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 cytokine-mediated stimulation of sphingomyelin (SM) metabolism is emerging as an important signal transduction pathway via the generation of ceramide and sphingosine, products which have been shown to affect a wide variety of biological processes. Because SM-mediated signal transduction is initiated via the hydrolysis of an integral membrane phospholipid by a phospholipase C-like enzyme (sphingomyelinase) to yield lipids which modulate protein kinase C activity, the SM and phosphatidylinositol (PI) signaling pathways share certain similarities. The present study was undertaken to examine the potential for interplay between SM and PI turnover by testing the effects of sphingosine, sphingosine-1-phosphate, and ceramide on PI turnover. In dermal fibroblasts, sphingosine stimulated a rapid dose-dependent hydrolysis of PI, yielding inositol 1,4,5-triphosphate, followed by increased levels of intracellular calcium. Sphingosine-induced inositol phosphate (IP) accumulation was observed between 5 and 30 microM sphingosine with a maximal accumulation of 2.7-fold over control levels. Enhanced IP formation was measured as early as 5 s following sphingosine treatment and IP levels remained elevated for more than 60 min. Intracellular calcium mobilization accompanied the dose-dependent accumulation of IPs in response to sphingosine, although this effect was not apparent until after a 30-40-s lag period. Interestingly, sphingosine-1-phosphate stimulated a more rapid release of intracellular Ca2+ than sphingosine, but it had no effect on PI turnover. DL-threo-Dihydrosphingosine, a competitive inhibitor of sphingosine kinase, stimulates both PI turnover and Ca2+ flux, but does not block the action of sphingosine relative to those two processes. Ceramide (added as C2-ceramide), N-stearylamine, and stearoyl-D-sphingosine did not affect PI turnover or Ca2+ mobilization. Pretreatment of intact cells with pertussis toxin partially inhibited sphingosine-mediated IP accumulation, suggesting a role for guanine nucleotide binding protein(s) (G protein) in sphingosine-stimulated PI turnover. Furthermore, guanosine 5'-O-(3-thiotriphosphate) stimulated, whereas guanosine 5'-O-(2-thiodiphosphate) inhibited, sphingosine-induced IP accumulation in permeabilized cells. Collectively, these data suggest that sphingosine enhances PI turnover by stimulating phospholipase C activity, and the activation of this process may be modulated by G protein interactions. Thus, the regulation of PI turnover and Ca2+ mobilization by sphingosine may represent another mechanism by which sphingosine modulates cell function and that these effects can be distinguished from those of ceramide.
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PMID:Sphingosine-mediated phosphatidylinositol metabolism and calcium mobilization. 811 27

Aggregation of receptors specific for the constant region of immunoglobulin G activates a repertoire of monocyte responses that can lead ultimately to targeted cell killing via antibody-directed cellular cytotoxicity. The high affinity receptor, FcgammaRI, contains no recognized signaling motif in its cytoplasmic tail but rather utilizes the gamma-chain of FcepsilonRI as an accessory molecule to recruit tyrosine kinases for signal transduction. We show here that, in a human monocytic cell line primed with interferon-gamma, FcgammaRI mobilizes intracellular calcium stores using a novel pathway that involves tyrosine kinase coupling to phospholipase D and resultant downstream activation of sphingosine kinase. Moreover, FcgammaRI is not coupled to phospholipase C; hence, calcium release from intracellular stores occurred in the absence of any measurable rise in inositol triphosphate. Finally, as this novel activation pathway is also shown to be responsible for mediating the vesicular trafficking of internalized immune complexes for degradation, it is likely to play a key role in controlling intracellular events triggered by FcgammaRI.
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PMID:FcgammaRI coupling to phospholipase D initiates sphingosine kinase-mediated calcium mobilization and vesicular trafficking. 954 63

