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)

The mechanism whereby gastrin triggers phosphoinositide breakdown was investigated in an enriched preparation of isolated rabbit parietal cells (approx. 75%). In a permeabilized preparation of myo-[3H]inositol-labelled cells, GTP[S], a non-hydrolysable GTP analogue, enhanced [3H]inositol trisphosphate ([3H]InsP3 accumulation in a dose-dependent manner; submaximal concentrations of GTP[S] (less than 10 microM), potentiated gastrin-induced [3H]InsP3 release; preincubation for 5 min with GDP[S], a non-hydrolysable GDP analogue, dose-dependently reduced [3H]InsP3 accumulation stimulated by gastrin even in presence of GTP[S]. Exposure of intact parietal cells for 3 h to pertussis toxin (PTx) (200 ng/ml) led to a 15-50% reduction in gastrin-induced [14C]aminopyrine [(14C]AP) uptake (an index of in vitro acid secretion) and [3H]inositol phosphate ([3H]InsP) accumulation. A decrease in the accumulation of the different [3H]inositol phosphate occurred in gastrin-stimulated parietal cells treated with PTx. A rightward shift of gastrin dose-response curves in the presence of PTx was observed for [14C]AP uptake (EC50 values: 0.125 +/- 0.045 nM without PTx and 1.05 +/- 0.63 nM with PTx), for [3H]InsP accumulation (EC50 values: 0.16 +/- 0.08 nM without PTx and 1.56 +/- 0.58 nM with PTx) and [125I]gastrin binding (IC50 values: 0.247 +/- 0.03 nM without PTx and 2.38 +/- 0.56 nM with PTx). In contrast, cholera toxin (CTx) treatment (100 ng/ml) for 3 h was without effect on gastrin-induced [3H]InsP accumulation. CTx induced a pronounced potentiation of gastrin-stimulated [14C]AP uptake; this effect can be mimicked by IBMX (a phosphodiesterase inhibitor) and by forskolin (an activator of adenylyl cyclase). We conclude that: (i) one or more than one G protein appeared to be involved in gastrin receptor coupling to phospholipase C (PL-C); (ii) these G proteins are not substrates for CTx; (iii) one of these appeared to be a PTx-sensitive 'Gi-like' protein which could be involved in hormone-induced acid secretion, (iiii) the potentiating effect of CTx observed on AP uptake stimulated by gastrin suggests the existence of a cooperative effect between cAMP pathway (CTx) and the gastrin-induced phosphoinositide breakdown in acid secretory activity of parietal cells.
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PMID:Involvement of a pertussis toxin-sensitive G protein in the action of gastrin on gastric parietal cells. 212 30

The alpha-adrenergic receptors mediate the effects of epinephrine and norepinephrine on cellular signaling systems via guanine nucleotide binding regulatory proteins (G-proteins). Three alpha-adrenergic receptor subtypes have been cloned: the alpha 1, the alpha 2-C10, and the alpha 2-C4 adrenergic receptors. To investigate functional differences between the different subtypes, we assessed the ability of each to interact with adenylyl cyclase and polyphosphoinositide metabolism by permanently and transiently expressing the DNAs encoding the alpha 1, the alpha 2-C10, and the alpha 2-C4 adrenergic receptors in cells lacking endogenous alpha-adrenergic receptors. Both alpha 2-C10 and alpha 2-C4 couple primarily to inhibition of adenylyl cyclase and to a lesser extent to stimulation of polyphosphoinositide hydrolysis. alpha 2-C10 inhibits adenylyl cyclase more efficiently than alpha 2-C4. Effects of the alpha 2-adrenergic receptors on adenylyl cyclase inhibition and on polyphosphoinositide hydrolysis are both mediated by pertussis toxin-sensitive G-proteins. The major coupling system of the alpha 1-adrenergic receptor is activation of phospholipase C via a pertussis toxin-insensitive G-protein. alpha 1-Adrenergic receptor stimulation can also increase intracellular cAMP by a mechanism that does not involve direct activation of adenylyl cyclase. As with the muscarinic cholinergic receptor family our results show that each of the alpha-adrenergic receptor subtypes can couple to multiple signal transduction pathways and suggest several generalities about the effector coupling mechanisms of G-protein-coupled receptors.
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PMID:Multiple second messenger pathways of alpha-adrenergic receptor subtypes expressed in eukaryotic cells. 215 28

