Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.24 (mitogen-activated protein kinase)
 95,810 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Both protein kinase C (PKC) and extracellular signal-regulated kinases (ERK1/2) are involved in mediating vascular smooth muscle contraction. We tested the hypotheses that in addition to PKC activation of ERK1/2, by negative feedback ERKs modulate PKC-induced contraction, and that their interactions modulate both thick and thin myofilament pathways. In ovine middle cerebral arteries (MCA), we measured isometric tension and intracellular free calcium concentration ([Ca(2+)](i)) responses to PKC stimulation [phorbol 12,13-dibutyrate (PDBu), 3 x 10(-6) M] in the absence or presence of ERK1/2 inhibition (U-0126, 10(-5) M). After PDBu +/- ERK1/2 inhibition, we also examined by Western immunoblot the levels of total and phosphorylated ERK1/2, caldesmon(Ser789), myosin light chain(20) (MLC(20)), and CPI-17. PDBu induced significant increase in tension in the absence of increased [Ca(2+)](i). PDBu also increased phosphorylated ERK1/2 levels, a response blocked by U-0126. In turn, U-0126 augmented PDBu-induced contractions. PDBu also was associated with significant increases in phosphorylated caldesmon(Ser789) and MLC(20) levels, each of which peaked at 5 to 10 min. PDBu also increased phosphorylated CPI-17 levels, which peaked at 2 to 3 min. Rho kinase inhibition (Y-27632, 3 x 10(-7) M) did not alter PDBu-induced contraction. These results support the idea that PKC activation can increase CPI-17 phosphorylation to decrease myosin light chain phosphatase activity. In turn, this increases MLC(20) phosphorylation in the thick filament pathway and increases Ca(2+) sensitivity. In addition, ERK1/2-dependent phosphorylation of caldesmon(Ser789) was not necessary for PDBu-induced contraction and appears not to be involved in the reversal of caldesmon's inhibitory effect on actin-myosin ATPase.
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PMID:PKC-induced ERK1/2 interactions and downstream effectors in ovine cerebral arteries. 1595 60

Vascular contraction is an important determinant of the peripheral vascular resistance and blood pressure. The mechanisms underlying vascular smooth muscle (VSM) contraction and the pathological changes that occur in hypertension have been the subject of numerous studies and interpretations. Activation of VSM by vasoconstrictor stimuli at the cell surface causes an increase in [Ca(2+)](i), Ca(2+)-dependent activation of myosin light chain (MLC) kinase, MLC phosphorylation, actin-myosin interaction and VSM contraction. Additional signaling pathways involving Rho-kinase and protein kinase C (PKC) may increase the myofilament force sensitivity to [Ca(2+)](i) and MLC phosphorylation, and thereby maintain vascular contraction. PKC is a particularly intriguing protein kinase as it comprises a family of Ca(2+)-dependent and Ca(2+)-independent isoforms, which have different tissue and subcellular distribution, and undergo differential translocation during cell activation. PKC translocation to the cell surface may trigger a cascade of protein kinases, such as mitogen-activated protein kinase (MAPK) and MAPK kinase (MEK) that ultimately interact with the contractile myofilaments and cause VSM contraction. Also, PKC translocation to the nucleus may promote VSM growth and proliferation. Increased PKC expression and activity have been identified in several forms of hypertension. The subcellular location of PKC may determine the state of VSM activity, and may be useful in the diagnosis/prognosis of hypertension. Vascular PKC isoforms may represent specific targets for modulation of VSM hyperactivity, and isoform-specific PKC inhibitors may be useful in treatment of Ca(2+) antagonist-resistant forms of hypertension.
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PMID:Protein kinase C isoforms as specific targets for modulation of vascular smooth muscle function in hypertension. 1613 52

