EC:2.7.11.13 - FACTA Search

Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.13 (protein kinase C)
49,245 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Depending on the vascular bed considered, the actions of ATP on the endothelium are mediated by either P2Y or P2U receptors. The two types of receptors seem to coexist on bovine aortic endothelial cells, where they are both coupled to phospholipase C. In this study, we have investigated whether they are truly coexpressed on the same cells and whether their signaling pathways diverge beyond phospholipase C activation. Measurements of [Ca2+]i in single cells showed that almost all bovine aortic endothelial cells are responsive to both 2-methylthio-ATP (2MeSATP), an agonist of P2Y receptors, and UTP, an agonist of P2U receptors. UTP stimulated the release of prostacyclin from freshly isolated bovine aortic endothelial cells, even when they were exposed to cycloheximide at the time of their collection: this indicates that P2U receptors must already be expressed on endothelial cells in situ and do not appear during cell culture. The time course of inositol phosphate (InsP) accumulation and the relative proportion of Ins(1,4,5)P3, Ins(1,3,4,5)P4, and Ins(1,3,4)P3 were similar in cells stimulated by 2MeSATP or UTP. UTP and 2MeSATP both stimulated the hydrolysis of phosphatidylcholine by phospholipase D, as reflected by the release of [3H]choline from prelabeled cells. The responses to both agents were blocked after downregulation of protein kinase C, resulting from a prolonged exposure to phorbol 12-myristate 13-acetate: this blockade occurred at a step distal to phospholipase C activation. A single difference between the two pathways has been identified: the effect of 2MeSATP on InsP3 was significantly more inhibited after a short exposure to phorbol 12-myristate 13-acetate than that of UTP.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Coexpression of P2Y and P2U receptors on aortic endothelial cells. Comparison of cell localization and signaling pathways. 783 29

Intracellular Ca2+ responses to extracellular matrix molecules were studied in suspensions of pancreatic acinar cells loaded with Fura-2. Collagen type I, laminin, fibrinogen and fibronectin were unable to raise cytosolic free Ca2+ concentration ([Ca2+]i), whereas collagen type IV, at concentrations from 5 to 50 micrograms/ml, significantly increased it. The effect of collagen type IV was not due to possible contamination with type-I transforming growth factor beta or plasminogen, as neither of these agents was able to increase [Ca2+]i. Using highly specific mass assays, concentrations of inositol lipids, 1,2-diacylglycerol (DAG) and Ins(1,4,5) P3 were measured in pancreatic acinar cells stimulated with collagen type IV. A decrease in the concentrations of PtdIns(4,5) P2 and PtdIns4 P with a concomitant increase in the concentrations of DAG and InsP3 mass were observed, showing that collagen type IV increases [Ca2+]i by activation of phospholipase C. The observed [Ca2+]i signals had two components, the first resulting from Ca2+ release from the intracellular stores, and the second resulting from Ca2+ flux from the extracellular medium through the verapamil-insensitive channels. A tyrosine kinase inhibitor (tyrphostine) was able to block inositol lipid signalling caused by collagen type IV, which together with the insensitivity of this pathway to cholera toxin and pertussis toxin or to preactivation of protein kinase C, the longer duration of the increase in [Ca2+]i and a longer lag period needed for observation of increases in DAG and InsP3 concentration with collagen type IV than with carbachol (50 mM) suggest that activation of phospholipase C by collagen type IV is caused by tyrosine kinase activation. Inositol lipid signalling and increases in [Ca2+]i were also observed with Arg-Gly-Asp (RGD)-containing peptide but not with Arg-Asp-Gly (RDG)-containing peptide. Collagen type IV and RGD-containing peptide, but not carbachol, competed in increasing [Ca2+]i and DAG concentration, suggesting that the binding site of collagen type IV responsible for phospholipase C activation contains the RGD sequence. Together the present results suggest that, in pancreatic acinar cells, RGD sequence(s) within collagen type IV molecules cause activation of tyrosine kinase, probably through one of the integrin receptors, which then stimulates phospholipase C and increases [Ca2+]i.
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PMID:Collagen type IV stimulates an increase in intracellular Ca2+ in pancreatic acinar cells via activation of phospholipase C. 819 49

