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:3.1.4.3 (phospholipase C)
18,461 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Arachidonic acid activates isolated Rho-kinase and contracts permeabilized smooth muscle fibres. Various assays were carried out to examine the mechanism of this activation. Native Rho-kinase was activated 5-6 times by arachidonic acid but an N-terminal, constitutively-active fragment of Rho-kinase, expressed as a glutathione-S-transferase (GST) fusion protein and including the catalytic subunit (GST-Rho-kinase-CAT), was not. GST-Rho-kinase-CAT was inhibited by a C-terminal fragment of Rho-kinase and arachidonic acid removed this inhibition. These results suggest that the C-terminal part of Rho-kinase, containing the RhoA binding site and the pleckstrin homology domain, acts as an autoinhibitor. It is suggested further that activation by arachidonic acid is due to its binding to the autoinhibitory region and subsequent release from the catalytic site. Arachidonic acid, at concentrations greater than 30 microM, increases force in alpha-toxin-permeabilized femoral artery but not in Triton X-100-skinned fibres. The content of Rho-kinase in the latter was lower than in alpha-toxin-treated or intact fibres. The arachidonic acid-induced contraction was not observed at a pCa above 8.0 and was inhibited by Y-27632 and wortmannin, inhibitors of Rho-kinase and myosin light-chain kinase (MLCK), respectively. The activation of Rho-kinase and subsequent phosphorylation of the myosin phosphatase target subunit inhibits myosin phosphatase and increases myosin phosphorylation.
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PMID:Arachidonic acid-induced Ca2+ sensitization of smooth muscle contraction through activation of Rho-kinase. 1129 40

This article provides an update of a minireview published in 1996 (Abdel-Latif AA. Proc Soc Exp Biol Med 211:163-177, 1996), the purpose of which was to examine in nonvascular smooth muscle the biochemical and functional cross talk between the sympathetic nervous system, which governs the formation of cAMP and muscle relaxation, and the parasympathetic nervous system, which governs the generation of IP3 and diacylglycerol, from the polyphosphoinositides, Ca2+ mobilization, and contraction. This review examines further evidence, both from nonvascular and vascular smooth muscle, for cross talk between the cyclic nucleotides, cAMP and cGMP via their respective protein kinases, and the Ca2+-dependent- and Ca2+-independent-signaling pathways involved in agonist-induced contraction. These include the IP3-Ca2+-CaM- myosin light chain kinase (MLCK) pathway and the Ca2+-independent pathways, including protein kinase C-, MAP kinase-, and Rho-kinase. In addition, MLC phosphorylation and contraction can also be increased by a decrease in myosin phosphatase activity. A summary of the cross talk between the cyclic nucleotides and these signaling pathways was presented. In smooth muscle, there are several targets for cyclic nucleotide inhibition and consequent relaxation, including the receptor, G proteins, phospholipase C-beta1-4 isoforms, IP3 receptor, Ca2+ mobilization, MLCK, MAP kinase, Rho-kinase, and myosin phosphatase. While significant progress has been made in the past four years on this cross talk, the precise mechanisms underlying the biochemical basis for the cyclic nucleotide inhibition of Ca2+ mobilization and consequently muscle contraction remain to be established. Although it is well established that second-messenger cross talk plays an important role in smooth muscle relaxation, the many sources which exist in smooth muscle for Ca2+ mobilization, coupled with the multiple signaling pathways involved in agonist-induced contraction, contribute appreciably to the difficulties found by many investigators in identifying the targets for cyclic nucleotide inhibition and consequent relaxation. Better methodology and more novel interdisciplinary approaches are required for elucidating the mechanism(s) of cAMP- and cGMP-inhibition of smooth muscle contraction.
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PMID:Cross talk between cyclic nucleotides and polyphosphoinositide hydrolysis, protein kinases, and contraction in smooth muscle. 1136 Oct 33

