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 PTH/PTH-related peptide (PTHrP) receptor and the calcitonin receptor mediate the action of their physiological ligands by activating two different effectors, adenylyl cyclase and phospholipase C. Whereas regulation of adenylyl cyclase via both receptors is thought to involve the G protein G(s), it is not known whether activation of phospholipase C results from coupling of the receptors to G(q) family members or whether beta gamma-subunit released from receptor-activated G(s) lead to phospholipase C activation. To elucidate the mechanism of this type of dual signaling, we reconstituted the signal transduction of the PTH/PTHrP and the calcitonin receptor in COS-7 and HEK293 cells. In COS-7 cells expressing the receptor alone, addition of the respective ligands resulted in the accumulation of cAMP and inositol phosphates. When cells were cotransfected with the cDNAs of receptor and different alpha-subunits of the Gq family (G alpha q, G alpha 11, G alpha 14, G alpha 15, and G alpha 16, a severalfold increase in the ligand-dependent inositol phosphate production could be observed, indicating that the receptors functionally interacted with all alpha-subunits of the G alpha q family. Additionally, whereas PTH treatment of HEK293 cells coexpressing both the PTH/PTHrP receptor and G alpha q increased both second messengers, the same treatment in cells expressing the PTH/PTHrP receptor alone increased only cAMP. Under all conditions tested, activation of phospholipase C via the PTH/PTHrP and calcitonin receptor required higher ligand concentrations than receptor-mediated adenylyl cyclase activation. Our data strongly support the idea that dual signaling of the PTH/PTHrP and calcitonin receptors is due to the a activation of different G proteins belonging to the G(s) and G(q) families.
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PMID:G alpha q family members couple parathyroid hormone (PTH)/PTH-related peptide and calcitonin receptors to phospholipase C in COS-7 cells. 873 87

Due to the importance of Ca2+ in the regulation of vital cellular and tissue functions, the concentration of Ca2+ in body fluids is closely guarded by an efficient feedback control system. This system includes Ca(2+)-transporting subsystems (bone, and kidney), Ca2+ sensing, possibly by a calcium-sensing receptor, and calcium-regulating hormones (parathyroid hormone [PTH], calcitonin [CT], and 1,25-dihydroxyvitamin D3 [1,25(OH)2D3]). In humans and birds, acute Ca2+ perturbations are handled mainly by modulation of kidney Ca2+ reabsorption and by bone Ca2+ flow under PTH and possibly CT regulation, respectively. Chronic perturbations are also handled by the more sluggish but economic regulatory action of 1,25(OH2)D3 on intestinal calcium absorption. Peptide hormone secretion is modulated by Ca2+ and several secretagogues. The hormones' signal is produced by interaction with their respective receptors, which evokes the cAMP and phospholipase C-IP3-Ca2+ signal transduction pathways. 1,25 (OH)2D3 operates through a cytoplasmic receptor in controlling transcription and through a membrane receptor that activates the Ca2+ and phospholipase C messenger system. The calciotropic hormones also influence processes not directly associated with Ca2+ regulation, such as cell differentiation, and may thus affect the calcium-regulating subsystems also indirectly.
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PMID:Homeostatic control of plasma calcium concentration. 874 55

