leptin-dependent tyrosine phosphorylation of JAK2, and this was abolished with OBRs when the Box 1 motif was mutated. In cells expressing receptors, JAK2 and IRS-1, leptin induced tyrosine phosphorylation of IRS-1 through OBRs and OBRl. In COS cells expressing hemagglutinin-ERK1 and receptors, leptin increased ERK1 kinase activity through OBRl, with the magnitude increased by co-expression of JAK1 or JAK2, and to a lesser degree through OBRs, despite greater receptor expression. In stable Chinese hamster ovary cell lines expressing OBRs or OBRl, leptin stimulated endogenous ERK2 phosphorylation. Whereas leptin stimulated tyrosine phosphorylation of hemagglutinin-STAT3 and induction of a c-fos luciferase reporter plasmid through OBRl, OBRs was without effect in these assays. In conclusion, OBRl is capable of signaling to IRS-1 and mitogen-activated protein kinase via JAK, in addition to activating STAT pathways. Although substantially weaker than OBRl, OBRs is capable of mediating signal transduction via JAK, but these activities are of as yet unknown significance for leptin biology in vivo.
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PMID:Divergent signaling capacities of the long and short isoforms of the leptin receptor. 940 87

Recent results indicate that a fluoroalumino complex (AlFx) is probably the molecule responsible for the mitogenic effect of fluoride in MC3T3-E1 osteoblast-like cells. Initial analysis suggested that a tyrosine phosphorylation (tyr phos) process similar to that induced by thrombin and activation of the p42 MAP kinase (ERK 2) mediate this cellular response. In the present study, the signaling mechanism activated by AlFx was further investigated. The results indicated that AlFx dose-dependently enhanced the tyr phos of the cell adhesion proteins FAK and paxillin, as well as of the adaptor molecules p46shc, p52shc, and p66shc and their association with GRB2. Pretreatment of MC3T3-E1 cells with cytochalasin D completely prevented FAK and paxillin tyr phos without any alteration in the tyr phos of Shc proteins and activation of ERK2 induced by AlFx. This observation suggests that in confluent MC3T3-E1 cells, there is no link between the activation of FAK induced by AlFx and the stimulation of ERK2. Pretreatment of the cells with pertussis toxin inhibited Shc phosphorylation, activation of ERK2, and markedly reduced cell replication induced by AlFx. This toxin also significantly reduced the stimulation of Pi transport activity induced by AlFx in these cells. Alteration in tyr phos induced by AlFx was not associated with any detectable inhibition of tyrosine phosphatase activity in MC3T3-E1 cell homogenates, suggesting that enhanced tyr phos induced by AlFx probably resulted from activation of a tyrosine kinase. In conclusion, the results of this study suggest that the mitogenic effect of fluoride in MC3T3-E1 osteoblast-like cells is mediated by the activation of a pertussis toxin-sensitive Gi/o protein and suggest an important role for these heterotrimeric G proteins in controlling the growth and differentiation of bone-forming cells.
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PMID:Mechanism of the mitogenic effect of fluoride on osteoblast-like cells: evidences for a G protein-dependent tyrosine phosphorylation process. 942 Dec 30