Thrombin and proteinase-activated receptors (PAR) specifically regulate several functions that markedly enhance the transformation phenotype such as inflammation, cell proliferation, tumor growth, and metastasis. We recently reported that thrombin inhibits cellular invasion induced by src, hepatocyte growth factor (HGF), and leptin in kidney and colonic epithelial cells via predominant activation of the pertussis toxin (PTx) -sensitive G-proteins Galphao/Galphai. We provide pharmacological and biochemical evidence that in the presence of PTx, PAR-1 induced cellular invasion through Galpha12/Galpha13- and RhoA/Rho kinase (ROCK) -dependent signaling. However, inhibition of the endogenous small GTPase RhoA by the C3 exoenzyme, dominant-negative N19-RhoA, activated G26V-RhoD, and activators of the nitric oxide/cGMP pathways conferred invasive activity to PAR-1 via a signaling cascade using Galphaq, phospholipase C (PLC), Ca(2+)/calmodulin myosin light chain kinase (CaM-MLCK), and phosphorylation of MLC. We found that cellular invasion induced by the src oncogene is abrogated by inhibitors of the RhoA/ROCK pathway and is independent of PLC/CaM-MLCK signaling. Our data demonstrate that the RhoA and RhoD small GTPases are acting as a molecular switch of cellular invasion and reveal a novel critical mechanism by which PAR-1 bypass Galphao/i and RhoA inhibition via differential coupling to heterotrimeric G-proteins linked to divergent or convergent biological responses. Our data also indicate that Rho GTPases and ROCK mediate a src-dependent invasion signal in kidney and colonic cancer cells. We conclude that dynamic regulation of Rho GTPases activation and inactivation by oncogenes, growth factors, cGMP-inducing agents, and adhesion molecules can initiate convergent invasion signals controlled by the thrombin PAR-1 in cancer cells.-Nguyen, Q.-D., Faivre, S., Bruyneel, E., Rivat, C., Seto, M., Endo, T., Mareel, M., Emami, S., Gespach, C. RhoA- and RhoD-dependent regulatory switch of Galpha subunit signaling by PAR-1 receptors in cellular invasion.
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PMID:RhoA- and RhoD-dependent regulatory switch of Galpha subunit signaling by PAR-1 receptors in cellular invasion. 1191 59

Respiratory tract infections caused by Bordetella pertussis are occasionally accompanied by severe neurologic disorders and encephalopathies. For these sequelae to occur the integrity of cerebral barriers needs to be compromised. The influence of pertussis toxin, a decisive virulence factor in the pathogenesis of pertussis disease, on barrier integrity was investigated in model systems for blood-liquor (epithelial) and blood-brain (endothelial) barriers. While pertussis toxin did not influence the barrier function in Plexus chorioideus model systems, the integrity of cerebral endothelial monolayers was severely compromised. Cellular intoxication by pertussis toxin proceeds via ADP-ribosylation of alpha-G(i) proteins, which not only interferes with the homeostatic inhibitory regulation of adenylate cyclase stimulation but also results in a modulation of the membrane receptor coupling. Increasing intra-endothelial cAMP levels by employing cholera toxin or forskolin even inhibited the pertussis toxin-induced permeabilization of endothelial barriers. Therefore, pertussis-toxin-induced permeabilization has to be mediated via a cAMP-independent pathway. To investigate potential signalling pathways we employed several well established cellular drugs activating or inhibiting central effectors of signal transduction pathways, such as phosphatidylinositol 3-kinase, adenylate cyclase, phospholipase C, myosin light chain kinase and protein kinase C. Only inhibitors and activators of protein kinase C and phosphatidylinositol 3-kinase affected the pertussis toxin-induced permeability. In summary, we conclude that permeabilization of cerebral endothelial monolayers by pertussis toxin does not depend on elevated cAMP levels and proceeds via the phosphokinase C pathway.
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PMID:Permeabilization in a cerebral endothelial barrier model by pertussis toxin involves the PKC effector pathway and is abolished by elevated levels of cAMP. 1266 64