A midregion fragment of PTH-related protein (PTHrP), which is intensively conserved across species, has been identified as a secretory product of several different cell types, including keratinocytes and squamous carcinomas. As recent data suggest that a midregion PTHrP fragment may be biologically active, we hypothesized that midregion PTHrPs interact with unique cell surface receptors that mediate autocrine or paracrine action. Dose-dependent transient elevations in intracellular calcium ([Ca2-]i) were observed in fura-2-loaded SqCC/Y1 squamous carcinoma cells exposed to human (h) PTHrP-(67-86)NH2, [Tyr36]hPTHrP-(1-36)NH2, and hPTHrP-(1-141) at concentrations ranging from 1 pM to 1 microM. The effects of maximal stimulatory concentrations of [Tyr36]PTHrP-(1-36)NH2 and PTHrP-(67-86)NH2 on [Ca2+]i were additive. The inhibitory PTH analog, [D-Trp12,Tyr34]bovine PTH-(7-34)NH2, attenuated the [Ca2+]i response to [Tyr36]hPTHrP-(1-36)NH2, but not that to PTHrP-(67-86)NH2. These data suggest that PTHrP-(67-86)NH2 activates a different receptor pathway in SqCC/Y1 cells from the one activated by [Tyr36]hPTHrP-(1-36)NH2. Radiolabeled PTHrP-(67-86)NH2 did not bind to SqCC/Y1 cells, and PTHrP-(67-86)NH2 did not compete for binding of 125I-labeled [Tyr36]PTHrP-(1-36)NH2 to PTH/PTHrP receptors on SaOS-2 osteosarcoma cells. Activation of the phospholipase C pathway by PTHrP-(67-86)NH2 was confirmed by exposing SqCC/Y1 cells to peptide for 1 min and measuring the accumulation of inositol trisphosphates. PTHrP-(67-86)NH2 treatment (100 nM) resulted in maximal stimulation of inositol trisphosphates of 3.1 +/- 0.1-fold over the control value, with an EC50 of 1.5 +/- 1.2 nm. In contrast, PTHrP-(67-86)NH2 (0.1 nM to 1 microM) did not stimulate adenylyl cyclase in SqCC/Y1 cells despite vigorous stimulation of cAMP formation by isoproterenol (1 microM) to 66-fold over the basal value. To determine whether messenger RNA (mRNA) prepared from SqCC/Y1 cells would direct the translation of a receptor protein that mediated a [Ca2+]i response to PTHrP-(67-86)NH2, we performed expression studies in Xenopus oocytes. Fluo-3 fluorescence in Xenopus oocytes expressing SqCC/Y1 mRNA was visualized by confocal video microscopy after exposure to 1 microM PTHrP-(67-86)NH2. Clear increases in [Ca2+]i were detected in mRNA-injected, but not in sham-injected, oocytes. Finally, we examined the effect of PTHrP-(67-86)NH2 treatment on fibronectin secretion from SqCC/YN1 cells. A significant 3.5-fold increase in fibronectin secretion into conditioned medium was observed when SqCC/Y1 cells were exposed to 100 nM PTHrP-(67-86)NH2, and this effect was dose dependent, with an EC50 of 0.1 nM. We conclude that PTHrP-(67-86)NH2 activates phospholipase C-dependent pathways in SqCC/Y1 cells through a receptor distinct from that activated by PTHrP-(1-36) in the same cells. As a midregion secretory fragment of PTHrP has been partially purified from several different cell types, this receptor may have broad biological significance.
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PMID:A midregion parathyroid hormone-related peptide mobilizes cytosolic calcium and stimulates formation of inositol trisphosphate in a squamous carcinoma cell line. 894 Mar 60

PTH-induced mobilization of cytosolic Ca2+ in a human kidney cell line (HEK/W) occurring in the absence of cAMP stimulation was characterized and compared with that obtained in the same cells stably transfected by the PTH/PTH-related peptide (PTHrp) receptor (HEK/T). In both cell lines, N-terminal fragments of PTH and PTHrp induced a concentration-dependent biphasic stimulation in [Ca2+]i: a transient peak followed by a slow linear increase. These increases in [Ca2+]i were inhibited by the PTH antagonist [Nle(8,18),Tyr(34)]bPTH(3-34). The transient peaks were due to calcium release from intracellular stores, as they resisted quenching of calcium in the extracellular buffer and were abolished by prior emptying of intracellular stores. These peaks differed, however, both in latency period and in magnitude, in the two cell lines. The phospholipase C inhibitor U73122 inhibited the PTH-induced increase in [Ca2+]i in HEK/T cells, but not in HEK/W. Similarly, PTH-induced inositol phosphate (InsPs) production was detected in HEK/T but not in HEK/W cells. PTH-induced calcium release in HEK/W cells was inhibited by the simultaneous presence of ryanodine and U73122. Low level PTH/PTHrp receptor messenger RNA expression was demonstrated by ribonuclease protection in HEK/W cells, although no specific binding of [125I]PTHrP(1-34) could be detected. Amplification products for the PTH/PTHrp receptor 1, but no other isoforms, were detected by RT-PCR in HEK/W cells. As expected, HEK/T cells responded to PTH by a 500-fold stimulation in cAMP production and expressed large numbers of PTH/PTHrp receptors, as shown by [125I]PTHrp binding. These results demonstrate that the signal transduction pathways activated by PTH in HEK/W and HEK/T cells are different. Because the major difference in these cell lines is the number of PTH/PTHrp receptors expressed, these results suggest that the transduction of signals by the PTH/PTHrp receptor is controlled by receptor number in such a way that PTH stimulates an increase in intracellular calcium in the absence of stimulation of InsPs and cAMP production in cells expressing low levels of PTH/PTHrp receptor, but stimulates calcium release through an InsPs pathway and induces cAMP production in cells expressing large numbers of PTH/PTHrp receptors. The control of receptor number may be one of the mechanisms through which PTH effects are regulated.
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PMID:Parathyroid hormone-induced calcium release from intracellular stores in a human kidney cell line in the absence of stimulation of cyclic adenosine 3',5'-monophosphate production. 938 12

