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)

Rabbit ileum strips were functionally skinned by exposure to staphylococcal alpha-toxin. Incubation of the strips in the ATP analog ATP gamma S or [35S]ATP gamma S in the presence of Ca2+ (but not in the absence of Ca2+) resulted in a maximal Ca2+-insensitive activated tension that persisted following removal of Ca2+. Correlated with this tension was 35S-labeling of the 20,000-dalton myosin light chain, LC20, that persisted even after removal of Ca2+. Tension in these strips partially relaxed when exposed to ATP (alpha,beta-methylene). In contrast, alpha-toxin-treated strips exposed to ATP or [gamma-32P]ATP showed Ca2+-sensitive, reversible activated tension and reversible 32P-labeling of the LC20. These results are consistent with a currently proposed model of Ca2+ control of smooth muscle contraction involving a myosin light chain kinase-phosphatase system.
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PMID:Irreversible thiophosphorylation and activation of tension in functionally skinned rabbit ileum strips by [35S]ATP gamma S. 50 Jun 32

Protein tyrosine kinase (PTK) blockers (tyrphostins) inhibit in a dose-dependent fashion thrombin-induced aggregation and serotonin release with IC50 values in the 10-35 microM concentration range. The inhibition of thrombin-induced aggregation correlates with their potency in inhibiting phosphorylation of proteins on tyrosine residues. Using metabolically 32P-labelled human platelets, it was found that the tyrphostins have no effect on the decrease in [32P]phosphatidylinositol bisphosphate but prevent the replenishment of [32P]polyphosphoinositide. Tyrphostins decreased [32P]phosphatidic acid production induced by thrombin, although never by more than 50%, and only delayed the peak of diacylglycerol, suggesting that phospholipase C was still activated. Tyrphostins inhibited the thrombin-elicited early phosphorylation of p43 and p20, substrates for protein kinase C (PKC) and myosin light chain kinase, respectively, at short times of activation. This inhibition, however, was overcome after 1 min of stimulation with thrombin. Tyrphostin AG213 also inhibited platelet aggregation and tyrosine protein phosphorylation induced by phorbol myristate acetate (PMA), but did not inhibit pleckstrin phosphorylation. These results suggest that thrombin induces the phosphorylation of proteins on tyrosine residues which most probably results in the activation of phosphoinositide kinases. The ability of tyrphostins to inhibit phosphorylation of p43 and p20 when induced by thrombin but not when induced by PMA confirms that PTKs may be involved subsequent to PKC activation.
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PMID:Inhibition of platelet activation by tyrosine kinase inhibitors. 138 25

The effects of norepinephrine (NE), carbachol (CCh), NaF, 3-isobutyl-1-methylxanthine (IBMX), and high K+ concentration (80 mM) depolarization on inositol trisphosphate (IP3) accumulation, cyclic AMP (cAMP) formation, and contraction were investigated in the dilator and sphincter smooth muscles of the sympathetically denervated as well as the normal rabbit eye. (a) In the denervated dilator muscle, NE-stimulated IP3 production and contraction are enhanced. (b) In the sphincter muscle of rabbits that have undergone sympathetic denervation. CCh-stimulated IP3 production and contraction are attenuated. (c) The increase in tension by a maximal effective dose of NaF (209 mM) in the dilator was 12.5 and 18 mg of tension/mg wet weight in normal and denervated tissue, respectively, and in the sphincter was 33.8 and 15.2 mg of tension/mg wet weight in normal and denervated tissue, respectively. NaF had no effect on cAMP formation. (d) Addition of NE had no effect on cAMP formation in both the normal and denervated dilator, whereas basal and IBMX-induced cAMP formation increased. in the denervated sphincter over that of the normal tissue by 15 and 60%, respectively. (e) Isoproterenol (5 microM) increased cAMP formation in the normal and denervated sphincter by 47 and 91%, respectively. (f) Whereas CCh inhibits cAMP formation in the normal sphincter, it lost its inhibitory effect in the sphincter with denervation. (g) IBMX (0.1 mM) attenuated the CCh-stimulated IP3 production and contraction of the sphincter by approximately 30% of their respective controls. (h) High K+ concentration depolarization attenuated contraction in both dilator and sphincter muscles with denervation. These observations suggest that an increase in the level of cAMP in the iris sphincter due to sympathetic denervation could lead to inhibition of phospholipase C (or other target sites, such as phosphorylation of the muscarinic receptor, Gp protein itself, myosin light chain kinase, or the IP3 receptor), IP3 production, and contraction. In conclusion, we suggest that the supersensitivity and subsensitivity observed after surgical sympathetic denervation of the iris dilator and sphincter muscles, respectively, are caused by alterations in the efficiency of coupling, probably through the Gp proteins, between their respective receptors and the breakdown of polyphosphoinositides by phospholipase C. In addition, we propose that the sympathetic nervous system can regulate, through alterations in cAMP levels, the muscarinic stimulation of IP3 accumulation and contraction in the iris sphincter. These findings add further support to the hypothesis that there are reciprocal interactions between the cAMP and IP3-Ca2+ signaling systems and the contractile response in the iris smooth muscle.
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PMID:Effects of surgical sympathetic denervation on myo-inositol trisphosphate production and contraction in the dilator and sphincter smooth muscles of the rabbit iris: evidence for interaction between the cyclic AMP and calcium signaling systems. 171 29