In pancreatic islets stimulated with carbamylcholine (carbachol), hydrolysis of both phosphatidylinositol (PtdIns) and phosphatidylinositol bisphosphate (Ptd-InsP2) occurs and can be measured as the inositol monophosphates Ins(1)P1, or Ins(4)P1, respectively (Biden, T. J., Prugue, M. L., and Davison, A. G. M. (1992) Biochem. J. 285, 541-549). Our current aim was to establish whether these two events were independently regulated. Rats islets were labeled with either [3H]inositol or [3H]arachidonic acid for measurement of InsP1s by high performance liquid chromatography or diacylglycerol by TLC, respectively. The rise in Ins(1)P1 due to carbachol (1 min) was inhibited by 50% by concomitantly raising extracellular KCl ([K+]e) from 6 to 30 mM, thereby depolarizing the islets. Similar results, obtained in the absence of extracellular Ca2+, exclude the involvement of voltage-gated Ca2+ channels. Conversely, hyperpolarization, by lowering [K+]e to 3 mM, increased by 30% the rise in Ins(1)P1. In fact, over the [K+]e range of 3 to 48 mM, stimulated Ins(1)P1 accumulation was directly proportional to the calculated membrane potential. In contrast, raising [K+]e from 6 to 48 mM exerted no significant effect on carbachol-stimulated Ins(4)P1 levels, and both Ins(1)P1 and Ins(4)P1 were unaffected in the absence of carbachol. The rises in Ins(1)P1 (but not Ins(4)P1) were also inhibited by depolarization with the sodium pump inhibitor, ouabain, or the K+ channel blocker, tolbutamide. Stimulated diacylglycerol accumulation and insulin secretion (20 min) showed a biphasic dependency on [K+]e, being less pronounced at 6 mM than at either 3 or 30 mM KCl. This reflects a selective potentiation of PtdIns and PtdInsP2 hydrolysis, due, respectively, to hyperpolarization and the gating of voltage-dependent Ca2+ channels. The differential regulation of these two hydrolytic events is probably important for independent control of the activation of protein kinase C and Ca2+ mobilization and might play a role in modulating the secretory response in vivo.
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PMID:Regulation by membrane potential of phosphatidylinositol hydrolysis in pancreatic islets. 849 68

There is now good evidence that in the AD brain, a number of neurotransmitter effector systems are defective. Such abnormalities include defective G, protein and protein kinase C function as well as a drastically reduced level of receptors for the second messenger Ins(1,4,5) P3. Such changes are probably not restricted to the late stages of the disease, and are found in regions of the brain that show little histopathological abnormality, such as the cerebellum. Whether these changes precede or are secondary to primary histopathological changes such as beta-amyloid deposition is not as yet clear. What is clear, however, is that such signal transduction abnormalities are likely to negate therapeutic benefits in clinical strategies based upon the tenet of neurotransmitter replacement.
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PMID:Receptor-effector coupling dysfunctions in Alzheimer's disease. 868 30

Age-related functional alterations in a variety of neurotransmitter systems result in modulation of interneuronal communications which has some relevance in neurological deficits observed in the aging process. The synergistic interactions between protein kinase and inositol 1,4,5-trisphosphate (insP3)/Ca2+ pathways underlie a variety of cellular responses to external stimuli. To determine whether age-dependent changes occur in the regulation of protein kinase C and inositol 1,4,5-trisphosphate/Ca2+ pathways, insP3 contents as a marker for the release of intracellular calcium, saturation binding analysis of Ins P3 receptor using [3H]inositol 1,4,5-trisphosphate, slot/northern blot analysis of Ins P3 receptor-encoding mRNA transcripts, and the activities of Ca2+/phospholipid-dependent protein kinase C isozymes were investigated in the rat spinal cord. Inositol 1,4,5-trisphosphate content and [3H]inositol 1,4,5-trisphosphate binding site density (Bmax) were quantified in the spinal cords of young (three months old), adult (12 months old) and senescent (25 months old) male Fischer 344 rats. Spinal cord content of inositol 1,4,5-trisphosphate was increased (P < 0.01) in the 25-month old compared to the three- and 12-month old animals. The density of Ins P3 receptor in particulate membranes derived from the 25-month old rats was reduced (P < or = 0.01), but the binding affinity (Kd) was increased (P < or = 0.04) by a factor of 2.2 and 3.2 at 25 months of age when compared with three- and 12-month old animals, respectively. Young and middle-aged animals showed no differences in both inositol 1,4,5-trisphosphate contents and [3H]inositol 1,4,5-trisphosphate binding site density. The quantity of Ins P3 receptor mRNA was significantly increased with age in the order 25 >> 12 > 3 months of age. Total functional cytosolic and membrane-associated PKC activities were decreased (P < or = 0.05) in the 25-month compared to the three- and 12-month old rats in which activity remained unchanged. Total membrane/cytosolic activity ratios were unchanged by the aging process. In all cases, the activities of membrane-associated conventional protein kinase C isozymes (alpha, beta and gamma), determined by immunoprecipitation followed by in situ quantification of protein kinase C activities in the immunoprecipitates, showed age-dependent decline. The activities of protein kinase C-alpha and beta were significantly decreased in age-related manner. However, the activity of the gamma-isozyme was not significantly changed at 12- and 25-months of age, although it was higher (P < or = 0.03) in young rats. Western blot analyses using affinity purified polyclonal antibodies specific for each isozyme indicated a single protein with an apparent molecular mass of approximately 80 x 10(3) molec. weight for all isozymes except for the beta isozyme that also had an appreciable immunoreactive band at approximately 36 x 10(3) molec. weight. Overall, the aging process did not affect the electropheretic mobility of each isozyme. With decreased protein kinase C activity, the present data suggest that the aging process would decrease protein kinase C-induced phosphorylation of membrane proteins including Ins P3 receptor. A significant change in Ins P3 receptor affinity combined with increased levels of Ins P3 receptor mRNA-encoding transcripts in senescent rats suggests not only a modification (possibly by phosphorylation) of Ins P3 receptor protein but also the existence of multiple (spliced) variants of Ins P3 receptor in spinal neurons with increasing age. The present data indicate that the spinal contents of inositol 1,4,5-trisphosphate increased with age, but with decreased efficacy and number of inositol 1,4,5-trisphosphate-activatable Ca2+ channels in the spinal cord of senescent rats. These age-related changes may contribute to the attenuated responsiveness of spinal cord neurons by phosphoinositide-coupled receptors during the aging process.
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PMID:Regulation of phosphatidylinositide transduction system in the rat spinal cord during aging. 884 10