The present study was performed to characterize the possible involvement of cAMP synthesis and protein kinase C (PKC) activation in the DNA synthesis-stimulating effect of parathyroid hormone-related protein (PTHrP) in proximal tubule cells. We found that DNA synthesis was stimulated by 10 microM 8BrcAMP, and 1 microM Sp-cDBIMPS, two cAMP analogs, and also by 1 microM phorbol 12-myristate 13-acetate (PMA) and 100 microM 1,2-dioctanoyl-sn-glycerol, two PKC activators, and 10 nM [Cys23] human (h)PTHrP (24-35) amide in rabbit proximal tubule cells (PTC). Both Sp-cDBIMPS and PMA, at 1 microM, also increased DNA synthesis in SV40-immortalized mouse proximal tubule cells MCT. Human PTHrP (7-34) amide [PTHrP (7-34)] dose dependently stimulated DNA synthesis in a similar manner as [34Tyr]PTHrP (1-34) amide [PTHrP (1-34)], in PTC. PMA pre-treatment for 20 h, which downregulates PKC, completely blocked the effect induced by PTHrP (7-34), but not that of PTHrP (1-34), in the latter cells. In contrast, the same PMA pre-treatment abolished the DNA synthesis stimulation by PTHrP (1-34) and PTHrP (7-34) in MCT cells, which appear to have PTH receptors mainly coupled to phospholipase C and not adenylate cyclase. Our results indicate that the stimulatory effect of PTHrP on DNA synthesis in proximal tubule cells is mediated by a cAMP- and PKC-dependent mechanism.
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PMID:Parathyroid hormone-related protein increases DNA synthesis in proximal tubule cells by cyclic AMP- and protein kinase C-dependent pathways. 965 Nov 15

The carboxyl(C)-truncated human (h) PTH (hPTH) analog hPTH(1-31), which activates adenylyl cyclase (AC), but not protein kinase C, in rat osteosarcoma cells, exerts an anabolic effect on rat bone in vivo similar to that of hPTH(1-34). It has been proposed, therefore, that this action of PTH(1-34) is mediated exclusively by stimulation of AC via the rat type-1 PTH/PTH-related peptide (PTHrP) receptor (PTH1R). To determine whether this selective signaling pattern also might be a property of the hPTH1R, we studied signal transduction via heterologously expressed hPTH1Rs in response to activation by hPTH(1-34), hPTH(1-31), and a C-truncated analog that does not increase rat bone mass in vivo, hPTH(1-30). In porcine LLC-PK1 cells that stably expressed recombinant hPTH1Rs, these three peptides activated AC identically (EC50 = 1-2 nM). In cells with comparable expression of rat PTH1Rs, AC activation by hPTH(1-34) and hPTH(1-31) again was identical, whereas full activation by hPTH(1-30) required higher concentrations (EC50 = 10 nM vs. 1 nM). Surprisingly, hPTH(1-31) fully stimulated phospholipase C (PLC), via both species of PTH1Rs, with potency that was similar (hPTH1Rs) or slightly reduced (rat PTH1Rs), relative to that of hPTH(1-34). hPTH(1-30), however, was 5-fold less potent than hPTH(1-34) in activating PLC via hPTH1Rs and showed weak and only partial activity via the rat PTH1R. Comparable results were obtained when human and rat PTH1Rs were transiently expressed heterologously in COS-7 cells or homologously in HEK 293 and UMR 106-01 cells, respectively. Binding affinities of these C-truncated peptides to human and rat PTH1Rs were concordant with their relative potencies in activating PLC. We conclude that hPTH(1-31) and, to a lesser extent, hPTH(1-30) can activate PLC, as well as AC, via both rat and human PTH1Rs. Accordingly, a role for PLC activation in the anabolic action of PTH in vivo cannot be excluded.
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PMID:Type-1 parathyroid hormone (PTH)/PTH-related peptide (PTHrP) receptors activate phospholipase C in response to carboxyl-truncated analogs of PTH(1-34). 975 12

