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:2.7.12.2 (MEK)
18,161 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We characterized the tracheal and bronchial relaxation caused by proteinase-activated receptor-2 (PAR-2) activation in ddY mice and/or in wild-type and PAR-2-knockout mice of C57BL/6 background. Ser-Leu-Ile-Gly-Arg-Leu-amide (SLIGRL-NH(2)) and Thr-Phe-Leu-Leu-Arg-amide, PAR-2- and PAR-1-activating peptides, respectively, caused relaxation in the isolated ddY mouse trachea and main bronchus. The relaxation was abolished by specific inhibitors of cyclooxygenase (COX)-1, COX-2, mitogen-activated protein kinase kinase (MEK), and p38 MAP kinase. The MEK and p38 MAP kinase inhibitors did not affect prostaglandin E(2)-induced relaxation. Inhibitors of cytosolic Ca(2+)-dependent phospholipase A(2) (PLA), Ca(2+)-independent PLA(2), diacylglycerol lipase, tyrosine kinase, and protein kinase C exhibited no or only minor inhibitory effects on the PAR-mediated relaxation. Trypsin, a PAR-2 activator, and 2-furoyl-Leu-Ile-Gly-Arg-Leu-amide, a potent PAR-2-activating peptide, in addition to SLIGRL-NH(2), caused airway relaxation in wild-type C57BL/6 mice, as in ddY mice. In PAR-2-knockout mice, the peptide effects were absent and the potency of trypsin decreased. Desensitization of PAR-2 and/or PAR-1 greatly suppressed the relaxant effect of trypsin. The bronchial and tracheal tissues displayed distinct sensitivities toward trypsin and the PAR-2-activating peptides. Our data indicate an involvement of both COX-1 and COX-2, and the MEK-extracellular signal-regulated kinase and p38 MAP kinase signaling pathways in the PAR-2- and PAR-1-triggered relaxation of mouse airway tissue, and substantiate a role for PAR-2 in regulating both the trachea and bronchial responsiveness in the mouse lung.
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PMID:Proteinase-activated receptor-2-mediated relaxation in mouse tracheal and bronchial smooth muscle: signal transduction mechanisms and distinct agonist sensitivity. 1519 93

Protease-activated receptor-2 (PAR-2) is activated by trypsin-like serine proteases and can promote cell migration through an ERK1/2-dependent pathway, involving formation of a scaffolding complex at the leading edge of the cell. Previous studies also showed that expression of a dominant negative fragment of beta-arrestin-1 reduces PAR-2-stimulated internalization, ERK1/2 activation, and cell migration; however, this reagent may block association of many proteins, including beta-arrestin-2 with clathrin-coated pits. Here we investigate the role of PAR-2 in the constitutive migration of a metastatic breast cancer cell line, MDA MB-231, and use small interfering RNA to determine the contribution of each beta-arrestin to this process. We demonstrate that a trypsin-like protease secreted from MDA MB-231 cells can promote cell migration through autocrine activation of PAR-2 and this correlates with constitutive localization of PAR-2, beta-arrestin-2, and activated ERK1/2 to pseudopodia. Addition of MEK-1 inhibitors, trypsin inhibitors, a scrambled PAR-2 peptide, and silencing of beta-arrestins with small interfering RNA also reduce base-line migration of MDA MB-231 cells. In contrast, a less metastatic PAR-2 expressing breast cancer cell line does not exhibit constitutive migration, pseudopodia formation, or trypsin secretion; in these cells PAR-2 is more uniformly distributed around the cell periphery. These data demonstrate a requirement for both beta-arrestins in PAR-2-mediated motility and suggest that autocrine activation of PAR-2 by secreted proteases may contribute to the migration of metastatic tumor cells through beta-arrestin-dependent ERK1/2 activation.
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PMID:Constitutive protease-activated receptor-2-mediated migration of MDA MB-231 breast cancer cells requires both beta-arrestin-1 and -2. 1548 20