Formation of inositol 1,4,5-trisphosphate (IP3) by phospholipase C (PLC) with subsequent release of Ca2+ from intracellular stores, is one of the major Ca2+ signalling pathways triggered by G-protein-coupled receptors (GPCRs). However, in a large number of cellular systems, Ca2+ mobilization by GPCRs apparently occurs independently of the PLC-IP3 pathway, mediated by an as yet unknown mechanism. The present study investigated whether sphingosine kinase activation, leading to production of sphingosine-1-phosphate (SPP), is involved in GPCR-mediated Ca2+ signalling as proposed for platelet-derived growth factor and FcepsilonRI antigen receptors. Inhibition of sphingosine kinase by DL-threo-dihydrosphingosine and N,N-dimethylsphingosine markedly inhibited [Ca2+]i increases elicited by m2 and m3 muscarinic acetylcholine receptors (mAChRs) expressed in HEK-293 cells without affecting mAChR-induced PLC stimulation. Activation of mAChRs rapidly and transiently stimulated production of SPP in HEK-293 cells. Finally, intracellular injection of SPP induced a rapid and transient Ca2+ mobilization in HEK-293 cells which was not antagonized by heparin. We conclude that mAChRs utilize the sphingosine kinase-SPP pathway in addition to PLC-IP3 to mediate Ca2+ mobilization. As Ca2+ signalling by various, but not all, GPCRs in different cell types was likewise attenuated by the sphingosine kinase inhibitors, we suggest a general role for sphingosine kinase, besides PLC, in mediation of GPCR-induced Ca2+ signalling.
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PMID:Sphingosine kinase-mediated Ca2+ signalling by G-protein-coupled receptors. 958 76

The early signaling mechanism of sphingosine 1-phosphate (S1P) on extracellular signal-regulated kinase (ERK) activation was investigated in C6 glioma cells. S1P activated the enzyme in association with a shift in the mobility on electrophoresis reflecting phosphorylation of both ERK1/ERK2 at as low as 10 nM. The lipid-induced ERK1/2 activation was partially inhibited by treatment of the cells with either phorbol 12-myristate 13-acetate (a long-term treatment to desensitize protein kinase C) or pertussis toxin (PTX) and was completely inhibited by a simultaneous treatment with both agents. Similarly, either calphostin C, an inhibitor of protein kinase C, or U73122, an inhibitor of phospholipase C, partially inhibited the S1Pinduced ERK1/2 activation in the nontreated cells with PTX and completely in the toxin-treated cells. On the other hand, the S1P-induced ERK activation was hardly affected by ethanol, which switched the product of phospholipase D from phosphatidic acid to metabolism-resistant phosphatidylethanol. S1P was able to activate ERK1/2 without a detectable increase in the intracellular content of the lipid, but sphingosine, a substrate of sphingosine kinase, which is an enzyme for S1P generation in the cells, hardly affected the ERK1/2 activation in spite of a marked elevation of intracellular S1P accumulation. This indicates that intracellular increase in S1P is not necessary for the S1P-induced ERK activation, and hence suggests the extracellular action mechanism of S1P. Supporting this idea, mRNAs of recently identified S1P specific receptors, Edg-1 and AGR16/H218, were expressed in C6 cells. Taken together, these results suggested that S1P acts on C6 cells extracellularly possibly through S1P receptors which are linked to at least two signaling pathways, i.e., the PTX-sensitive Gi/Go protein pathway and the toxin-insensitive Gq/G11-phospholipase C-PKC pathway, resulting in the activation of ERK.
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PMID:Possible involvement of cell surface receptors in sphingosine 1-phosphate-induced activation of extracellular signal-regulated kinase in C6 glioma cells. 988 6

Sphingosine-1-phosphate (SPP) produced from sphingosine by sphingosine kinase has recently been reported to act as intracellular second messenger for a number of plasma membrane receptors. In the present study, we investigated whether the sphingosine kinase/SPP pathway is involved in cellular signaling of the Gi protein-coupled formyl peptide receptor in myeloid differentiated human leukemia (HL-60) cells. Receptor activation resulted in rapid and transient production of SPP by sphingosine kinase, which was abolished after pertussis toxin treatment. Direct activation of heterotrimeric G proteins by AlF4- also rapidly increased SPP formation in intact HL-60 cells. In cytosolic preparations of HL-60 cells, sphingosine kinase activity was stimulated by the stable GTP analog, guanosine 5'-O-(3-thiotriphosphate). Inhibition of sphingosine kinase by DL-threo-dihydrosphingosine and N,N-dimethylsphingosine did not affect phospholipase C stimulation and superoxide production but markedly inhibited receptor-stimulated Ca2+ mobilization and enzyme release. We conclude that the formyl peptide receptor stimulates through Gi-type G proteins SPP production by sphingosine kinase, that the enzyme is also stimulated by direct G protein activation, and that the sphingosine kinase/SPP pathway apparently plays an important role in chemoattractant signaling in myeloid differentiated HL-60 cells.
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PMID:Formyl peptide receptor signaling in HL-60 cells through sphingosine kinase. 993 90