PTH is an 84-amino acid protein. Occupancy of its cognate receptor generally results in activation of adenylyl cyclase and/or phosphoinositide-specific phospholipase Cbeta (PLCbeta). In the kidney, PTH receptors are present on proximal and distal tubule cells. In proximal tubules, PTH induces calcium signaling, typified by a transient rise in intracellular calcium ([Ca2+]i) and inositol trisphosphate formation, but does not affect calcium absorption. By contrast, in distal tubules, PTH increases calcium absorption that is associated with a slow and sustained rise in [Ca2+]i, but does not stimulate phospholipase C (PLC) or cause inositol trisphosphate accumulation. Nonetheless, stimulation of distal calcium transport requires activation of protein kinase C (PKC) and protein kinase A. We now characterize the origin of the differential effects of ligand occupancy by using synthetic human PTH analogs that preferentially activate adenylyl cyclase and/or PLCbeta. We further tested the hypothesis that phospholipase D is responsible for PKC activation in distal tubule cells. PTH-(1-31) increased [Ca2+]i in distal tubule but not in proximal tubule cells, whereas PTH-(3-34) caused a partial increase in [Ca2+]i in proximal cells, but had no effect in distal cells. PTH-(7-34) blocked increases in [Ca2+]i in distal tubule cells stimulated by PTH-(1-34) and PTH-(1-31). The PLC inhibitor U73122 abolished the PTH-induced rise in [Ca2+]i and inositol trisphosphate formation by proximal tubule cells, but had no effect on PTH-stimulated Ca2+ uptake by distal tubule cells. These results support the view that activation of PKC by PTH in distal tubule cells does not involve PLCbeta. PTH did, however, activate phospholipase D with attendant formation of diacylglycerol in distal cells. As activation of PKC is required for induction of calcium transport by PTH, we conclude that PTH receptors are capable of activating multiple phospholipases and that the structural requirements for such activation differ in proximal and distal tubule cells.
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PMID:Cell-specific signaling and structure-activity relations of parathyroid hormone analogs in mouse kidney cells. 988 39

The effects of OST-766, an inhibitor of vacuolar H+-ATPase activity, on adenylyl cyclase and phospholipase C activity were explored in the osteoblast cell line ROS 17/2.8. In fresh homogenates of ROS 17/2.8 cells, OST-766 inhibited adenylyl cyclase activity (ACA) in response to guanine nucleotide and forskolin but had no effect on basal ACA. OST-766 enhanced the basal generation of IP2, but not that formed in response to Ca2+ or guanine nucleotides. In marked contrast, incubation of intact ROS 17/2.8 cells with OST-766 for at least 48 hours resulted in an increase in basal ACA as well as in response to PTH, guanine nucleotides and forskolin. Under similar conditions, the compound also increased IP1, IP2 and IP3 generation in response to guanine nucleotides and Ca2+. Levels of the guanine nucleotide binding proteins Gs and Gi were also increased in OST-766-treated cells. The results suggest that the actions of this H+-ATPase inhibitor include effects on osteoblasts through PTH-sensitive signal transduction pathways.
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PMID:Influence of antiresorptive agent OST-766 on signal transduction pathways involved in parathyroid hormone action. 991 23