Studies were performed to examine interactions between the adenylyl cyclase (AC) and phospholipase C (PLC) signaling systems in cultured rat inner medullary collecting duct cells. Stimulation of AC by either arginine vasopressin (AVP) or forskolin or addition of exogenous cAMP inhibits epidermal growth factor (EGF)-stimulated PLC. This inhibition is mediated by activation of cAMP-dependent kinase as it is prevented by pretreatment with the A-kinase inhibitor, N-[2-(methylamino)ethyl]-5-isoquinoline-sulfonamide (H8) but not by the C-kinase inhibitor, 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine (H7). Exposure to EGF eliminates AVP-stimulated cAMP generation. This is not mediated by a cyclooxygenase product as inhibition by EGF is observed even in the presence of the cyclooxygenase inhibitor, flurbiprofen. Inhibition by EGF is not due to an increase in inositol trisphosphate (IP3) as exposure of saponin-permeabilized cells to exogenous IP3 is without effect. Inhibition by EGF is prevented by pretreatment with the C-kinase inhibitor, H7, but not by the A-kinase inhibitor, H8. Exposure to the synthetic diacylglycerol (DAG), dioctanoylglycerol, also inhibits AVP-stimulated AC activity; therefore, inhibition by EGF is due to activation of protein kinase C. Thus, in cultured rat inner medullary collecting duct cells, cAMP and DAG function as mutually inhibitory second messengers with each impairing formation of the other.
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PMID:Cyclic adenosine monophosphate and diacylglycerol. Mutually inhibitory second messengers in cultured rat inner medullary collecting duct cells. 216 48

In order to evaluate the role of phosphoinositide turnover in growth factor action, we expressed human M1 muscarinic acetylcholine (Hm1) receptors in Chinese hamster lung fibroblasts (CCL39 cell line). In the transfected cells (39M1-81 clone), but not in wild type fibroblasts, the muscarinic agonist carbachol induced a release of inositol phosphates as strong as alpha-thrombin, a very potent growth factor and activator of phosphoinositide-specific phospholipase C (PLC) in this cell system. In contrast to thrombin, carbachol-stimulated PLC activity was not inhibited by pertussis toxin treatment of cells. At concentrations that elicited a comparable initial rate of inositol phosphate release (10 nM for thrombin and 0.1 mM for carbachol), both agents gave rise to an identical calcium signal and equally stimulated Na+/H+ exchange and the transcription of the early genes c-jun, c-fos, and c-myc. Surprisingly, however, carbachol is not a mitogen for 39M1-81 cells, and even if tested in association with insulin or fibroblast growth factor, its effects on cell proliferation remained weak when compared with thrombin. Also, the muscarinic agonist did not stimulate soft agar colony forming capacity and did not prevent growth arrest in Go upon serum deprivation of cycling 39M1-81 cells. The failure of carbachol to induce cell proliferation could not be attributed to rapid and complete desensitization of Hm1 receptors nor to the activation of inhibitory pathways like adenylyl cyclase stimulation. We conclude that strong and persistent activation of phosphoinositide turnover elicits early biochemical events generally associated with mitogenesis, but is not sufficient to stimulate or maintain continuous cell proliferation. On the basis of our results, we postulate that thrombin mitogenesis depends critically on signaling events different from phosphoinositide turnover, possibly the stimulation of a receptor tyrosine kinase or a Gi protein-activated tyrosine kinase.
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PMID:Strong and persistent activation of inositol lipid breakdown induces early mitogenic events but not Go to S phase progression in hamster fibroblasts. Comparison of thrombin and carbachol action in cells expressing M1 muscarinic acetylcholine receptors. 217 13

