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

The effects of epidermal growth factor (EGF) on intracellular calcium ([Ca(2+)](i)) responses to the muscarinic agonist carbachol were studied in a human salivary cell line (HSY). Carbachol (10(-4) M)-stimulated [Ca(2+)](i) mobilization was inhibited by 40% after 48-h treatment with 5 x 10(-10) M EGF. EGF also reduced carbachol-induced [Ca(2+)](i) in Ca(2+)-free medium and Ca(2+) influx following repletion of extracellular Ca(2+). Under Ca(2+)-free conditions, thapsigargin, an inhibitor of Ca(2+) uptake to internal stores, induced similar [Ca(2+)](i) signals in control and EGF-treated cells, indicating that internal Ca(2+) stores were unaffected by EGF; however, in cells exposed to thapsigargin, Ca(2+) influx following Ca(2+) repletion was reduced by EGF. Muscarinic receptor density, assessed by binding of the muscarinic receptor antagonist L-[benzilic-4,4'-(3)HCN]quinuclidinyl benzilate ([(3)H]QNB), was decreased by 20% after EGF treatment. Inhibition of the carbachol response by EGF was not altered by phorbol ester-induced downregulation of protein kinase C (PKC) but was enhanced upon PKC activation by a diacylglycerol analog. Phosphorylation of mitogen-activated protein kinase (MAP kinase) and inhibition of the carbachol response by EGF were both blocked by the MAP kinase pathway inhibitor PD-98059. The results suggest that EGF decreases carbachol-induced Ca(2+) release from internal stores and also exerts a direct inhibitory action on Ca(2+) influx. A decline in muscarinic receptor density may contribute to EGF inhibition of carbachol responsiveness. The inhibitory effect of EGF is mediated by the MAP kinase pathway and is potentiated by a distinct modulatory cascade involving activation of PKC. EGF may play a physiological role in regulating muscarinic receptor-stimulated salivary secretion.
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PMID:EGF inhibits muscarinic receptor-mediated calcium signaling in a human salivary cell line. 1100 83

We have previously shown that m1 and m2 muscarinic receptors were expressed on human peripheral blood lymphocytes (hPBL) and that pre-stimulation of these receptors enhanced phytohemagglutinin (PHA)-induced interleukin-2 (IL-2) production. Possible intracellular signal pathways of muscarinic receptors to regulate IL-2 production were examined in human T cell line Jurkat cells. Pretreatment of the cells with muscarinic receptor agonist, oxotremorine M (Oxo-M), enhanced IL-2 production induced by phorbol 12-myristate 13-acetate (PMA)/A23187, while Oxo-M by itself did not affect IL-2 production. The enhancement of IL-2 production by Oxo-M was inhibited by 4-diphenylacetoxy-N-methylpiperidine methiodide (4-DAMP) an ml/m3 receptor antagonist. When the cells were pretreated with AF-DX116, an m2 antagonist, the IL-2 production enhanced by Oxo-M was further stimulated. Reverse transcription-polymerase chain reaction (RT-PCR) revealed that ml and m2 muscarinic receptors exist on Jurkat cells. The stimulation of ml receptors enhanced the PMA/A23187-induced binding activity to AP-1 consensus sequences in IL-2 promoter and production of c-Fos and c-Jun protein. The stimulation of ml receptors did not modify the DNA binding of NF-kappaB, NF-AT or Oct-1. When ml receptors were stimulated, activities of mitogen-activated protein kinase (MAPK)/extracellular signal regulated protein kinase (ERK) and c-Jun N-terminal kinase (JNK) were increased, while p38 MAPK was not affected. Incubation with Oxo-M induced a transient increase in [Ca2+]i, which was abolished by pretreatment with 4-DAMP. Treatment with cyclosporin A markedly decreased the PMA/A23187-induced IL-2 promoter activity. This treatment, however, did not affect the enhancement of the promoter activity induced by ml receptor stimulation. The results suggest that transcription factor AP-1 is involved in the ml receptor-mediated enhancement of IL-2 transcript in Jurkat cells, and that pathways via MAPK/ERK and JNK, but not via p38 MAPK, are involved in the ml receptor-mediated enhancement of IL-2 promoter activity.
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PMID:Extracellular signal regulated protein kinase and c-jun N-terminal kinase are involved in ml muscarinic receptor-enhanced interleukin-2 production pathway in Jurkat cells. 1104 Dec 51