We examined downstream signaling events that followed the exposure of PC12 cells to extracellular ATP and UTP, and we compared the effects of these P2 receptor agonists with those of growth factors and other stimuli. Based on early findings, we focused particular attention on the mitogen-activated protein (MAP) kinase pathway. ATP and/or UTP produced increases in tyrosine phosphorylation of multiple proteins, including p42 MAP (ERK2) kinase, related adhesion focal tyrosine kinase (RAFTK) (PYK2, CAKbeta), focal adhesion kinase (FAK), Shc, and protein kinase Cdelta (PKCdelta). MAP (ERK2) kinase activity (quantified by substrate phosphorylation) was increased by UTP, ATP, phorbol 12-myristate 13-acetate, ionomycin, and growth factors. UTP and ATP were equipotent (EC50 approximately 25 microM) in stimulating MAP kinase activity, suggesting that these effects were mediated via the Gi-linked P2Y2 (P2U) receptor. Consistent with this, the UTP- and ATP-promoted activation of MAP kinase was diminished in pertussis toxin-treated cells. Treatment of cells with pertussis toxin also reduced both the UTP-dependent increases in intracellular calcium ion concentration ([Ca2+]i) and the tyrosine phosphorylation of RAFTK. Similarly, when [Ca2+]i elevation was prevented using BAPTA and EGTA, the activation of MAP kinase by UTP and ionomycin was blocked, and the tyrosine phosphorylation of RAFTK was reduced. The UTP-promoted increase in MAP kinase activity was partially reduced in cells in which PKC was down-regulated, suggesting that both PKC-dependent and PKC-independent pathways were involved. PKCdelta, which increases MAP kinase activity in some systems, became tyrosine-phosphorylated within 15 s of exposure of cells to ATP or UTP; but epidermal growth factor, nerve growth factor, and insulin had little effect. UTP also promoted the association of Shc with Grb2. These results suggest that the P2Y2 receptor-initiated activation of MAP kinase was dependent on the elevation of [Ca2+]i, involved the recruitment of Shc and Grb2, and was mediated by RAFTK and PKC.
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PMID:Activation of P2Y2 receptors by UTP and ATP stimulates mitogen-activated kinase activity through a pathway that involves related adhesion focal tyrosine kinase and protein kinase C. 944 69

In vitro megakaryocytic differentiation of the pluripotent K562 human leukemia cell line is induced by PMA. Treatment of K562 cells with PMA results in growth arrest, polyploidy, morphological changes, and increased cell-cell and cell-substrate adhesion. These PMA-induced changes in K562 cells are preceded by a rapid rise in the activity of MEK (MAP kinase/extracellular regulated kinases) that leads to a sustained activation of ERK2 (extracellular regulated kinase; MAPK). Blockade of MEK1 activation by PD098059, a recently described specific MEK inhibitor [D. T. Dudley et al. (1995). Proc. Natl. Acad. Sci. USA 92, 7686-7689], reverses both the growth arrest and the morphological changes of K562 cells induced by PMA treatment. These changes are not associated with a disruption of PMA-induced down-regulation of BCR-ABL kinase or early integrin signaling events but are associated with a block of the cell-surface expression of the gpIIb/IIIa (CD41) integrin, a cell marker of megakaryocytic differentiation. These results demonstrate that the PMA-induced signaling cascade initiated by protein kinase C activation requires the activity of the MEK/ERK signaling complex to regulate cell cycle arrest, thus regulating the program that leads to the cell-surface expression of markers associated with megakaryocytic differentiation.
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PMID:A role for the MEK/MAPK pathway in PMA-induced cell cycle arrest: modulation of megakaryocytic differentiation of K562 cells. 947 49

Protein tyrosine phosphorylation and other biochemical events have been shown to occur after cross-linking of Fc epsilonRI in rodent mast cells. To investigate the mechanism of Fc epsilonRI signal transduction in human mast cells, we used human cultured mast cells (HCMC) generated from cord blood cells in the presence of recombinant human stem cell factor and IL-6. We found that on cross-linking of Fc epsilonRI: 1) HCMC released histamine; 2) rapid tyrosine phosphorylation of multiple cellular substrates, including Syk, HS1, c-Cbl, ERK-1, and ERK-2, was observed; 3) intracellular Ca2+ and inositol phosphate production were increased within the first minute after Fc epsilonRI cross-linking; and 4) genistein, a tyrosine kinase inhibitor, inhibited both protein tyrosine phosphorylation and histamine release in a dose-dependent manner. These results were consistent with previous studies in rodent mast cells. In contrast, no tyrosine phosphorylation of phospholipase C gamma1 and Btk (Bruton's tyrosine kinase) were observed in our experimental conditions. These results suggest that the greater part of the early and late signaling events in HCMC is similar to those obtained with rodent mast cells and indicated that the requirement of tyrosine phosphorylation in the activation process of each of the signaling molecules might be different in HCMC and rodent mast cells. Our finding indicates that HCMC may be useful for analysis of Fc epsilonRI-mediated signal transduction in human mast cells.
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PMID:Early and late events in Fc epsilon RI signal transduction in human cultured mast cells. 955 Mar 84