Contribution of sphingosine kinase (SPK)-catalyzed production of sphingosine-1-phosphate (SPP), in comparison to phospholipase C (PLC), to Ca(2+) signalling by epidermal growth factor (EGF) was studied in two HEK-293 cell clones (HEK2 and HEK3), expressing functional EGF receptors and exhibiting release of stored Ca(2+) by intracellular SPP. In HEK3 cells, EGF increased [Ca(2+)](i) and stimulated both, SPK and PLC. [Ca(2+)](i) increase, but not PLC stimulation, was strongly reduced by SPK inhibition. In HEK2 cells, EGF similarly stimulated PLC, but did not increase [Ca(2+)](i) or stimulate SPK, suggesting that intracellular SPP production plays a major role for Ca(2+) signalling by EGF in HEK-293 cells.
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PMID:Role of sphingosine kinase in Ca(2+) signalling by epidermal growth factor receptor. 1056

Vascular endothelial growth factor (VEGF) induces adhesion molecules on endothelial cells during inflammation. Here we examined the mechanisms underlying VEGF-stimulated expression of intercellular adhesion molecule 1 (ICAM-1), vascular cell adhesion molecule 1 (VCAM-1), and E-selectin in human umbilical vein endothelial cells. VEGF (20 ng/ml) increased expression of ICAM-1, VCAM-1, and E-selectin mRNAs in a time-dependent manner. These effects were significantly suppressed by Flk-1/kinase-insert domain containing receptor (KDR) antagonist and by inhibitors of phospholipase C, nuclear factor (NF)-kappaB, sphingosine kinase, and protein kinase C, but they were not affected by inhibitors of mitogen-activated protein/extracellular signal-regulated kinase kinase (MEK) 1/2 or nitric-oxide synthase. Unexpectedly, the phosphatidylinositol (PI) 3'-kinase inhibitor wortmannin enhanced both basal and VEGF-stimulated adhesion molecule expression, whereas insulin, a PI 3'-kinase activator, suppressed both basal and VEGF-stimulated expression. Gel shift analysis revealed that VEGF stimulated NF-kappaB activity. This effect was inhibited by phospholipase C, NF-kappaB, or protein kinase C inhibitor. VEGF increased VCAM-1 and ICAM-1 protein levels and increased leukocyte adhesiveness in a NF-kappaB-dependent manner. These results suggest that VEGF-stimulated expression of ICAM-1, VCAM-1, and E-selectin mRNAs was mainly through NF-kappaB activation with PI 3'-kinase-mediated suppression, but was independent of nitric oxide and MEK. Thus, VEGF simultaneously activates two signal transduction pathways that have opposite functions in the induction of adhesion molecule expression. The existence of parallel inverse signaling implies that the induction of adhesion molecule expression by VEGF is very finely regulated.
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PMID:Vascular endothelial growth factor expression of intercellular adhesion molecule 1 (ICAM-1), vascular cell adhesion molecule 1 (VCAM-1), and E-selectin through nuclear factor-kappa B activation in endothelial cells. 1110 18

Recently, a family of G-protein-coupled receptors named endothelial differentiation gene (Edg) receptor family has been identified, which are specifically activated by the two serum lipids, sphingosine-1-phosphate and lysophosphatidic acid. Sphingosine-1-phosphate can also act intracellularly to release Ca2+ from intracellular stores. Since in several cell types, G-protein-coupled lysophosphatidic acid or sphingosine-1-phosphate receptors mobilize Ca2+ in the absence of a measurable phospholipase C stimulation, it was analysed here whether intracellular sphingosine-1-phosphate production was the signalling mechanism used by extracellular sphingosine-1-phosphate for mobilization of stored Ca2+. Sphingosine-1-phosphate and the low affinity sphingosine-1-phosphate receptor agonist, sphingosylphosphorylcholine, induced a rapid, transient and nearly complete pertussis toxin-sensitive Ca2+ mobilization in human embryonic kidney (HEK-293) cells. The G-protein-coupled sphingosine-1-phosphate receptors, Edg-1, Edg-3 and Edg-5, were found to be endogenously expressed in these cells. Most interestingly, sphingosine-1-phosphate and sphingosylphosphorylcholine did not induce a measurable production of inositol-1,4,5-trisphosphate or accumulation of inositol phosphates. Instead, sphingosine-1-phosphate and sphingosylphosphorylcholine induced a rapid and transient increase in production of intracellular sphingosine-1-phosphate with a maximum of about 1.4-fold at 30 s. Stimulation of sphingosine-1-phosphate formation by sphingosine-1-phosphate and sphingosylphosphorylcholine was fully blocked by pertussis toxin, indicating that extracellular sphingosine-1-phosphate via endogenously expressed G(i)-coupled receptors induces a stimulation of intracellular sphingosine-1-phosphate production. As sphingosine-1-phosphate- and sphingosylphosphorylcholine-induced increases in intracellular Ca2+ were blunted by sphingosine kinase inhibitors, this sphingosine-1-phosphate production appears to mediate Ca2+ signalling by extracellular sphingosine-1-phosphate and sphingosylphosphorylcholine in HEK-293 cells.
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PMID:Stimulation of intracellular sphingosine-1-phosphate production by G-protein-coupled sphingosine-1-phosphate receptors. 1123 14