The mast cell product tryptase, via protease-activated receptor 2 (PAR2), induces cyclooxygenase-2 (COX2) and 15-deoxy-prostaglandin J2 (15d-PGJ2) synthesis. 15d-PGJ2, through the nuclear peroxisome proliferator activated receptor gamma (PPARgamma), subsequently causes fibroblast proliferation. In this study we attempted to determine initial events of the tryptase/PAR2 signaling pathway leading to COX2 induction and fibroblast proliferation. In human fibroblasts (HFFF2), cDNA array, RT-PCR and Western blotting studies demonstrated that tryptase, but not 15d-PGJ2, up-regulates c-jun, c-fos and COX2 expression, and phosphorylates the extracellular signal-regulated kinase isoforms 1 and 2 (erk1/2). Furthermore, tryptase effects on erk1/2, c-jun, c-fos, COX2 and cell proliferation were prevented by PD98059, an inhibitor of the mitogen-activated protein kinase kinase (MEK). Other kinases [P38, stress-activated protein kinase/c-jun N-terminal kinase (SAPK/JUNK), erk5], intracellular Ca(2+) or cAMP were not affected by tryptase/PAR2. Our study identifies crucial intracellular events leading to induction of COX2 and fibroblast proliferation, i.e. a cornerstone of fibrosis.
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PMID:The action of the mast cell product tryptase on cyclooxygenase-2 (COX2) and subsequent fibroblast proliferation involves activation of the extracellular signal-regulated kinase isoforms 1 and 2 (erk1/2). 1560 29

We found that striptease-positive mast cells were abundant in the invasive front of human colon adenocarcinoma by examining 30 cases. Because tryptase has been suggested to be the agonist proteinase for protease-activated receptor-2 (PAR-2), we investigated the effects of stimulation of PAR-2 by tryptase on the cell signaling and proliferation of DLD-1, a human colon carcinoma cell line. PAR-2 stimulation by tryptase induced the increase in [Ca(2+)](i), which was desensitized by the prior application of PAR-2 activating peptide (AP). The proliferative responses of DLD-1 to tryptase and PAR-2 AP were associated with the phosphorylation of MEK and MAP kinase. Inhibition of MEK by PD98059 completely inhibited the proliferation-enhancing effects of tryptase and PAR-2 AP as well as phosphorylation of MAP kinase. Moreover, tryptase and PAR-2 AP stimulated the production of prostaglandin E2 and the inhibition of prostaglandin synthesis by indomethacin or NS398 resulted in the complete inhibition of the proliferative responses to tryptase and PAR-2 AP. Furthermore, the tryptase-stimulated proliferation of DLD-1 was concentration-dependently inhibited by nafamostat mesilate, a specific inhibitor of tryptase. These results as a whole indicated that tryptase has proliferative effects on DLD-1 through cyclooxygenase- and MAP kinase-dependent manners acting on PAR-2 by its proteolytic activity.
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PMID:Mast cell tryptase stimulates DLD-1 carcinoma through prostaglandin- and MAP kinase-dependent manners. 1609 13

Human airway trypsin-like protease (HAT) was isolated from airway secretions and localized to bronchial epithelial cells by immunohistochemistry. In the present study, we examined whether HAT could stimulate DNA synthesis and proliferation of primary human bronchial fibroblasts (HBF). HAT significantly stimulated the proliferation of HBF by 20-55%, a level similar to that of the mitogenic activity of lung mast cell tryptase (MCT). HAT also stimulated the incorporation of [3H]thymidine in HBF, and this HAT-induced DNA synthesis was abolished by leupeptin. Protease-activated receptor-2 (PAR-2) mRNA was expressed and localized to the cell surface in HBF. PAR-2 activating peptide (AP) also enhanced DNA synthesis, and both HAT and PAR-2 AP induced receptor internalization, similar to the response to trypsin. Pretreatment of HBF with anti-PAR-2 antibody significantly suppressed both HAT and PAR-2 AP-induced DNA synthesis. In addition, HAT and PAR-2 AP induced intracellular Ca2+ mobilization in HBF. The HAT-induced increase in Ca2+ was desensitized by pretreatment with trypsin or PAR-2 AP. U0126, a specific MAPK inhibitor, completely inhibited HAT-induced DNA synthesis as well as HAT-induced phosphorylation of MAPK. The effect of HAT and MCT together was additive, whereas the effect of HAT and insulin together on HBF DNA synthesis was synergistic. These results indicate that HAT stimulates fibroblast proliferation in bronchial airways through a PAR-2-dependent MEK-MAPK mediated pathway and that HAT is linked to airway processes involving fibroblasts.
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PMID:Human airway trypsin-like protease stimulates human bronchial fibroblast proliferation in a protease-activated receptor-2-dependent pathway. 1619 37