Fibronectin receptor integrin-mediated cell adhesion triggers intracellular signaling events such as the activation of the Ras/mitogen-activated protein (MAP) kinase cascade. In this study, we show that the nonreceptor protein-tyrosine kinases (PTKs) c-Src and focal adhesion kinase (FAK) can be independently activated after fibronectin (FN) stimulation and that their combined activity promotes signaling to extracellular signal-regulated kinase 2 (ERK2)/MAP kinase through multiple pathways upstream of Ras. FN stimulation of NIH 3T3 fibroblasts promotes c-Src and FAK association in the Triton-insoluble cell fraction, and the time course of FN-stimulated ERK2 activation paralleled that of Grb2 binding to FAK at Tyr-925 and Grb2 binding to Shc. Cytochalasin D treatment of fibroblasts inhibited FN-induced FAK in vitro kinase activity and signaling to ERK2, but it only partially inhibited c-Src activation. Treatment of fibroblasts with protein kinase C inhibitors or with the PTK inhibitor herbimycin A or PP1 resulted in reduced Src PTK activity, no Grb2 binding to FAK, and lowered levels of ERK2 activation. FN-stimulated FAK PTK activity was not significantly affected by herbimycin A treatment and, under these conditions, FAK autophosphorylation promoted Shc binding to FAK. In vitro, FAK directly phosphorylated Shc Tyr-317 to promote Grb2 binding, and in vivo Grb2 binding to Shc was observed in herbimycin A-treated fibroblasts after FN stimulation. Interestingly, c-Src in vitro phosphorylation of Shc promoted Grb2 binding to both wild-type and Phe-317 Shc. In vivo, Phe-317 Shc was tyrosine phosphorylated after FN stimulation of human 293T cells and its expression did not inhibit signaling to ERK2. Surprisingly, expression of Phe-925 FAK with Phe-317 Shc also did not block signaling to ERK2, whereas FN-stimulated signaling to ERK2 was inhibited by coexpression of an SH3 domain-inactivated mutant of Grb2. Our studies show that FN receptor integrin signaling upstream of Ras and ERK2 does not follow a linear pathway but that, instead, multiple Grb2-mediated interactions with Shc, FAK, and perhaps other yet-to-be-determined phosphorylated targets represent parallel signaling pathways that cooperate to promote maximal ERK2 activation.
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PMID:Multiple Grb2-mediated integrin-stimulated signaling pathways to ERK2/mitogen-activated protein kinase: summation of both c-Src- and focal adhesion kinase-initiated tyrosine phosphorylation events. 956 77

Cells grown in 3-dimensional collagen gels adopt a nonproliferative, contractile phenotype which is more characteristic of cells in vivo than cells grown in 2-dimensional culture. The floating collagen gel contraction assay is a well-defined system used to study cell-extracellular matrix interactions grown in 3-dimensional culture. Although the cell biology of this system is well defined, the cell signaling associated with gel contraction has not been well characterized. In this study we demonstrate that fetal bovine (FBS) and platelet-derived growth factor (PDGF)-induced mesangial cell-collagen gel contraction is associated with increased tyrosine phosphorylation of a number of proteins including focal adhesion kinase (FAK) and the 42-kDa isoform of MAPK (ERK2). FBS-induced gel contraction is not affected by the presence of the MEK inhibitor PD098059. Low concentrations of PDGF-BB (10 ng/ml) induce gel contraction; however, at higher PDGF-BB concentrations (80 ng/ml) gel contraction is not observed. PDGF-BB-induced gel contraction as well as tyrosine phosphorylation of FAK are inhibited in the presence of the PI-3 kinase inhibitor wortmanin. Minimal autophosphorylation of the PDGF-beta receptor is observed under 3-dimensional culture conditions following PDGF-BB stimulation; however, when mesangial cells grown in 2-dimensional culture are exposed to PDGF-BB, the PDGF-beta receptor was prominently phosphorylated. We conclude that induction of collagen gel contraction by FBS and PDGF-BB is associated with tyrosine kinase phosphorylation and that these responses differ substantially from what occurs in 2-dimensional cultures in the presence of the same agonists.
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PMID:Tyrosine kinase cell signaling pathways of rat mesangial cells in 3-dimensional cultures: response to fetal bovine serum and platelet-derived growth factor-BB. 957 Sep 28