To better understand the molecular mechanisms that underlie the exaggerated bradykinin (BK)-stimulated release of Ins(1,4,5)P3 in fibroblasts from Alzheimer patients, the role of G-proteins, protein kinase C (PKC) and cyclic AMP in BK-induced Ins(1,4,5)P3 formation was determined. A role for G-proteins in the coupling of the BK receptor to intracellular signals was indicated by guanosine 5'-(3-O-thio)triphosphate (GTP gamma S) enhanced BK-stimulated Ins(1,4,5)P3 release. The coupling of G-proteins to Ins(1,4,5)P3 formation was sensitive to cholera toxin (CTX), but not pertussis toxin (PTX), and was not altered by PKC activation. The inhibition by CTX appeared to be secondary to its ability to increase cyclic AMP, because forskolin also inhibited the BK-mediated Ins (1,4,5)P3 release. Activation of PKC with TPA diminished the number of BK receptors by 33% and proportionally decreased BK-mediated Ins(1,4,5)P3 formation by 28%. The latter response was abolished by PKC inhibitors. Depletion of PKC by prolonged TPA treatment did not further alter the number of BK receptors but further decreased the Ins(1,4,5)P3 response by 65%. Thus, changes in PKC probably do not underlie the enhanced BK-induced Ins(1,4,5)P3 formation in AD fibroblasts, because both activation and depletion of the PKC diminished the Ins(1,4,5)P3 response.
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PMID:Regulation of bradykinin-induced Ins(1,4,5)P3 formation by protein kinase C in human fibroblasts. 889 Sep 34

The discovery of new pharmacologic and biochemical tools has prompted intensive research on the intracellular mechanisms conveying the physiologic message carried by angiotensin II (A II). Virtually all the cardiovascular effects of A II are activated by mobilization of the calcium messenger system through the AT1-receptor subtype. The AT2 subtype, which is highly expressed in fetal tissues, appears to be silent in adult tissues but may play a role in growth-related functions. Several functional domains that are involved in distinct processes have been identified in the AT1 receptor. Through a GTP-binding protein (Gq), A II activates a phospholipase C, which generates inositol 1,4,5-trisphosphate (Ins[1,4,5]P3) and diacylglycerol. Ins(1,4,5)P3 releases calcium from intracellular stores, which is a signal for a "capacitative" calcium influx. The net result of the various processes of calcium trafficking is an initial transient peak of cytosolic calcium concentration ([Ca2+]c) followed by a sustained response. A II also induces a translocation of protein kinase C (PKC) from the cytosol to the cell membrane. PKC can either potentiate or counteract the responses elicited by the [Ca2+]c changes. A II also alters the activity of voltage-gated calcium channels and of the sodium-calcium exchanger. Finally, the activity of adenylyl cyclase can also be affected. By contrast, the signaling mechanisms linked to the AT2-receptor subtype are poorly understood. The integration of these multiple and variable signals, as well as the cell's enzymatic repertory, eventually determine the specific cellular response. The unraveling of these complex mechanisms opens new perspectives for the development of therapeutic tools that could interfere more specifically with the intracellular processes of A II and its effects on the cardiovascular system.
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PMID:Distribution and signal transduction of angiotensin II AT1 and AT2 receptors. 891 39