During culture, smooth-muscle cells obtained from rabbit basilar arteries were examined for contractile activity by means of differential interference microscopy with a video analysis system (digital imaging microscopy system). This system proved useful for observing the contraction and ultrastructural changes of the living cells. Hemolysate-treated cells showed augmented responses to 5-hydroxytryptamine and leukotriene C4, but not to KCl. This augmented response diminished gradually during the culture period. Both a phospholipase C blocking agent, 2-nitro-4-carboxyphenyl-n,n-diphenylcarbamate (NCDC), and a myosin light chain kinase blocking agent, 1-(5-chloronaphthalenesulfonyl)-1H-hexahydro-1,4-diazepine (ML-9), suppressed this augmented response. Protein kinase C activity of the cells, as measured by Western blot analysis, did not increase during the period of culture with hemolysate. The results obtained suggest that hemolysate had the following effects on the cells: 1) acute but gradual contraction of the cells; 2) augmentation of cellular responses to vasoactive agents; and 3) progressive contraction and morphological alteration of the cells. Possible mechanisms by which hemolysate exerts these effects are discussed, taking into consideration the interrelationship between these effects.
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PMID:Altered reactivity of hemolysate-treated cultured smooth-muscle cells from rabbit basilar artery determined by digital imaging microscopy. 204 25

In primary hypertension, phospholipase C (PLC) is hypersensitive in several target tissues (platelets, vascular smooth muscle cells, aortic fibroblasts). Protein kinase C (PKC) and myosin light chain kinase (MLCK), which are physiologically activated by PLC-triggered second messengers (diacylglycerol and Ca2+ ions, respectively), phosphorylate specific proteins closely involved in the cell functional responses. In this study, we have examined and compared between platelets of spontaneously hypertensive rats (SHR) and their normotensive controls Wistar-Kyoto (WKY), the patterns of protein phosphorylation obtained either with the receptor-mediated agonist thrombin (i.e. which acts via PLC) or with direct activators of the protein kinases, PKC and MLCK. Activation by thrombin of 32P-prelabeled platelets induced incorporation of radioactivity into two proteins, P20 (myosin light chain) and P47. The curves obtained when platelets were challenged with either increasing doses of thrombin (0.025-0.3 U/ml) for 20 sec or with a low dose of the agent (0.1 U/ml) for up to 1 min, revealed that phosphorylation of the target proteins of PKC (P47) and of MLCK (P20) were significantly enhanced in platelets of SHR compared to WKY. In contrast, direct activation of PKC by phorbol ester and of MLCK by the calcium ionophore A23187 evoked the selective phosphorylation of the respective target proteins, P47 and P20, to a similar extent in platelets of SHR and WKY. Taken together, these results demonstrate that a physiological agonist (thrombin) induces an enhanced phosphorylation of intracellular proteins in platelets of SHR.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:[Increased phosphorylations of proteins involved in the expression of the physiologic response of platelets in SHR rats]. 212 75

1. Flash photolysis of caged compounds of phenylephrine, inositol 1, 4, 5 trisphosphate (InsP3), GTP gamma S, ATP, and CTP has been successfully used to study excitation-contraction coupling, contractile regulation, and contraction in smooth muscle. Major processes explored with this method were (a) the delay between agonist-receptor interaction and contraction and between the rise in InsP3, Ca2+ release and contraction; (b) the effect of myosin light chain phosphorylation on the rate of force development and the respective contributions of phosphorylation and crossbridge kinetics to differences between phasic and tonic smooth muscles; (c) the kinetics of the crossbridge cycle. We have also reviewed recent results obtained by other methods and bearing on the mechanisms of pharmacomechanical Ca2+ release and modulation of the Ca2+ sensitivity of the regulatory/contractile apparatus. 2. The long delay (1.5 at 22 degrees C) following activation of alpha 1-adrenergic receptors through photolysis of caged phenylephrine and the high Q10 of this process are consistent with the hypothesis that activation of phospholipase C is the major mechanism of alpha-adrenergic pharmacomechanical Ca2+ release. 3. The delay between photolysis of caged InsP3 and Ca2+ release is short: 30 ms or less, while the latency of contraction is significant (0.3-0.5 s at 22 degrees C) and similar to the lag between the rise in [Ca2+]i and force development in intact smooth muscles. The latency of contraction following photolysis of caged ATP in permeabilized muscles in rigor, in the presence of Ca2+ and calmodulin, is similar, about 0.2-0.5 s at 22 degrees C. 4. In muscles in which the myosin light chains are maintained in a phosphorylated state during rigor, photolysis of caged ATP initiates contractions with a short delay (10 ms or less). This result and those summarized above (2 and 3) suggest that the major portion of the delay between agonist-receptor interaction and contraction is due to activation of phospholipase C and InsP3 production, and about 0.2-0.5 s of the delay (22 degrees C) can be ascribed to prephosphorylation reactions between Ca2+, calmodulin, and myosin light chain kinase, and/or to mechanical processes, or to the chemical kinetics of two-step reactions. 5. Force development from rigor, initiated by photolysis of caged ATP in the presence of Ca2(+)-calmodulin, is rate-limited by myosin light chain phosphorylation; it is significantly accelerated if the myosin light chains are already phosphorylated prior to photolysis.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Flash photolysis studies of excitation-contraction coupling, regulation, and contraction in smooth muscle. 218 79