In preadipocytes, alpha2-adrenergic receptor (alpha2-AR) stimulation leads to a Gi/Go-dependent rearrangement of actin cytoskeleton. This is characterized by a rapid cell spreading, the formation of actin stress fibers, and the increase in tyrosyl phosphorylation of the focal adhesion kinase (pp125(FAK)). These cellular events being tightly controlled by the small GTPase p21(rhoA), the existence of a Gi/Go-dependent coupling of alpha2-AR to p21(rhoA) in preadipocytes was proposed. In alpha2AF2 preadipocytes (a cell clone derived from the 3T3F442A preadipose cell line and which stably expresses the human alpha2C10-adrenergic receptor) alpha2-adrenergic-dependent induction of cell spreading, formation of actin stress fibers, and increase in tyrosyl phosphorylation of pp125(FAK) were abolished by pretreatment of the preadipocytes with the C3 exoenzyme, a toxin which impairs p21(rhoA) activity by ADP-ribosylation. Conversely, C3 exoenzyme had no effect on the alpha2-adrenergic-dependent increase in tyrosyl phosphorylation and shift of ERK2 mitogen-activated protein kinase. alpha2-Adrenergic stimulation also led to an increase in GDP/GTP exchange on p21(rhoA), as well as to an increase in the amount of p21(rhoA) in the particulate fraction of alpha2AF2 preadipocytes. Stable transfection of alpha2AF2 preadipocytes with the COOH-terminal domain of betaARK1 (betaARK-CT) (a blocker of Gbeta gamma-action), strongly inhibited the alpha2-adrenergic-dependent increase in tyrosyl phos- phorylation and shift of ERK2, without modification of the tyrosyl phosphorylation of pp125(FAK) and spreading of preadipocytes. These results show that alpha2-adrenergic-dependent reorganization of actin cytoskeleton requires the activation of p21(rhoA) in preadipocytes. Conversely to the activation of the p21(ras)/mitogen-activated protein kinase pathway, the alpha2-adrenergic activation of p21(rhoA)-dependent pathways are independent of the beta gamma-subunits of heterotrimeric G proteins.
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PMID:Gbeta gamma-independent coupling of alpha2-adrenergic receptor to p21(rhoA) in preadipocytes. 962 80

The organization of the actin cytoskeleton can be regulated by soluble factors that trigger signal transduction events involving the Rho family of GTPases. Since adhesive interactions are also capable of organizing the actin-based cytoskeleton, we examined the role of Cdc42-, Rac-, and Rho-dependent signaling pathways in regulating the cytoskeleton during integrin-mediated adhesion and cell spreading using dominant-inhibitory mutants of these GTPases. When Rat1 cells initially adhere to the extracellular matrix protein fibronectin, punctate focal complexes form at the cell periphery. Concomitant with focal complex formation, we observed some phosphorylation of the focal adhesion kinase (FAK) and Src, which occurred independently of Rho family GTPases. However, subsequent phosphorylation of FAK and paxillin occurs in a Rho-dependent manner. Moreover, we found Rho dependence of the assembly of large focal adhesions from which actin stress fibers radiate. Initial adhesion to fibronectin also stimulates membrane ruffling; we show that this ruffling is independent of Rho but is dependent on both Cdc42 and Rac. Furthermore, we observed that Cdc42 controls the integrin-dependent activation of extracellular signal-regulated kinase 2 and of Akt, a kinase whose activity has been demonstrated to be dependent on phosphatidylinositol (PI) 3-kinase. Since Rac-dependent membrane ruffling can be stimulated by PI 3-kinase, it appears that Cdc42, PI 3-kinase, and Rac lie on a distinct pathway that regulates adhesion-induced membrane ruffling. In contrast to the differential regulation of integrin-mediated signaling by Cdc42, Rac, and Rho, we observed that all three GTPases regulate cell spreading, an event that may indirectly control cellular architecture. Therefore, several separable signaling pathways regulated by different members of the Rho family of GTPases converge to control adhesion-dependent changes in the organization of the cytoskeleton, changes that regulate cell morphology and behavior.
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PMID:Integrin-mediated signals regulated by members of the rho family of GTPases. 967 53


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