The activation of a variety of cell surface receptors results in a biphasic increase in the cytoplasmic Ca2+ concentration due to the release or mobilization of Ca2+ from intracellular stores and to the entry of Ca2+ from the extracellular space. It is well established that phosphatidylinositol 4,5-bisphosphate hydrolysis is responsible for the changes in Ca2+ homeostasis. Stimulation of Ca2(+)-mobilizing receptors also results in the phospholipase C-catalyzed hydrolysis of the minor plasma membrane phospholipid, phosphatidylinositol 4,5-bisphosphate, with the concomitant formation of inositol (1,4,5) trisphosphate [1,4,5)IP3) and diacylglycerol. Analogous to the adenylyl cyclase signaling system, receptor-mediated stimulation of phospholipase C also appears to be mediated by one or more intermediary guanine nucleotide-dependent regulatory proteins. There is strong evidence that (1,4,5)IP3 stimulates Ca2+ release from intracellular stores. The Ca2(+)-releasing actions of (1,4,5)IP3 are terminated by its metabolism through two distinct pathways. (1,4,5)IP3 is dephosphorylated by a 5-phosphatase to inositol (1,4) bisphosphate; alternatively, (1,4,5)IP3 can be phosphorylated to inositol (1,3,4,5) tetrakisphosphate by a 3-kinase. Whereas the mechanism of Ca2+ mobilization is understood, the precise mechanisms involved in Ca2+ entry are not known. A recent proposal that (1,4,5)IP3 secondarily elicits Ca2+ entry by emptying an intracellular Ca2+ pool will be considered. This review summarizes our current understanding of the mechanisms by which inositol phosphates regulate cytoplasmic Ca2+ concentrations.
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PMID:Inositol phosphate formation and its relationship to calcium signaling. 219 Aug 8

The last few years have evidenced a tremendous expansion in our appreciation of the role of regulatory GTP-binding proteins in cellular activation. The availability of cholera and pertussis toxins to detect G proteins as well as methodological advances in the study of cellular function has afforded the opportunity to examine G protein participation in many cellular events. Regulation of adenylyl cyclase and cyclic GMP phosphodiesterase by G proteins has been demonstrated. Phosphatidylinositol-4,5-biphosphate specific phospholipase C activity appears to be subject to G protein control. G proteins regulate inward K+ and Ca2+ channels through a mechanism which may be independent of effects on the above mentioned enzymes. Certainly, the number of G proteins which have been identified from sequencing of complementary DNA affords the potential for G protein involvement in many cellular events. Only three G proteins have however been isolated and functionally characterized, Gs, Gi and transducin. Whether all the functions of these proteins have been identified remains to be seen.
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PMID:Regulatory GTP-binding proteins: emerging concepts on their role in cell function. 243 67

cDNA cloning has identified the presence in the human genome of three genes encoding alpha subunits of pertussis toxin substrates, generically called "Gi." They are named alpha i-1, alpha i-2 and alpha i-3. However, none of these genes has been functionally identified with any of the alpha subunits of several possible G proteins, including pertussis toxin-sensitive Gp's, stimulatory to phospholipase C or A2, Gi, inhibitory to adenylyl cyclase, or Gk, stimulatory to a type of K+ channels. We now report the nucleotide sequence and the complete predicted amino acid sequence of human liver alpha i-3 and the partial amino acid sequence of proteolytic fragments of the alpha subunit of human erythrocyte Gk. The amino acid sequence of the proteolytic fragment is uniquely encoded by the cDNA of alpha i-3, thus identifying it as alpha k. The probable identity of alpha i-1 with alpha p and possible roles for alpha i-2, as well as additional roles for alpha i-1 and alpha i-3 (alpha k) are discussed.
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PMID:Alpha i-3 cDNA encodes the alpha subunit of Gk, the stimulatory G protein of receptor-regulated K+ channels. 245 65

