Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.1 (protein kinase)
 81,284 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Phospholipase Cepsilon (PLCepsilon) is a novel class of phosphoinositide-specific PLC characterized by possession of CDC25 homology and Ras/Rap1-associating domains. We and others have shown that human PLCepsilon is translocated from the cytoplasm to the plasma membrane and activated by direct association with Ras at its Ras/Rap1-associating domain. In addition, translocation to the perinuclear region was induced upon association with Rap1.GTP. However, the function of the CDC25 homology domain remains to be clarified. Here we show that the CDC25 homology domain of PLCepsilon functions as a guanine nucleotide exchange factor for Rap1 but not for any other Ras family GTPases examined including Rap2 and Ha-Ras. Consistent with this, coexpression of full-length PLCepsilon or its N-terminal fragment carrying the CDC25 homology domain causes an increase of the intracellular level of Rap1.GTP. Concurrently, stimulation of the downstream kinases B-Raf and extracellular signal-regulated kinase is observed, whereas the intracellular level of Ras.GTP and Raf-1 kinase activity are unaffected. In wild-type Rap1-overexpressing cells, epidermal growth factor induces translocation of PLCepsilon to the perinuclear compartments such as the Golgi apparatus, which is sustained for at least 20 min. In contrast, PLCepsilon lacking the CDC25 domain translocates to the perinuclear compartments only transiently. Further, the formation of Rap1.GTP upon epidermal growth factor stimulation exhibits a prolonged time course in cells expressing full-length PLCepsilon compared with those expressing PLCepsilon lacking the CDC25 homology domain. These results suggest a pivotal role of the CDC25 homology domain in amplifying Rap1-dependent signal transduction, including the activation of PLCepsilon itself, at specific subcellular locations such as the Golgi apparatus.
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PMID:Role of the CDC25 homology domain of phospholipase Cepsilon in amplification of Rap1-dependent signaling. 1139 6

Epac1 is a guanine nucleotide exchange factor for Rap1 that is activated by direct binding of cAMP. In vitro studies suggest that cAMP relieves the interaction between the regulatory and catalytic domains of Epac. Here, we monitor Epac1 activation in vivo by using a CFP-Epac-YFP fusion construct. When expressed in mammalian cells, CFP-Epac-YFP shows significant fluorescence resonance energy transfer (FRET). FRET rapidly decreases in response to the cAMP-raising agents, whereas it fully recovers after addition of cAMP-lowering agonists. Thus, by undergoing a cAMP-induced conformational change, CFP-Epac-YFP serves as a highly sensitive cAMP indicator in vivo. When compared with a protein kinase A (PKA)-based sensor, Epac-based cAMP probes show an extended dynamic range and a better signal-to-noise ratio; furthermore, as a single polypeptide, CFP-Epac-YFP does not suffer from the technical problems encountered with multisubunit PKA-based sensors. These properties make Epac-based FRET probes the preferred indicators for monitoring cAMP levels in vivo.
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PMID:Detecting cAMP-induced Epac activation by fluorescence resonance energy transfer: Epac as a novel cAMP indicator. 1555 Sep 31

Cyclic AMP (cAMP) is a well-known intracellular signaling molecule improving barrier function in vascular endothelial cells. Here, we delineate a novel cAMP-triggered signal that regulates the barrier function. We found that cAMP-elevating reagents, prostacyclin and forskolin, decreased cell permeability and enhanced vascular endothelial (VE) cadherin-dependent cell adhesion. Although the decreased permeability and the increased VE-cadherin-mediated adhesion by prostacyclin and forskolin were insensitive to a specific inhibitor for cAMP-dependent protein kinase, these effects were mimicked by 8-(4-chlorophenylthio)-2'-O-methyladenosine-3', 5'-cyclic monophosphate, a specific activator for Epac, which is a novel cAMP-dependent guanine nucleotide exchange factor for Rap1. Thus, we investigated the effect of Rap1 on permeability and the VE-cadherin-mediated cell adhesion by expressing either constitutive active Rap1 or Rap1GAPII. Activation of Rap1 resulted in a decrease in permeability and enhancement of VE-cadherin-dependent cell adhesion, whereas inactivation of Rap1 had the counter effect. Furthermore, prostacyclin and forskolin induced cortical actin rearrangement in a Rap1-dependent manner. In conclusion, cAMP-Epac-Rap1 signaling promotes decreased cell permeability by enhancing VE-cadherin-mediated adhesion lined by the rearranged cortical actin.
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PMID:Cyclic AMP potentiates vascular endothelial cadherin-mediated cell-cell contact to enhance endothelial barrier function through an Epac-Rap1 signaling pathway. 1560 37