Stimulation of bovine iris sphincter muscle with carbachol (10 microM) increased accumulation of Ins(1,4,5)P3 (InsP3) and Ins(1,3,4,5)P4 (InsP4) by 86 and 32% respectively. Addition of isoproterenol (5 microM) to muscle pretreated with carbachol reduced the 3H-radioactivity in InsP3 by 30% and increased that of InsP4 by 41%. InsP3 3-kinase was predominantly localized in the soluble fraction (110,000 g supernatant) of the iris sphincter. The enzyme was purified from this fraction by sequential chromatography on DEAE-cellulose, calmodulin (CAM)-agarose affinity, and Mono-Q anion-exchange columns. The specific activity of the purified enzyme was 1.94 mumol/min per mg protein with a purification of 114-fold, compared with the cytosolic fraction of the muscle. SDS/PAGE showed the enzyme to be associated with a protein band corresponding to 50 kDa. In the presence of 10 microM Ca2+, CaM dose-dependently stimulated the enzyme. InsP3 3-kinase specifically phosphorylated InsP3 with an apparent K(m) of 0.56 microM and a Vmax. of 2.5 mumol/min per mg protein. The stimulatory effect of CaM was due to a change in Vmax. and not in its K(m). The enzyme was maximally active at pH 7.0-7.5. Phosphorylation of the purified InsP3 3-kinase with protein kinase A increased its activity; in contrast, phosphorylation with protein kinase C inhibited the enzyme activity. Treatment of the intact iris sphincter with isoproterenol or phorbol 12,13-dibutyrate resulted in stimulation of InsP3 3-kinase activity in the soluble fraction and this activation was preserved on SDS/PAGE and renaturation. These results indicate that the bovine iris sphincter contains a Ca-CaM-dependent InsP3 3-kinase which can be differentially regulated, both in vitro and in intact muscle, by protein kinases A and C.
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PMID:Purification and properties of D-myo-inositol 1,4,5-trisphosphate 3-kinase from bovine iris sphincter smooth muscle: effects of protein phosphorylation in vitro and in intact muscle. 894 63

Thrombopoietin (TPO) is the major regulator of the proliferation and differentiation of megakaryocyte precursors through interaction with its receptor encoded by the c-mpl protooncogene. We established the human TPO-dependent leukemia cell line, UT-7/TPO (Blood 87, 4552, 1996). In these cells, TPO activated protein kinase C (PKC) in a time dependent manner. Subsequently, the c-myc gene was transiently induced to a maximal level 60-90 minutes after TPO exposure. In addition, we found that stimulating UT-7/TPO cells with TPO rapidly induces the significant accumulation of inositol 1, 4, 5-trisphosphate (Ins-P3), leading to the mobilization of calcium from intracellular stores. Taken together, the activation of PKC and subsequent c-myc gene induction are involved in the TPO-induced cellular response(s), presumably through the activation of PLC.
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PMID:Protein kinase C and c-myc gene activation pathways in thrombopoietin signal transduction. 907 Feb 65

We performed experiments to investigate the actions of protein kinase C(PKC) on mechanical contraction during agonist stimulation in guinea pig stomach. We used carbachol and high K condition to enhance mechanical contraction by mobilizing intracellular Ca2+ and increasing Ca2+ influx through voltage-dependent Ca(2+)-channel, respectively. Phorbol 12, 13-dibutyrate (PDBu) increased spontaneous contractions sensitive to verapamil (438 +/- 82.2%, n = 7) and potentiated high K-induced contraction (189 +/- 22.5%, n = 5). However, carbachol (CCh)-induced contractions in PDBu-treated condition depended on extracellular Ca2+. In the presence of extracellular Ca2+, CCh-induced contraction was potentiated, while it was suppressed in the absence of extracellular Ca(2+)-preloaded muscle strips. To prove the hypothesis that such phenomena might be related with changes of myoplasmic Ca2+ concentration, we investigated the effect of PDBu on voltage-dependent Ca2+ current(ICa) and CCh-induced Ca2+ activated K current(IK(Ca)) transient using whole-cell voltage clamp technique. For recording voltage-dependent Ca2+ current(ICa), 10 mM Ba2+, instead of Ca2+, was used to enhance the current size. Voltage-dependent Ba2+ current(IBa) was increased by PDBu (212 +/- 32.2% of steady state currents, n = 5), while CCh-induced increase of IK(Ca) transient was inhibited by PDBu (n = 5), the changes of which were similar to those of muscle contractions. To analyze the steps involved in the inhibition of CCh-induced IK(Ca) transient by PDBu, we investigated the effect of PDBu on IK(Ca) in the cells perfused with Ins (1, 4, 5)P3. However, Ins (1, 4, 5)P3 induced IK(Ca) was not inhibited by the treatment with phorbol ester. From these results, it is concluded that inhibition of phosphatidylinositol phospholipase C(PI-PLC) system and potentiation of ICa by PKC are important regulatory mechanisms in agonist-induced muscle contraction.
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PMID:Dual roles of phorbol 12, 13-dibutyrate in the regulation of guinea-pig gastric contraction. 912 43

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