When PC12 cells are deprived of trophic support they undergo apoptosis. We have previously shown that survival of trophic factor-deprived PC12M1 cells can be promoted by activation of the G protein-coupled muscarinic receptors. The mechanism whereby muscarinic receptors inhibit apoptosis is poorly understood. In the present study we investigated this mechanism by examining the effect of muscarinic receptor activation on the serum deprivation-induced activity of key players in apoptosis, the caspases, in PC12M1 cells. The results showed that m1 muscarinic activation inhibits caspase activity induced by serum deprivation. This effect appeared to be caused by the prevention of activation of caspases such as caspase-2 and caspase-3, and not by the inhibition of existing activity. Muscarinic receptor activation also stimulated the mitogen-activated protein kinase/extracellular signaling-regulated kinase (MAPK/ERK) and phosphoinositide (PI) 3-kinase signaling pathways. The PI 3-kinase pathway inhibitors wortmannin and LY294002, as well as the MAPK/ERK pathway PD98059 inhibitor, did not however suppress the inhibitory effect of the muscarinic receptors on caspase activity. The results therefore suggested that the muscarinic survival effect is mediated by a pathway that leads to caspase inhibition by MAPK/ERK- and PI 3-kinase-independent signaling cascades.
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PMID:M1 muscarinic receptors block caspase activation by phosphoinositide 3-kinase- and MAPK/ERK-independent pathways. 1104 77

We investigated the role played by agonist-mediated phosphorylation of the G(q/11)-coupled M(3)-muscarinic receptor in the mechanism of activation of the mitogen-activated protein kinase pathway, ERK-1/2, in transfected Chinese hamster ovary cells. A mutant of the M(3)-muscarinic receptor, where residues Lys(370)-Ser(425) of the third intracellular loop had been deleted, showed a reduced ability to activate the ERK-1/2 pathway. This reduction was evident despite the fact that the receptor was able to couple efficiently to the phospholipase C second messenger pathway. Importantly, the ERK-1/2 responses to both the wild-type M(3)-muscarinic receptor and DeltaLys(370)-Ser(425) receptor mutant were dependent on the activity of protein kinase C. Our results, therefore, indicate the existence of two mechanistic components to the ERK-1/2 response, which appear to act in concert. First, the activation of protein kinase C through the diacylglycerol arm of the phospholipase C signaling pathway and a second component, absent in the DeltaLys(370)-Ser(425) receptor mutant, that is independent of the phospholipase C signaling pathway. The reduced ability of the DeltaLys(370)-Ser(425) receptor mutant to activate the ERK-1/2 pathway correlated with an approximately 80% decrease in the ability of the receptor to undergo agonist-mediated phosphorylation. Furthermore, we have previously shown that M(3)-muscarinic receptor phosphorylation can be inhibited by a dominant negative mutant of casein kinase 1alpha and by expression of a peptide corresponding to the third intracellular loop of the M(3)-muscarinic receptor. Expression of these inhibitors of receptor phosphorylation reduced the wild-type M(3)-muscarinic receptor ERK-1/2 response. We conclude that phosphorylation of the M(3)-muscarinic receptor on sites in the third intracellular loop by casein kinase 1alpha contributes to the mechanism of receptor activation of ERK-1/2 by working in concert with the diacylglycerol/PKC arm of the phospholipase C signaling pathway.
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PMID:Phosphorylation of the Gq/11-coupled m3-muscarinic receptor is involved in receptor activation of the ERK-1/2 mitogen-activated protein kinase pathway. 1108 74

Receptors for many neurotransmitters including catecholamines and acetylcholine (ACh) have been detected on the cell surface of lymphocytes. It has been demonstrated that a human T cell line synthesizes ACh and suggested that ACh may be an autacoid modulating T cell-dependent immune responses. However, the biochemical interactions of the ACh system with the immune system have not been elucidated in detail. We have shown that m1 and m2 muscarinic receptor mRNAs are expressed in human peripheral blood lymphocytes and in human T cell line Jurkat cells and that pretreatment of these cells with a muscarinic receptor agonist enhances interleukin-2 (IL-2) production. We also postulated possible intracellular signaling pathways via which muscarinic receptors regulate IL-2 production in Jurkat cells. The findings suggest that M1 muscarinic receptors are involved in muscarinic receptor-mediated enhancement of IL-2 production in Jurkat cells and that the transcription factor AP-1 and pathways via mitogen-activated protein kinase (MAPK)/extracellular signal regulated protein kinase and c-Jun N-terminal kinase, but not via p38 MAPK, may be involved in the muscarinic receptor-mediated enhancement of IL-2 production. Our findings demonstrate a neuro-immune interaction through muscarinic receptor signaling in immune cells.
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PMID:Roles of muscarinic acetylcholine receptors in interleukin-2 synthesis in lymphocytes. 1124 67