A member of a new subfamily of G protein-coupled receptors, protease-activated receptor 2 (PAR2), is highly expressed on endothelial cells and plays an important role in inflammation. The purpose of this study was to determine the molecular mechanism used by PAR2 to induce IL-8 production and thereby mediate cell adhesion. We observed that PAR2-activating peptide (PAR2-AP) significantly increase peripheral blood mononuclear cells adhere to endothelial cells. Both PAR2-AP and the endogenous PAR2 activator trypsin caused concentration- and time-dependent increase in endothelial IL-8 production, and this effect was concentration dependently and selectively attenuated by the p38 mitogen-activated protein kinase (MAPK) inhibitor SB203580. Western blotting analysis showed that PAR2-AP induced phosphorylation of p38 MAPK and its upstream protein kinase MAPK kinase 3/6 (MKK3/6) in a time-dependent manner. Using reverse-transcriptase-polymerase chain reaction and enzyme-linked immunosorbent assay, PAR2-AP was found to cause an increase in IL-8 mRNA expression and its transcription factor activating transcription factor 2, respectively,. As expected, these signals were suppressed by SB203580 in a concentration-dependent manner. Furthermore, introduction of dominant-negative vectors targeting p38 MAPK, MKK3, and MKK6 abolished PAR2-AP-mediated IL-8 production and cell adhesion function. In conclusion, PAR2 via p38 MAPK signaling regulates IL-8 production and thereby mediates cell adhesion.
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PMID:The p38 mitogen-activated protein kinase pathway plays a critical role in PAR2-induced endothelial IL-8 production and leukocyte adhesion. 1827 46

Proteinase-activated receptor (PAR)(2) is activated by trypsin-like serine proteinases and has been implicated in intestinal inflammation. However, its role in the regulation of intestinal mucosal function remains unclear. Using the intestinal epithelial cell line, SCBN, we have studied the stimulus-secretion coupling mechanisms of PAR(2)-induced epithelial chloride transport, focusing on cyclooxygenase (COX)-1 and COX-2 activities and prostaglandin (PG) E(2) secretion. SCBN monolayers were grown on Snapwell supports, mounted in modified Ussing chambers, and exposed to the activating peptide, SLIGRL-NH(2) (50 microM), to activate PAR(2). Pretreatment with inhibitors of cytosolic PLA(2) (cPLA(2)) (AACOCF3, arachidonyltrifluoromethyl ketone), COX-1 [SC560, 5-(4-chlorophenyl)-1-(4-methoxyphenyl)-3-(trifluoromethyl)-1H-pyrazole], and COX-2 (celecoxib) resulted in a significant concentration-dependent attenuation of PAR(2)-induced changes in short-circuit current. Immunoblot analysis showed a PAR(2)-induced increase in cPLA(2) phosphorylation that was blocked by the mitogen-activated protein kinase kinase inhibitor, PD98059 [2-(2-amino-3methoxyphenyl)-4H-1benzopyran-4-one, C(16)H(13)NO(3)], and the pan-protein kinase C inhibitor, GFX (bisindolylmaleimide). PAR(2) stimulation also resulted in a large increase in the production of PGE(2) as determined by enzyme-linked immunosorbent assay and was also blocked by PD98059 and GFX. Immunofluorescence and immunoblot analysis determined that EP2 and EP4 are expressed at the basolateral membrane of SCBN cells. Through the use of selective inhibitors (EP2, AH6809 [6-isopropoxy-9-oxoxanthene-2-carboxylic acid]; EP4, GW627368X [N-[2[4,9-diethoxy-1-oxo-1,3-dihydro-2H-benzo[f]isoindol-2-yl)phenyl] acetyl]benzene sulphonamide]), it was found that both EP2 and EP4 were involved in mediating the PAR(2)-induced chloride secretory response. We conclude that basolateral PAR(2) activation induces epithelial chloride secretion that is mediated by cPLA(2), COX-1, COX-2, and the subsequent release of PGE(2). The production of PGE(2) results in an autocrine secretory response that is dependent on basolateral EP2 and EP4 receptors.
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PMID:Prostaglandin E2 derived from cyclooxygenases 1 and 2 mediates intestinal epithelial ion transport stimulated by the activation of protease-activated receptor 2. 1919 Feb 38