In SH-SY5Y cells, activation of delta-opioid receptors with [D-Pen(2,5)]-enkephalin (DPDPE; 1 microM) did not alter the intracellular free Ca(2+) concentration [Ca(2+)](i). However, when DPDPE was applied during concomitant Gq-coupled m3 muscarinic receptor stimulation by carbachol or oxotremorine-M, it produced an elevation of [Ca(2+)](i). The DPDPE-evoked increase in [Ca(2+)](i) was abolished when the carbachol-sensitive intracellular Ca(2+) store was emptied. There was a marked difference between the concentration-response relationship for the elevation of [Ca(2+)](i) by carbachol (EC(50) 13 microM, Hill slope 1) and the concentration-response relationship for carbachol's permissive action in revealing the delta-opioid receptor-mediated elevation of [Ca(2+)] (EC(50) 0.7 mM; Hill slope 1.8). Sequestration of free G protein beta gamma dimers by transient transfection of cells with a beta gamma binding protein (residues 495-689 of the C terminal tail of G protein-coupled receptor kinase 2) reduced the ability of delta opioid receptor activation to elevate [Ca(2+)](i). However, DPDPE did not elevate either basal or oxotremorine-M-evoked inositol phosphate production indicating that delta-opioid receptor activation did not stimulate phospholipase C. Furthermore, delta-opioid receptor activation did not result in the reversal of muscarinic receptor desensitization, membrane hyperpolarization or stimulation of sphingosine kinase. There was no coincident signalling between the delta-opioid receptor and the lysophosphatidic acid receptor which couples to elevation of [Ca(2+)](i) in SH-SY5Y cells by a PLC-independent mechanism. In SH-SY5Y cells the coincident signalling between the endogenously expressed delta-opioid and m3 muscarinic receptors appears to occur in the receptor activation-Ca(2+) release signalling pathway at a step after the activation of phospholipase C.
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PMID:Coincident signalling between the Gi/Go-coupled delta-opioid receptor and the Gq-coupled m3 muscarinic receptor at the level of intracellular free calcium in SH-SY5Y cells. 1125 87

Based on the finding that G protein-coupled receptors (GPCRs) can induce Ca2+ mobilization, apparently independent of the phospholipase C (PLC)/inositol-1,4,5-trisphosphate (IP3) pathway, we investigated whether sphingosine kinase, which generates sphingosine-1-phosphate (SPP), is involved in calcium signaling by mAChR and other GPCRs. Inhibition of sphingosine kinase by DL-threo-dihydrosphingosine and N,/N-dimethylsphingosine markedly inhibited [Ca2+]i increases elicited by M2 and M3 mAChRs in HEK-293 cells without affecting PLC activation. Activation of M2 and M3 mAChR rapidly and transiently stimulated production of SPP. Furthermore, microinjection of SPP into HEK-293 cells induced rapid and transient Ca2+ mobilization. Pretreatment of HEK-293 cells with the calcium chelator BAPTA/AM fully blocked mAChR-induced SPP production. On the other hand, incubation of HEK-293 cells with calcium ionophores activated SPP production. Similar findings were obtained for formyl peptide and P2Y2 purinergic receptors in HL-60 cells. On the basis of these studies we propose, that following initial IP3 production by receptor-mediated PLC activation, a local discrete increase in [Ca2+]i induces sphingosine kinase stimulation, which ultimately leads to full calcium mobilization. Thus, sphingosine kinase activation most likely represents an amplification system for calcium signaling by mAChRs and other GPCRs.
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PMID:Sphingosine kinase-mediated calcium signaling by muscarinic acetylcholine receptors. 1139 23


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