Two heterozygous PTH/PTH-related peptide (PTHrP) receptor missense mutations were previously identified in patients with Jansen's metaphyseal chondrodysplasia (JMC), a rare form of short limb dwarfism associated with hypercalcemia and normal or undetectable levels of PTH and PTHrP. Both mutations, H223R and T410P, resulted in constitutive activation of the cAMP signaling pathway and provided a plausible explanation for the abnormalities in skeletal development and mineral ion homeostasis. In the present study we analyzed genomic DNA from four additional sporadic cases with JMC to search for novel activating mutations in the PTH/PTHrP receptor, to determine the frequency of the two previously identified missense mutations, H223R and T410P, and to determine whether different mutations present with different severity of the disease. The H223R mutation was identified in three novel JMC patients and is, therefore, to date the most frequent cause of JMC. In the fourth patient, a novel heterozygous missense mutation was found that changes isoleucine 458 in the receptor's seventh membrane-spanning region to arginine (I458R). In COS-7 cells expressing the human PTH/PTHrP receptor with the I458R mutation, basal cAMP accumulation was approximately 8 times higher than that in cells expressing the wild-type receptor despite impaired surface expression of the mutant receptor. Furthermore, the I458R mutant showed higher responsiveness to PTH than the wild-type receptor in its ability to activate both downstream effectors, adenylyl cyclase and phospholipase C. Like the H223R and the T410P mutants, the I458R mutant had no detectable effect on basal inositol phosphate accumulation. Overall, the patient with the I458R mutation exhibited clinical and biochemical abnormalities similar to those in patients with the previously identified H223R and T410P mutations.
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PMID:A novel parathyroid hormone (PTH)/PTH-related peptide receptor mutation in Jansen's metaphyseal chondrodysplasia. 1048 64

The initial steps involved in mediating the transduction of PTH signal via its G protein-coupled receptors are well understood and occur through the activation of cAMP and phospholipase C pathways. However, the cellular and molecular mechanisms for subsequent receptor desensitization are less well understood. Recently, a new family of GTPase activating proteins known as regulators of G protein signaling (RGS), has been implicated in desensitization of several G protein-coupled ligand-induced processes. At present, it is not known whether any of the RGS proteins play a role in PTH signaling. Using the differential display method, we screened for genes that are selectively expressed after a single s.c. injection of human PTH (1-38) (8 microg/100 g) in osteoblast-enriched femoral metaphyseal spongiosa of young male rats (3-4 weeks old). We found and cloned one full-length complementary DNA that encodes a 211-amino acid RGS protein and shares 97% sequence identity with mouse and human RGS2. Based on sequence similarity, we have designated this clone as rat RGS2. Northern blot analysis confirmed that the expression of RGS2 messenger RNA (mRNA) is rapidly and transiently increased by human PTH (1-38) in both metaphyseal (4-to 5-fold) and diaphyseal (2- to 3-fold) bone, as well as in cultured osteoblast cultures (2- to 37-fold). In vitro, forskolin and dibutyryl cAMP similarly elevated RGS2 mRNA. In vivo, PTH analog (1-31) [which stimulates intracellular cAMP accumulation, PTHrP (1-34), and prostaglandin E2] induced RGS2 mRNA expression; whereas PTH analogs (3-34) and (7-34), which do not stimulate cAMP production, had no effect on expression. In tissue distribution analysis, RGS2 is widely expressed and was detected in all tissues examined (heart, spleen, liver, skeletal muscle, kidney, and testis), with significant expression in two nonclassical PTH-sensitive tissues: the brain, and the heart. After PTH injection, RGS2 mRNA expression was induced in rat bone but not in any of the other tissues examined. These findings demonstrate that RGS2 is regulated by PTH, prostaglandin E2, and PTHrP and that regulation by PTH in bone occurs via the cAMP pathway. Additionally, these results suggest the exciting possibility that increased RGS2 expression in osteoblasts may be one of the early events influencing PTH signaling.
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PMID:Dynamic regulation of RGS2 in bone: potential new insights into parathyroid hormone signaling mechanisms. 1061 20


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