Here, we demonstrate that elevation of intracellular cyclic AMP (cAMP) in vascular endothelial cells (ECs) by either a direct activator of adenylyl cyclase or endogenous cAMP-mobilizing G protein-coupled receptors inhibited the tyrosine phosphorylation of STAT proteins by an interleukin 6 (IL-6) receptor trans-signaling complex (soluble IL-6Ralpha/IL-6). This was associated with the induction of suppressor of cytokine signaling 3 (SOCS-3), a bona fide inhibitor in vivo of gp130, the signal-transducing component of the IL-6 receptor complex. Attenuation of SOCS-3 induction in either ECs or SOCS-3-null murine embryonic fibroblasts abolished the inhibitory effect of cAMP, whereas inhibition of SHP-2, another negative regulator of gp130, was without effect. Interestingly, the inhibition of STAT phosphorylation and SOCS-3 induction did not require cAMP-dependent protein kinase activity but could be recapitulated upon selective activation of the alternative cAMP sensor Epac, a guanine nucleotide exchange factor for Rap1. Consistent with this hypothesis, small interfering RNA-mediated knockdown of Epac1 was sufficient to attenuate both cAMP-mediated SOCS-3 induction and inhibition of STAT phosphorylation, suggesting that Epac activation is both necessary and sufficient to observe these effects. Together, these data argue for the existence of a novel cAMP/Epac/Rap1/SOCS-3 pathway for limiting IL-6 receptor signaling in ECs and illuminate a new mechanism by which cAMP may mediate its potent anti-inflammatory effects.
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PMID:Exchange protein activated by cyclic AMP (Epac)-mediated induction of suppressor of cytokine signaling 3 (SOCS-3) in vascular endothelial cells. 1691 20

Signaling by the B cell antigen receptor (BCR) is essential for B lymphocyte homeostasis and immune function. In immature B cells, ligation of the BCR promotes growth arrest and apoptosis, and BCR-driven balancing between pro-apoptotic extracellular signal-regulated kinase 1 and 2 (ERK1/2) and anti-apoptotic phosphoinositide 3-kinase-dependent Akt seems to define the final cellular apoptotic response. Dysfunction of these late BCR signaling events can lead to the development of immunological diseases. Here we report on novel cyclic AMP-dependent mechanisms of BCR-induced growth arrest and apoptosis in the immature B lymphoma cell line WEHI-231. BCR signaling to ERK1/2 and Akt requires cyclic AMP-regulated Epac, the latter acting as a guanine nucleotide exchange factor for Rap1 and H-Ras independent of protein kinase A. Importantly, activation of endogenously expressed Epac by a specific cyclic AMP analog enhanced the induction of growth arrest (reduced DNA synthesis) and apoptosis (nuclear condensation, annexin V binding, caspase-3 cleavage and poly-ADP-ribose polymerase processing) by the BCR. Our data indicate that cyclic AMP-dependent Epac signals to ERK1/2 and Akt upon activation of Rap1 and H-Ras, and is involved in BCR-induced growth arrest and apoptosis in WEHI-231 cells.
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PMID:B cell receptor-induced growth arrest and apoptosis in WEHI-231 immature B lymphoma cells involve cyclic AMP and Epac proteins. 1916 86

The monomeric GTPase Rap1 controls functional activation of beta2 integrins in leukocytes. In this article, we describe a novel mechanism by which the chemoattractant fMLP activates Rap1 and inside-out signaling of beta2 integrins. We found that fMLP-induced activation of Rap1 in human polymorphonuclear leukocytes or neutrophils and differentiated PLB-985 cells was blocked by inhibitors of the NO/guanosine-3',5'-cyclic monophosphate-dependent protein kinase (cGKI) pathway [N-(3-(aminomethyl)benzyl)acetamidine, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one, DT-3 peptide, 8-(4-chlorophenylthio)guanosine 3',5'-cyclic monophosphothioate, Rp-isomer triethylammonium salt-guanosine-3',5'-cyclic monophosphate], indicating that the downstream signaling events in Rap1 activation involve the production of NO and guanosine-3',5'-cyclic monophosphate, as well as the activation of cGKI. Silencing the expression of vasodilator-stimulated phosphoprotein (VASP), a substrate of cGKI, in resting PLB-985 cells or mice neutrophils led to constitutive activation of Rap1. In parallel, silencing VASP in differentiated PLB-985 cells led to recruitment of C3G, a guanine nucleotide exchange factor for Rap1, to the plasma membrane. Expression of murine GFP-tagged phosphodeficient VASP Ser235Ala mutant (murine serine 235 of VASP corresponds to human serine 239) in PLB-985 cells blunted fMLP-induced translocation of C3G to the membrane and activation of Rap1. Thus, bacterial fMLP triggers cGKI-dependent phosphorylation of human VASP on serine 239 and, thereby, controls membrane recruitment of C3G, which is required for activation of Rap1 and beta2 integrin-dependent antibacterial functions of neutrophils.
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PMID:Vasodilator-stimulated phosphoprotein regulates inside-out signaling of beta2 integrins in neutrophils. 2048 41