In a wild tobacco plant, Nicotiana attenuata, two mitogen-activated protein kinases (MAPKs), salicylic acid-induced protein kinase (SIPK) and wound-induced protein kinase (WIPK), play central roles in modulating herbivory-induced phytohormone and anti-herbivore secondary metabolites. However, the identities of their upstream MAPK kinases (MAPKKs) were elusive. Ectopic overexpression studies in N. benthamiana and N. tabacum suggested that two MAPKKs, MKK1 and MEK2, may activate SIPK and WIPK. The homologues of MKK1 and MEK2 were cloned in N. attenuata (NaMKK1 and NaMEK2) and a virus-induced gene silencing approach was used to knock-down the transcript levels of these MAPKK genes. Plants silenced in NaMKK1 and NaMEK2 were treated with wounding or simulated herbivory by applying the oral secretions of the specialist herbivore Manduca sexta to wounds. MAPK activity assay indicated that after wounding or simulated herbivory NaMKK1 is not required for the phosphorylation of NaSIPK and NaWIPK; in contrast, NaMEK2 and other unknown MAPKKs are important for simulated herbivory-elicited activation of NaSIPK and NaWIPK, and after wounding NaMEK2 probably does not activate NaWIPK but plays a minor role in activating NaSIPK. Consistently, NaMEK2 and certain other MAPKKs, but not NaMKK1, are needed for wounding- and simulated herbivory-elicited accumulation of jasmonic acid (JA), JA-isoleucine, and ethylene. Furthermore, both NaMEK2 and NaMKK1 regulate the levels of trypsin proteinase inhibitors. The findings underscore the complexity of MAPK signalling pathways and highlight the importance of MAPKKs in regulating wounding- and herbivory-induced responses.
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PMID:Two mitogen-activated protein kinase kinases, MKK1 and MEK2, are involved in wounding- and specialist lepidopteran herbivore Manduca sexta-induced responses in Nicotiana attenuata. 2161 19

Osteosarcoma is the most common malignant primary bone tumor in children and adolescents and is characterized by a high metastatic potential. Its clinical outcome remains discouraging despite aggressive treatments. Thus, novel therapeutic approaches are needed. Recent results indicate that inorganic phosphate (Pi) is capable of affecting specific signal transduction pathways and of acting as an active regulator of cell behaviour. Previously, we found that Pi inhibits proliferation of human osteosarcoma U2OS cells via an adenylate cyclase/cAMP mediated mechanism. Here, we report that upon Pi treatment, U2OS cells become extremely hard to dislodge with trypsin. The lack of sensitivity to the trypsin action was paralleled by relevant changes in integrin subunits expression and accompanied by an increase of cell adhesion in cell-matrix adhesion assays. Interestingly, exposure of U2OS cells to Pi results also in a strong activation and protein level up-regulation of Rap1 small GTPase and in an early increase followed by a sustained inhibition of Erk1/2 phosphorylation. Importantly, the Pi-induced increase of cell adhesion was enforced by a cAMP analogue which specifically activated Epac/Rap1 and insensitive to PKA and MEK1/2 inhibitors. Our results enforce the evidences of inorganic phosphate as a signalling molecule, identify beta3 integrin, Rap1, ERK1/2 as proteins whose expression and function are relevantly affected by Pi in osteosarcoma U2OS cells The clinical significance and potential therapeutic applications by our data will be discussed.
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PMID:Novel molecular mechanisms by inorganic phosphate in osteosarcoma U2OS cells. 2162 31

Many motile processes are regulated such that movement occurs only upon activation of a signaling cascade. Sperm from a variety of species are initially quiescent and must be activated prior to beating. The signaling events leading to the activation and regulation of sperm motility are not well characterized. Mature seminal vesicle sperm from the water strider Aquarius remigis are immotile in vitro, but vigorous motility is activated by trypsin. Trypsin-activated motility was blocked by pretreatment of the sperm with BAPTA-AM to chelate intracellular Ca(2+) and was partially rescued by subsequent addition of A23187 and Ca(2+). Thapsigargin stimulated motility in the absence of trypsin, suggesting that intracellular Ca(2+) stores are available. In addition, motility could be fully activated by the phosphatase inhibitor calyculin A, suggesting that the immotile state is maintained by an endogenous phosphatase and that kinase activity is required for motility. The MEK1/2 inhibitor U0126 significantly reduced trypsin activated motility, and MPM-2, an antibody which recognizes proline-directed phosphorylation by kinases such as MAPK, recognized components of the water strider sperm flagellum. Antibodies specific for the mouse protease activated receptor PAR2 recognized an antigen on the sperm flagellum. These results suggest that trypsin stimulates a Ca(2+) and MAPK mediated signaling pathway and potentially implicate a PAR2-like protein in regulating motility.
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PMID:Protease activation and the signal transduction pathway regulating motility in sperm from the water strider Aquarius remigis. 2227 49


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