Activation of the m1 muscarinic receptor subtype in rat pheochromocytoma (PC12) cells stably expressing cloned m1 muscarinic acetylcholine receptors was previously shown to induce morphological changes and growth arrest. However, the signaling pathways which lead to these effects were not identified. In an attempt to characterize the intracellular signaling that might be involved in the muscarinic-induced effects, we investigated the role of reactive oxygen species in the regulation of these processes. Stimulation of the muscarinic receptor in these cells increased the intracellular concentrations of reactive oxygen species. Muscarinic activation induced intracellular signaling pathways that involve activation of Ras, extracellular signal-regulated kinase (ERK), and p38. These pathways were partially blocked when reactive oxygen species (ROS) production was prevented by the antioxidant N-acetylcysteine. Other muscarinic-induced signals, such as activation of c-Jun NH(2)-terminal kinase (JNK) or an increase in the binding activity of the transcription factors nuclear factor-kappa B and activator protein-1, were inhibited by the antioxidant dicoumarol. N-Acetylcysteine also blocked the growth arrest and changes in cell shape induced by stimulation of the muscarinic receptor in PC12M1 cells. These findings suggest that ROS act as second messengers in muscarinic-induced cellular signaling. Moreover, generation of ROS appears to be an early and critical intermediary event, which occurs immediately after stimulation of the muscarinic receptor and affects in a variety of mechanisms the muscarinic-mediated cellular signaling.
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PMID:Reactive oxygen species regulate signaling pathways induced by M1 muscarinic receptors in PC12M1 cells. 1125 88

Cholinergic agents are known to affect the epithelial transport of H2O and electrolytes in the kidney. In proximal tubule cells, cholinergic agonists increase basolateral Na-HCO(3) cotransport activity via M(1) muscarinic receptor activation. The signaling intermediates that couple these G protein-coupled receptors to cotransporter activation, however, are not well defined. We therefore sought to identify distal effectors of muscarinic receptor activation that contribute to increased NBC activity in cultured proximal tubule cells. As demonstrated previously for acute CO2-regulated cotransport activity, we found that inhibitors of Src family kinases (SFKs) or the classic mitogen-activated protein kinase (MAPK) pathway prevented the stimulation of NBC activity by carbachol. The ability of carbachol to activate Src, as well as the proximal (Raf) and distal [extracellular signal-regulated kinases 1 and 2 (ERK1/2)] elements of the classic MAPK module, was compatible with these findings. Cholinergic stimulation of ERK1/2 activity was also completely prevented by overexpression of a dominant negative mutant of Ras (N17-Ras). Taken together, these findings suggest a requirement for the sequential activation of SFKs, Ras, and the classic MAPK pathway [Raf-->MAPK/ERK kinase (MEK)-->ERK]. These findings provide important insights into the molecular mechanisms underlying cholinergic regulation of NBC activity in renal epithelial cells. They also suggest a specific mechanism whereby cholinergic stimulation of the kidney can contribute to pH homeostasis.
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PMID:SFKs, Ras, and the classic MAPK pathway couple muscarinic receptor activation to increased Na-HCO(3) cotransport activity in renal epithelial cells. 1129 27

Galpha(i)-coupled receptor stimulation results in epidermal growth factor receptor (EGFR) phosphorylation and MAPK activation. Regulators of G protein signaling (RGS proteins) inhibit G protein-dependent signal transduction by accelerating Galpha(i) GTP hydrolysis, shortening the duration of G protein effector stimulation. RGS16 contains two conserved tyrosine residues in the RGS box, Tyr(168) and Tyr(177), which are predicted sites of phosphorylation. RGS16 underwent phosphorylation in response to m2 muscarinic receptor or EGFR stimulation in HEK 293T or COS-7 cells, which required EGFR kinase activity. Mutational analysis suggested that RGS16 was phosphorylated on both tyrosine residues (Tyr(168) Tyr(177)) after EGF stimulation. RGS16 co-immunoprecipitated with EGFR, and the interaction did not require EGFR activation. Purified EGFR phosphorylated only recombinant RGS16 wild-type or Y177F in vitro, implying that EGFR-mediated phosphorylation depended on residue Tyr(168). Phosphorylated RGS16 demonstrated enhanced GTPase accelerating (GAP) activity on Galpha(i). Mutation of Tyr(168) to phenylalanine resulted in a 30% diminution in RGS16 GAP activity but completely eliminated its ability to regulate G(i)-mediated MAPK activation or adenylyl cyclase inhibition in HEK 293T cells. In contrast, mutation of Tyr(177) to phenylalanine had no effect on RGS16 GAP activity but also abolished its regulation of G(i)-mediated signal transduction in these cells. These data suggest that tyrosine phosphorylation regulates RGS16 function and that EGFR may potentially inhibit Galpha(i)-dependent MAPK activation in a feedback loop by enhancing RGS16 activity through tyrosine phosphorylation.
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PMID:RGS16 function is regulated by epidermal growth factor receptor-mediated tyrosine phosphorylation. 1160 4