Signalling via m3 and m2 receptors in smooth muscles involved activation of two G-protein-dependent pathways by each receptor. m2 receptors were coupled via Gbetagammai3 with activation of phospholipase C-beta3, phosphoinositide 3-kinase and Cdc42/Rac1 (where Cdc stands for cell division cycle) and p21-activated kinase 1 (PAK1), resulting in phosphorylation and inactivation of myosin light chain kinase (MLCK). Each step was inhibited by methoctramine and pertussis toxin. PAK1 activity was abolished in cells expressing both Cdc42-DN (where DN stands for dominant negative) and Rac1-DN. MLCK phosphorylation was inhibited by PAK1 antibody, and in cells expressing Cdc42-DN and Rac1-DN. m3 receptors were coupled via Galpha(q/11) with activation of phospholipase C-beta1 and via RhoA with activation of Rho-associated kinase (Rho kinase), phospholipase D and protein kinase C (PKC). Rho kinase and phospholipase D activities were inhibited by C3 exoenzyme and in cells expressing RhoA-DN. PKC activity was inhibited by bisindolylmaleimide, and in cells expressing RhoA-DN; PKC activity was also inhibited partly by Y27632 (44+/-5%). PKC-induced phosphorylation of PKC-activated 17 kDa inhibitor protein of type 1 phosphatase (CPI-17) at Thr38 was abolished by bisindolylmaleimide and inhibited partly by Y27632 (28+/-3%). Rho-kinase-induced phosphorylation of myosin phosphatase targeting subunit (MYPT1) and was abolished by Y27632. Sustained phosphorylation of 20 kDa regulatory light chain of myosin II (MLC20) and contraction were abolished by bisindolylmaleimide Y27632 and C3 exoenzyme and in cells expressing RhoA-DN. The results suggest that Rho-kinase-dependent phosphorylation of MYPT1 and PKC-dependent phosphorylation and enhancement of CPI-17 binding to the catalytic subunit of MLC phosphatase (MLCP) act co-operatively to inhibit MLCP activity, leading to sustained stimulation of MLC20 phosphorylation and contraction. Because Y27632 inhibited both Rho kinase and PKC activities, it could not be used to ascertain the contribution of MYPT1 to inhibition of MLCP activity. m2-dependent phosphorylation and inactivation of MLCK precluded its involvement in sustained MLC20 phosphorylation and contraction.
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PMID:Differential signalling by muscarinic receptors in smooth muscle: m2-mediated inactivation of myosin light chain kinase via Gi3, Cdc42/Rac1 and p21-activated kinase 1 pathway, and m3-mediated MLC20 (20 kDa regulatory light chain of myosin II) phosphorylation via Rho-associated kinase/myosin phosphatase targeting subunit 1 and protein kinase C/CPI-17 pathway. 1273 88

The mechanism of labour is not fully understood and further research into this important physiological process is needed. In some species, notably sheep, parturition is due to activation of the fetal hypothalamic-pituitary-adrenal axis. However, in primates, this axis appears to have a supportive, rather than essential role. Successful parturition requires an increase in coordinated uterine contractility together with changes in connective tissue that allow cervical ripening and dilatation. In most mammals, however, these changes are synchronised by a fall in maternal progesterone levels and a rise in oestrogens. This is not the case in women in whom the onset of labour occurs without apparent changes in circulating steroid levels. The basis of uterine contractility is the interaction between actin and myosin in myometrial smooth muscle cells. This is driven by calcium through Ca(2+)-calmodulin-dependent myosin light chain kinase (MLCK) activity. Moreover, calcium sensitisation occurs via activation of Rho kinase, a calcium-independent pathway that promotes contractility by inhibiting myosin phosphatase and probably by phosphorylating myosin on the same site as MLCK. Uterine activity can be modulated by many G-protein coupled receptors (GPCRs). For example, receptors coupled to Galpha(q) (oxytocin-, prostanoid FP and TP, endothelin-receptors) stimulate contractility by activating the phospholipase C/Ca(2+) pathway; receptors coupled to Galpha(s) (beta(2)-adrenoceptors, prostanoid EP2 and IP, some 5-hydroxytryptamine receptors e.g. 5-HT(7)) relax the uterus by increasing myometrial cyclic AMP levels; and receptors coupled to Galpha(i) (alpha(2)-adrenoceptors, muscarinic, 5-HT(1)) potentiate contractility, probably by inhibiting cAMP production. Because of its relative abundance in pregnant uterine tissue, the oxytocin receptor is an obvious target for tocolytic therapy. Oxytocin antagonists have been introduced into clinical practice for the management of preterm labour and offer the advantage of uterine selectivity and fewer side effects than conventional beta-agonist therapy.
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PMID:Mechanisms of labour--biochemical aspects. 1276 10