Retinal pigment epithelial cells (RPEs) are thought to be one of the main components of fibrous membrane observed in eyes with proliferative vitreo-retinopathy. We investigated the signalling mechanisms of TGF-beta2-dependent collagen gel contraction by RPEs. An in vitro type I collagen gel contraction assay was performed to evaluate the effect of TGF-beta2 on gel contraction. The expression of alpha-smooth muscle actin (alpha-SMA) and the phosphorylation state of myosin light chain (MLC) were analyzed by Western blotting. The involvement of protein kinases such as p44/42 mitogen-activated protein kinase (MAPK), protein kinase C (PKC), p38 MAPK and phosphatidylinositol-3 kinase was investigated. The contribution of Rho-kinase and/or MLC-kinase was also evaluated using respective kinase inhibitors (Y27632, hydroxyfasudil and ML7). Additionally, RPEs were immunostained to examine whether the expression of alpha-SMA detected in our western blotting correlated to the stress fiber formation within the cells. TGF-beta2 caused time (0-5 days)-and dose (0 10 ng ml(-1))-dependent gel contraction associated with overexpression of alpha-SMA and phosphorylation of MLC (p < 0.01, respectively). PKC inhibitor (GF109203X, 5 microM) and p38 MAPK inhibitor (SB203580, 10 microM) significantly attenuated TGF-beta2-elicited gel contraction via partial downregulation of both alpha-SMA expression and MLC phosphorylation (p < 0.01, respectively). The gel contraction was prominently inhibited in the presence of Y27632 (10 microM) or hydroxyfasudil (10 microM) with strong suppression of MLC phosphorylation but had no significant effect on alpha-SMA expression. Treatment with ML7, in contrast, resulted in a marginal inhibition of MLC phosphorylation and gel contraction. Finally, pretreatment of the cells with Y27632 or hydroxyfasudil prevented the formation of stress fiber within the cells. These results indicate that TGF-beta2-dependent myofibroblastic transdifferentiation and MLC phosphorylation by RPEs involve both PKC and p38 MAPK pathways at least in part. Myofibroblastic transdifferentiation of RPEs appears to be independent of the Rho-kinase pathway, and the presence of alpha-SMA does not necessarily reflect the contractile potential of a cell. While Rho-kinase inhibitors are incapable of preventing myofibroblastic transdifferentiation itself, this pathway could be one of the critical targets of cell-mediated contraction of the tissue containing fibrillar collagens by transdifferentiated RPEs.
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PMID:Critical role of the Rho-kinase pathway in TGF-beta2-dependent collagen gel contraction by retinal pigment epithelial cells. 1631 Jan 90

We evaluated the hypothesis that mechanical deformation of airway smooth muscle induces structural remodeling of airway smooth muscle cells, thereby modulating mechanical performance in subsequent contractions. This hypothesis implied that past experience of mechanical deformation was retained (or "memorized") as structural changes in airway smooth muscle cells, which modulated the cell's subsequent contractile responses. We termed this phenomenon mechanical strain memory. Preshortening has been found to induce attenuation of both force and isotonic shortening velocity in cholinergic receptor-activated airway smooth muscle. Rapid stretching of cholinergic receptor-activated airway smooth muscle from an initial length to a final length resulted in post-stretch force and myosin light chain phosphorylation that correlated significantly with initial length. Thus post-stretch muscle strips appeared to retain memory of the initial length prior to rapid stretch (mechanical strain memory). Cytoskeletal recruitment of actin- and integrin-binding proteins and Erk 1/2 MAPK appeared to be important mechanisms of mechanical strain memory. Sinusoidal length oscillation led to force attenuation during oscillation and in subsequent contractions in intact airway smooth muscle, and p38 MAPK appeared to be an important mechanism. In contrast, application of local mechanical strain to cultured airway smooth muscle cells induced local actin polymerization and cytoskeletal stiffening. It is conceivable that deep inspiration-induced bronchoprotection may be a manifestation of mechanical strain memory such that mechanical deformation from past breathing cycles modulated the mechanical performance of airway smooth muscle in subsequent cycles in a continuous and dynamic manner.
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PMID:Mechanisms of mechanical strain memory in airway smooth muscle. 1633 51

The Min/+ mouse is a model for APC-dependent colorectal cancer (CRC). We showed that tumorigenesis in this animal was associated with decreased E-cadherin adhesion and increased epidermal growth factor receptor (Egfr) activity in the non-tumor intestinal mucosa. Here, we tested whether these abnormalities correlated with changes in the actin cytoskeleton due to increased Rho-ROCK signaling. We treated Apc+/+ (WT) littermate small intestine with EGTA, an inhibitor of E-cadherin, and with LPA, an RhoA activator; both induced effects on adhesion and kinase activity that mimicked the Min/+ phenotype. GTP-bound Rho was increased in Min/+ enterocytes relative to WT. Since RhoA activity is associated with actomyosin contractility, markers of this signaling cascade were assessed including phosphorylated myosin light chain (MLC), cofilin, Pyk2, Src, and MAPK kinases. The increased actomyosin contractility characterizing Min/+ intestinal tissue was suppressed by the ROCK inhibitor, Y27632, but was inducible in the WT by EGTA or LPA. Finally, ultrastructural imaging revealed changes consistent with actomyosin contractility in Min/+ enterocytes. Thus, the positive regulation of E-cadherin adhesion provided by Apc+ in vivo allows proper negative regulation of Egfr, Src, Pyk2, and MAPK, as well as RhoA activities.
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PMID:Deficient E-cadherin adhesion in C57BL/6J-Min/+ mice is associated with increased tyrosine kinase activity and RhoA-dependent actomyosin contractility. 1636 33