Fluoride interacts with G proteins and, consequently, stimulates phospholipase C as measured by the formation of inositol phosphates and phosphatidic acid. In human platelets this paralleled platelet aggregation and the activation of phosphorylation of substrates of protein kinase C (47kDa protein) and myosin light chain kinase (20kDa protein). Phospholipase C activation by fluoride was inhibited by dibutyryl cyclic AMP and by agents that increase cyclic AMP levels such as iloprost and forskolin. This information suggest that cyclic AMP affects the G protein associated with the stimulation of phospholipase C.
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PMID:Activation of platelet phospholipase C by fluoride is inhibited by elevation of cyclic AMP. 246 3

The mechanism of vasodilator-induced inhibition of platelet aggregation was investigated in human platelets. Cyclic nucleotide-elevating vasodilators stimulated cAMP- or cGMP-dependent protein phosphorylation, inhibited the activation of both protein kinase C and myosin light chain kinase, and inhibited the thrombin-induced hydrolysis of phosphatidylinositol-4,5-bisphosphate without affecting its resynthesis. The results suggest that cAMP- and cGMP-elevating vasodilators both inhibit platelet aggregation at an early step of the activation cascade, presumably at the level of phospholipase C.
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PMID:Cyclic nucleotide elevating vasodilators inhibit platelet aggregation at an early step of the activation cascade. 253 41

The photomechanical response of the vertebrate iris sphincter pupillae isolated from irises of many species of vertebrates contract when light is shined on them. It appears that the cell membranes of the constituent smooth muscle cells contain rhodopsin which triggers the photomechanical response (PMR) when bleached. In amphibians and some fish this mechanism of pupillary control is more important than the more well-known retinal reflex. In the mammals the retinal reflex is more important; however, even in the mammals the exact role of the innervation is not understood. The PMR can be inhibited by beta adrenergic agonists but not by alpha adrenergic agonists. The activation sequence of the PM probably involves (1) rhodopsin activated G-protein, (2) phospholipase C, (3) inositol triphosphate, and (4) a calcium-calmodulin-myosin light chain kinase cascade. A simple mathematical version of the phosphorylation theory of smooth muscle contraction accurately predicts the time courses of PMRs to light stimuli of different durations and intensities.
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PMID:Photomechanical coupling in the vertebrate sphincter pupillae. 265 40

We have examined regulation by protein kinase C (Ca2+/phospholipid-dependent enzyme) of thrombin-induced inositol polyphosphate accumulation in human platelets. When platelets are exposed to thrombin for 10 s, the protein kinase C inhibitor staurosporine causes inositol phosphate elevations over control values of 2.7-fold (inositol 1,4,5-trisphosphate (Ins(1,4,5)P3], 1.9-fold (inositol 1,3,4,5-tetrakisphosphate (Ins(1,3,4,5)P4], and 1.2-fold (inositol 1,3,4-trisphosphate). In the same period, phosphatidic acid and diacylglycerol are unaffected. The myosin light chain kinase inhibitor ML-7 has no effect on inositol phosphate accumulations. Staurosporine does not inhibit Ins(1,4,5)P3 3-kinase and 5-phosphomonoesterase activities in saponin-permeabilized platelets incubated with exogenous Ins(1,4,5)P3 unless the platelets have been exposed to thrombin and protein kinase C is consequently activated. The protein kinase C agonist beta-phorbol 12,13-dibutyrate increases the Vmax of the 3-kinase 1.8-fold, with little effect on Km. Our results provide strong evidence for a role for protein kinase C in regulating inositol phosphate levels in thrombin-activated platelets. We propose that endogenously activated protein kinase C removes Ins(1,4,5)P3 by stimulating both 5-phosphomonoesterase and Ins(1,4,5)P3 3-kinase. Initial activation of phospholipase C does not appear to be affected by such protein kinase C. Inhibition of protein kinase C by staurosporine decreases 5-phosphomonoesterase activity. The resulting elevated Ins(1,4,5)P3, as substrate for Ins(1,4,5)P3 3-kinase, promotes production of Ins(1,3,4,5)P4, which also may accumulate through decreased 5-phosphomonoesterase activity and elevated Ca2+ levels. These factors apparently counteract the inhibitory effect on 3-kinase, yielding a net increase in Ins(1,3,4,5)P4.
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PMID:Inhibition of protein kinase C by staurosporine promotes elevated accumulations of inositol trisphosphates and tetrakisphosphate in human platelets exposed to thrombin. 270 80


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