Various neurotransmitters excite neurons by suppressing a ubiquitous, voltage-dependent, noninactivating K+ conductance called the M-conductance (gM). In bullfrog sympathetic ganglion neurons the suppression of gM by the P2Y agonist ATP involves phospholipase C (PLC). The present results are consistent with the involvement of the lipid and inositol phosphate cycles in the effects of both P2Y and muscarinic cholinergic agonists on gM. Impairment of resynthesis of phosphatidylinositol 4,5-bisphosphate (PIP2) with the phosphatidylinositol 4-kinase inhibitor wortmannin (10 microm) slowed or blocked the recovery of agonist-induced gM suppression. This effect could not be attributed to an action of wortmannin on myosin light chain kinase or on phosphatidylinositol 3-kinase. Inhibition of PIP2 synthesis at an earlier point in the lipid cycle by the use of R59022 (40 microm) to inhibit diacylglycerol kinase also slowed the rate of recovery of successive ATP responses. This effect required several applications of agonist to deplete levels of various phospholipid intermediates in the lipid cycle. PIP2 antibodies attenuated the suppression of gM by agonists. Intracellular application of 20 microm PIP2 slowed the rundown of KCNQ2/3 currents expressed in COS-1 or tsA-201 cells, and 100 microm PIP2 produced a small potentiation of native M-current bullfrog sympathetic neurons. These are the results that might be expected if agonist-induced activation of PLC and the concomitant depletion of PIP2 contribute to the excitatory action of neurotransmitters that suppress gM.
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PMID:Experiments to test the role of phosphatidylinositol 4,5-bisphosphate in neurotransmitter-induced M-channel closure in bullfrog sympathetic neurons. 1283 15

Interleukin-1beta (IL-1beta) is a proinflammatory cytokine that plays a central role in inflammatory bowel disease (IBD). In order to elucidate the mechanism of motility disorders frequently observed in IBD, we investigated the long term effects of IL-1beta on rat ileal smooth muscle contractility by using an organ culture system. When ileal smooth muscle strips were cultured with IL-1beta (10 ng/ml), contractions elicited by high K+ and carbachol were inhibited in a time-dependent manner. IL-1beta more strongly inhibited the carbachol-induced contractions than high K+ with decreasing myosin light chain phosphorylation. In the alpha-toxin-permeabilized ileal muscle, carbachol with GTP or guanosine 5'-3-O-(thio)triphosphate increased the Ca2+ sensitivity of contractile elements, and this G protein-coupled Ca2+ sensitization was significantly reduced in the IL-1beta-treated ileum. Among the functional proteins involved in the smooth muscle Ca2+ sensitization, CPI-17 expression was significantly reduced after the culture with IL-1beta, whereas the expressions of RhoA, ROCK-I, ROCK-II, MYPT-1, myosin light chain kinase, and myosin phosphatase (PP1) were unchanged. The phosphorylation level of CPI-17 by carbachol was low in accordance with the decrease in CPI-17 expression due to IL-1beta treatment. In contrast, constitutively phosphorylated MYPT-1 was also decreased in the IL-1beta-treated muscles. These results suggest that long term treatment with IL-1beta decreases either CPI-17 expression or MYPT-1 phosphorylation, which may result in an increase in myosin phosphatase activity to reduce force generation. Based on these findings, we consider IL-1beta to be an important mediator of gastrointestinal motility disorders in IBD, and CPI-17 and MYPT-1 are key molecules in the decreased smooth muscle contractility due to IL-1beta.
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PMID:Chronic treatment with interleukin-1beta attenuates contractions by decreasing the activities of CPI-17 and MYPT-1 in intestinal smooth muscle. 1451 13