Sustained smooth-muscle contraction or its experimental counterpart, Ca2+ sensitization, by G(q/13)-coupled receptor agonists is mediated via RhoA-dependent inhibition of MLC (myosin light chain) phosphatase and MLC20 (20 kDa regulatory light chain of myosin II) phosphorylation by a Ca2+-independent MLCK (MLC kinase). The present study identified the corresponding pathways initiated by G(i)-coupled receptors. Somatostatin acting via G(i)1-coupled sstr3 receptor, DPDPE ([D-Pen2,D-Pen5]enkephalin; where Pen is penicillamine) acting via G(i)2-coupled delta-opioid receptors, and cyclopentyl adenosine acting via G(i)3-coupled adenosine A1 receptors preferentially activated PI3K (phosphoinositide 3-kinase) and ILK (integrin-linked kinase), whereas ACh (acetylcholine) acting via G(i)3-coupled M2 receptors preferentially activated PI3K, Cdc42 (cell division cycle 42)/Rac1, PAK1 (p21-activated kinase 1) and p38 MAPK (mitogen-activated protein kinase). Only agonists that activated ILK induced sustained CPI-17 (protein kinase C potentiated inhibitor 17 kDa protein) phosphorylation at Thr38, MLC20 phosphorylation at Ser19, and contraction, consistent with recent evidence that ILK can act as a Ca2+-independent MLCK capable of phosphorylating the MLC phosphatase inhibitor, CPI-17, at Thr38. ILK activity, and CPI-17 and MLC20 phosphorylation were inhibited by LY294002 and in muscle cells expressing ILK(R211A) or treated with siRNA (small interfering RNA) for ILK. ACh acting via M2 receptors activated ILK, and induced CPI-17 and MLC20 phosphorylation and muscle contraction, but only after inhibition of p38 MAPK; all these responses were inhibited in cells expressing ILK(R211A). Conversely, ACh activated PAK1, a step upstream of p38 MAPK, whereas the three other agonists did so only in cells transfected with ILK(R211A) or siRNA for ILK. The results demonstrate reciprocal inhibition between two pathways downstream of PI3K, with ILK inhibiting PAK1, and p38 MAPK inhibiting ILK. Sustained contraction via G(i)-coupled receptors is dependent on CPI-17 and MLC20 phosphorylation by ILK.
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PMID:Gi-coupled receptors mediate phosphorylation of CPI-17 and MLC20 via preferential activation of the PI3K/ILK pathway. 1647 57

The invasive properties of cancer cells depend on their intrinsic motile potential and on their ability to breach the endothelial barrier. In the present work, we investigated the mechanisms by which adhesion of colon cancer cells to E-selectin expressed by endothelial cells regulates the barrier function of these cells and modulates transmigration of cancer cells. We found that the stimulation of E-selectin by activating antibodies or the adhesion of HT-29 cells results in an increase in the activity of extracellular signal-regulated kinase (ERK) and p38 mitogen-activated protein kinases. In turn, the activation of p38 and ERK enhances transendothelial permeability and migration of HT-29 cells. We also obtained evidence suggesting that p38-mediated increase in transendothelial migration of cancer cells depends on a myosin light chain phosphorylation-mediated formation of stress fibres. On the other hand, the activation of ERK by E-selectin modulates the opening of interendothelial spaces by initiating the activation of Src kinase activities and the dissociation of the VE-cadherin/beta-catenin complex. We conclude that activation of E-selectin by adhering cancer cells is an important process that regulates the extravasation of colon cancer cells by initiating p38- and ERK-dependent mechanisms that both contribute to regulate the integrity of the endothelial layer.
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PMID:Regulation of transendothelial migration of colon cancer cells by E-selectin-mediated activation of p38 and ERK MAP kinases. 1671 42