This study examines the mechanism of guanine nucleotide-binding protein (G protein) coupling of receptors to phospholipase C. The Xenopus oocyte has a muscarinic receptor-activated Cl- current that is mediated by inositol 1,4,5-trisphosphate. Modulation of the muscarinic receptor-evoked Cl- current was examined under voltage clamp in oocytes injected with resolved G-protein subunits. The presence of an alpha subunit of G proteins in oocytes was shown by pertussis toxin-labeling of a 41-kDa band in oocyte membranes. The presence of the beta subunit of G proteins was demonstrated by immunoblotting experiments with an antiserum (U-49) that is specific for the beta subunit. Pertussis toxin treatment of oocytes resulted in the uncoupling of muscarinic receptors from activation of the Cl- current. Cells microinjected with 1.5 ng of human erythrocyte beta gamma-subunit complex or 1.0 ng of bovine brain beta gamma-subunit complex showed approximately a 95% reduction in the evoked Cl- current. Cells injected with equal volumes of protein storage vehicle showed no change in response. Cells injected with boiled beta gamma subunits, bovine serum albumin, or resolved alpha subunits also showed no reduction in response. Cells injected with various concentrations of beta gamma subunits showed a concentration dependence with half-maximal inhibition of the muscarinic activated Cl- current at about 10 nM. Cells injected with 1.0 ng of bovine brain beta gamma subunits could not respond to bath-applied agonist but could generate the Cl- current on intracellular injection of inositol 1,4,5-trisphosphate. These observations suggest that there is a G protein responsible for muscarinic receptor-mediated signal transduction through phospholipase C and that it is an alpha beta gamma heterotrimer. It appears that the mode of action of the G protein in the phospholipase C system may be similar to that of the hormone-activated adenylyl cyclase.
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PMID:Beta gamma subunits of GTP-binding proteins inhibit muscarinic receptor stimulation of phospholipase C. 246 57

In membranes of myeloid differentiated HL-60 cells, the chemotactic peptide N-formyl-methionyl-leucyl-phenylalanine stimulates phospholipase C via a pertussis toxin-sensitive G-protein but does not inhibit adenylyl cyclase. In these membranes, the chemotactic peptide markedly stimulates the cholera toxin-dependent [32P]ADP-ribosylation of two proteins with approximate molecular masses of 40 and 41 kDa, respectively. The radiolabeled proteins comigrate on sodium dodecyl sulfate-polyacrylamide gels with the two pertussis toxin substrates present in HL-60 membranes, alpha i2 and alpha i3. The effect of the chemotactic peptide is blocked by treatment of intact HL-60 cells with pertussis toxin. Peptide mapping studies using Staphylococcus aureus protease V8 reveal that the two radiolabeled proteins are structurally distinct. Thus, the agonist-activated formyl peptide receptor functionally interacts with two distinct pertussis toxin substrates, most likely with Gi2 and Gi3. As the third Gi protein, Gi1, appears to be absent from both HL-60 cells and from systems that clearly reveal hormonal inhibition of adenylyl cyclase, the results strongly suggest that primary structure alone does not suffice to determine which effector mechanism is regulated by a given Gi-protein.
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PMID:Two distinct Gi-proteins mediate formyl peptide receptor signal transduction in human leukemia (HL-60) cells. 251 19

The signal transduction pathways of the cloned human 5-HT1A receptor have been examined in two mammalian cell lines transiently (COS-7) or permanently (HeLa) expressing this receptor gene. In both systems, 5-hydroxytryptamine (5-HT, serotonin) mediated a marked inhibition of beta 2-adrenergic agonist-stimulated (80% inhibition in COS-7 cells) or forskolin-stimulated cAMP formation (up to 90% inhibition in HeLa cells). This serotonin effect (EC50 = 20 nM) could be competitively antagonized by metitepine and spiperone (Ki = 81 and 31 nM, respectively) and could also be blocked by pretreatment of cells with pertussis toxin. In both cell types, 5-HT failed to stimulate adenylyl cyclase through the expressed receptors. In HeLa cells, 5-HT also stimulated phospholipase C (approximately 40-75% stimulation of formation of inositol phosphates). Again, this effect was inhibited by metitepine. However, the EC50 of 5-HT was considerably higher (approximately 3.2 microM) than that found for inhibition of adenylyl cyclase. Both pathways were demonstrated to be similarly affected by pertussis toxin. These findings indicate that like the M2 and M3 muscarinic cholinergic receptors, the 5-HT1A receptor can couple to multiple transduction pathways with varying efficiencies via pertussis toxin-sensitive G-proteins. The lack of stimulation of cAMP formation by this 5-HT1A receptor may suggest the existence of another pharmacologically closely related receptor.
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PMID:Effector coupling mechanisms of the cloned 5-HT1A receptor. 254 39


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