Neuropeptide Y (NPY) elevates the permeability of cultured rat aortic endothelial cells (RAECs) in monolayer cultures under hypoxic conditions (5% O(2)) possibly by binding to the NPY Y(3) receptor. The present study evaluated the effects of NPY compared to vascular endothelial growth factor (VEGF). RAECs were cultured on the upper chamber base of a double-chamber culture system, FITC-labeled albumin was introduced into the chamber, and permeation into the lower chamber was measured. Treatment was with 3 x 10(-7) M NPY or 10(-7) g/ml VEGF for 2 h along with specific inhibitors. The VEGF receptor-2 tyrosine kinase inhibitor tyrphostin SU-1498 and the protein kinase C inhibitor bis-indolylmaleimide I (GF-109203X) suppressed the VEGF-induced increase in monolayer permeability but not that caused by NPY. Furthermore, although the action of NPY was blocked in a concentration-dependent manner by phospholipase C inhibitor 1-(6-[[(17beta)-3-methoxyestra-1,3,5(10)-trien-17-yl]amino]hexyl)-1H-pyrrole-2,5-dione (U-73122), it was less sensitive than VEGF. However, the effects of both NPY and VEGF on the permeability of the RAEC monolayer were blocked with equal concentration dependence by STI571 (imatinib mesylate), which is an inhibitor of Abl tyrosine kinase in the nucleus and/or cytoplasm. The myosin light-chain kinase inhibitor 1-(5-chloronaphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine HCl (ML-9) suppressed both NPY- and VEGF-induced increment in permeability by approximately 70%, whereas the calmodulin-dependent kinase inhibitor DY-9760e could decrease to below the baseline. These results indicate that the NPY Y(3)-receptor subtype is specifically linked to the effects of STI571 on endothelial cells, and that NPY, a sympathetic coneurotransmitter, may increase vascular permeability in association with altered intracellular or nuclear signal transduction.
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PMID:Effects of specific signal transduction inhibitors on increased permeability across rat endothelial monolayers induced by neuropeptide Y or VEGF. 1497 29

To clarify whether cyclic AMP (cAMP)/cAMP-dependent protein kinase (PKA) activation and Rho-kinase inhibition share a common mechanism to decrease the Ca2+ sensitivity of airway smooth muscle contraction, we examined the effects of 8-bromoadenosine 3',5'-cyclic monophosphate (8-BrcAMP), a stable cAMP analog, and (+)-(R)-trans-4-(1-aminoethyl)-N-(4-pyridyl) cyclohexane carboxamide dihydrochloride, monohydrate (Y-27632), a Rho-kinase inhibitor, on carbachol (CCh)-, guanosine 5'-O-(3-thiotriphosphate) (GTPgammaS)-, 4beta-phorbol 12,13-dibutyrate (PDBu)-, and leukotriene D4 (LTD4)-induced Ca2+ sensitization in alpha-toxin-permeabilized rabbit tracheal and human bronchial smooth muscle. In rabbit trachea, CCh-induced smooth muscle contraction was inhibited by 8-BrcAMP and Y-27632 to a similar extent. However, GTPgammaS-induced smooth muscle contraction was resistant to 8-BrcAMP. In the presence of a saturating concentration of Y-27632, PDBu-induced smooth muscle contraction was completely reversed by 8-BrcAMP. Conversely, PDBu-induced smooth muscle contraction was resistant to Y-27632. In the presence of a saturating concentration of 8-BrcAMP, GTPgammaS-induced Ca2+ sensitization was also reversed by Y-27632. The 8-BrcAMP had no effect on the ATP-triggered contraction of tracheal smooth muscle that had been treated with calyculin A in rigor solutions. The 8-BrcAMP and Y-27632 additively accelerated the relaxation rate of PDBu- and GTPgammaS-treated smooth muscle under myosin light chain kinase-inhibited conditions. In human bronchus, LTD4-induced smooth muscle contraction was inhibited by both 8-BrcAMP and Y-27632. We conclude that cAMP/PKA-induced Ca2+ desensitization contains at least two mechanisms: 1) inhibition of the muscarinic receptor signaling upstream from Rho activation and 2) cAMP/PKA's preferential reversal of PKC-mediated Ca2+ sensitization in airway smooth muscle.
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PMID:8-Bromo-cAMP decreases the Ca2+ sensitivity of airway smooth muscle contraction through a mechanism distinct from inhibition of Rho-kinase. 1512 38


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