This study was designed to characterise the muscarinic receptor subtype responsible for acetylcholine-mediated in vitro pulmonary artery relaxation in rats and the importance of the presence of neostigmine (an anti-cholinesterase) during receptor characterisation. Cumulative administration of acetylcholine elicited concentration-dependent relaxation of phenylephrine (1 microM) precontracted preparations. Inclusion of neostigmine (10 microM) caused a parallel leftward shift with an increase of the pD(2) value (7.09 vs. 6.43) of the concentration-response curve of acetylcholine. The magnitude of maximum relaxation, however, was not affected. Using a range of conventional muscarinic receptor antagonists (atropine, pirenzepine, methoctramine, p-FHHSiD and tropicamide) and the highly selective Green Mamba muscarinic toxins (MT-3 and MT-7), it was found that muscarinic M(3) receptors are probably responsible for endothelium-dependent relaxation of the pulmonary artery upon acetylcholine challenge. Preincubation with N(G)-nitro-L-arginine methyl ester (L-NAME, 20 microM, a nitric oxide synthase inhibitor), but not N(G)-nitro-D-arginine methyl ester (D-NAME, 20 microM), abolished acetylcholine-elicited relaxation. Moreover, 6-anilino-5,8-quinolinedione (LY 83583, 1 microM) and methylene blue (1 microM) (both are guanylate cyclase inhibitors) markedly attenuated acetylcholine-elicited relaxation. However, the presence of indomethacin (3 microM, a cyclo-oxygenase inhibitor), (-)-perillic acid (30 microM, a p21(ras) blocker), 2-[2'-amino-3'-methoxy-phenyl]-oxana-phthalen-4-one (PD 98059) (10 microM, a p42/p44 mitogen-activated protein kinase inhibitor), 4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)1H-imidazole (SB 203580) (1 microM, a p38 mitogen-activated protein kinase blocker), wortmannin (500 nM, a phosphatidylinositol-3 kinase inhibitor) and genistein (10 microM, a tyrosine kinase blocker) failed to alter acetylcholine-provoked pulmonary arterial relaxation. These results suggest that acetylcholine caused pulmonary arterial relaxation through the activation of muscarinic M(3) receptors in the endothelium. Moreover, the p21(ras)/mitogen-activated protein kinase pathway seems to play no role in mediating acetylcholine-elicited relaxation.
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PMID:Role of mitogen-activated protein kinase pathway in acetylcholine-mediated in vitro relaxation of rat pulmonary artery. 1175 66

IK(SO) is a standing-outward potassium current found in cerebellar granule neurons which is inhibited by the activation of muscarinic M(3) receptors. However the pathway between muscarinic receptor activation and current inhibition is unknown. Using two structurally distinct inhibitors of the activation of MEK1 (mitogen activated protein (MAP) kinase kinase 1), PD 98059 and U 0126, we have shown that the MAP kinase signalling cascade does not appear to underlie muscarinic inhibition of IK(SO), recorded using whole-cell patch-clamp methods. Nevertheless, both PD 98059 and U 0126 caused an inhibition of IK(SO) when applied acutely with 30 microM of each compound producing around 50% inhibition of the current. In addition, U 0125, which is structurally related to U 0126 but has a much lower potency for inhibiting MEK1 activation, was also able to inhibit IK(SO) to a similar degree. Neither the inhibition by PD 98059 nor that by U 0126 was found to be voltage dependent. This was true whether the IK(SO) current was outward or inward. Block of IK(SO) by these two compounds may compromise interpretation of studies in intact neuronal preparations when they are used as MEK1 inhibitors.
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PMID:Inhibition of the potassium current IK(SO), in cerebellar granule cells, by the inhibitors of MEK1 activation, PD 98059 and U 0126. 1180 18


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