We previously showed that hydrogen peroxide (H2O2) induced resistance artery relaxation independent of endothelium. Thus, in this study we investigated the mechanism of relaxation induced by H2O2 on human renal vascular smooth muscle cell (HVSMC). HVSMC were stimulated with H2O2 and/or angiotensin II (Ang II), proline-rich-tyrosine-kinase-2 (PYK2), ERK1/2 MAP-Kinase, and myosin light chain 20 phosphorylation (Lc20) were assessed using Western blot analysis in the presence of potassium channel blockers, MAP-Kinase, and nitric oxide synthesis (NOS) inhibitors. H2O2 increased PYK2 and ERK1/2 phosphorylation, and at the same time decreased Lc20 phosphorylation. AngII increased phosphorylation of PYK2, ERK1/2 and Lc20, whereas, the pretreatment of HVSMC with H2O2 decreased Lc20 phosphorylation induced by AngII. MEK inhibition, decreased ERK1/2 phosphorylation, but had no effect on the inhibition of phosphorylation of Lc20 induced by H2O2. The inhibition of Ca2(+)-dependent K+ channels (BKCa) and NOS did not block the decrease of Lc20 phosphorylation in response to H2O2. On the other hand, pretreatment of HVSMC with 60 mM of KCl, increased rather than decreased Lc20 phosphorylation in response to H2O2. This study shows the evidence that H2O2 acts as a relaxing factor and as an activator of PYK2 and ERK1/2 in Human renal VSMC. The relaxation induced by H2O2 is independent of BKCa, ERK1/2 MAP-Kinase and NOS pathways. The relaxing effect to H2O2 changes to contracting effect when the potassium channels are compromised.
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PMID:Hydrogen peroxide acts as relaxing factor in human vascular smooth muscle cells independent of map-kinase and nitric oxide. 1672 Mar 30

The effect of granulocyte colony-stimulating factor (G-CSF) on human neutrophil motility was studied using videomicroscopy. Stimulation of neutrophils with G-CSF resulted in enhanced motility with morphological change and increased adherence. Enhanced neutrophil motility was detected within 3-5 min after G-CSF stimulation, reached a maximum at 10 min, and was sustained for approximately 35 min. The maximum migration rate was 84.4 +/- 2.9 microm/5 min. A study using the Boyden chamber method revealed that G-CSF-stimulated neutrophils exhibited random migration but not chemotaxis. Enhanced neutrophil motility and morphological change were inhibited by MEK [mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) kinase] inhibitors (PD98059 and U0126), and a phosphatidylinositol 3-kinase (PI3K) inhibitor (wortmannin), but not by a p38 MAPK inhibitor (SB203580). These findings are consistent with the fact that G-CSF selectively activates MEK/ERK and PI3K, but not p38, in neutrophils. MEK/ERK activation was associated with G-CSF-induced redistribution of F-actin and phosphorylated myosin light chain. Enhanced neutrophil motility was observed even in the presence of neutralizing anti-CD18 antibody, which prevented cell adherence. These findings indicate that G-CSF induces human neutrophil migration via activation of MEK/ERK and PI3K.
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PMID:Enhanced neutrophil motility by granulocyte colony-stimulating factor: the role of extracellular signal-regulated kinase and phosphatidylinositol 3-kinase. 1690 68

Recent studies in Cdc42 knockout mouse embryonic stem (ES) cells and ES-derived fibroblastoid cell lines raise concern on a body of literature derived by dominant mutant expression approach in a variety of cell lines implicating mammalian Cdc42 as a key regulator of filopodia induction, directional migration and cell cycle progression. To resolve the physiological function of mammalian Cdc42, we have characterized the Cdc42(-/-) and Cdc42GAP(-/-) primary mouse embryonic fibroblasts (MEFs) produced by gene targeting as the Cdc42 loss- or gain-of-activity cell model. The Cdc42(-/-) cells were defective in filopodia formation stimulated by bradykinin and in dorsal membrane ruffling stimulated by PDGF, whereas the Cdc42GAP(-/-) cells displayed spontaneous filopodia. The Cdc42 loss- or gain-of-activity cells were defective in adhesion to fibronectin, wound-healing, polarity establishment, and migration toward a serum gradient. These defects were associated with deficiencies of PAK1, GSK3beta, myosin light chain, and FAK phosphorylation. Furthermore, Cdc42(-/-) cells were defective in G1/S-phase transition and survival, correlating with deficient NF-kappaB transcription and defective JNK, p70 S6K, and ERK1/2 activation. These results demonstrate a different requirement of Cdc42 activity in primary MEFs from ES or ES-derived clonal fibroblastoid cells and suggest that Cdc42 plays cell-type-specific signaling roles.
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PMID:Gene targeting of Cdc42 and Cdc42GAP affirms the critical involvement of Cdc42 in filopodia induction, directed migration, and proliferation in primary mouse embryonic fibroblasts. 1691 16


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