UMLS:C0043167 - FACTA Search

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Query: UMLS:C0043167 (pertussis)
19,595 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We studied PGE2 specific binding sites in human myometrial microsomes prepared from uterine specimens obtained by hysterectomy (women between 38 and 55 years of age). Competition experiments showed that the potency order for various prostaglandins (PGs) was: PGE2 > or = PGE1 >> PGF2 alpha > Iloprost > or = Carbacyclin >> ZK 110841 (PDG2 analogue). These relative affinities indicated that the receptor was of the EP type. In kinetic experiments GTP, GppNHp and GTP gamma S increased the rate of PGE2 binding (steady state was reached more rapidly in the presence of nucleotides) but maximal specific binding was not significantly different. Complete dissociation could not be obtained, even in the presence of GTP. Only 50% of maximal binding was readily dissociable. The dissociation rate was 4.56.10(-4) sec-1 (half time of about 660 sec) and in the presence of GTP analogues it was slightly increased (k-1 = 7.16 10(-4) sec-1, half time 420 sec.). Scatchard analysis of saturation curves showed an increase in ligand receptor affinity in the presence of GTP or nucleotide analogues: the Kd shifted from 9.66 +/- 2.8.10(-9) M to 4.96 +/- 1.25.10(-9) M, but the number of binding sites did not change significantly (310 +/- 37 to 350 +/- 17 fmol/mgP). The effect of GTP was observed at a concentration of 5.10(-4)M. GppNHp and GTP gamma S were effective at 1.10(-5) M. Pretreatment of myometrial membranes with pertussis or cholera toxins had no effect on PGE2 binding to membrane sites. Our conclusion is that GTP induced conversion of a population of low affinity sites into a population of higher affinity sites. This effect of guanine nucleotides was described in adipocytes and kidney medulla. Competition studies with PGE2 analogues (sulprostone, 17-phenyl-omega-trinor PGE2, M&B 28,767, misoprostol, butaprost) showed that this receptor mediates a contractile response and is probably an EP3 subtype.
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PMID:Modulation of human myometrial PGE2 receptor by GTP characterization of receptor subtype. 823 33

We recently identified four isoforms of bovine prostaglandin E receptor EP3 subtype, which are coupled to different signaling pathways; EP3A is coupled to inhibition of adenylate cyclase, while EP3B and EP3C are coupled to its stimulation and EP3D is coupled to phosphatidylinositol turnover, in addition to the adenylate cyclase system (Namba, T., Sugimoto, Y., Negishi, M., Irie, A., Ushikubi, F., Kakizuka, Ito, S., A., Ichikawa, A., and Narumiya, S. (1993) Nature 365, 166-170). We examined here the identity of coupled G proteins and their regulation by one of the isoforms, EP3C, in the membranes of EP3C cDNA-transfected Chinese hamster ovary cells. M&B 28767, an EP3 agonist, stimulated the GTPase activity in the pertussis toxin (PT)-treated cell membrane, but inhibited it in the cholera toxin (CT)-treated cell membrane, while the agonist neither stimulated nor inhibited it in the both PT- and CT-treated cell membrane. In the PT- and CT-treated cell membrane reconstituted with various G proteins, M&B 28767 inhibited the GTPase activity of G(o), but stimulated that of Gs. On the other hand, M&B 28767 did not affect the GTPase activity of Gi1, Gi2, or Gi3. M&B 28767 increased the apparent affinity of G(o) for GDP without any change in that for GTP, as assessed by displacement of [35S]GTP gamma S (guanosine 5'-O-(3-thiotriphosphate)) binding to G(o). In contrast, M&B 28767 increased the apparent affinity of Gs for GTP but decreased that for GDP. These results demonstrated that the EP3 receptor isoform is coupled to two different G proteins, and oppositely regulates their activities, inhibition of G(o), and stimulation of Gs.
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PMID:Opposite coupling of prostaglandin E receptor EP3C with Gs and G(o). Stimulation of Gs and inhibition of G(o). 825 19

Prostaglandin E2 has been reported both to stimulate glycogen-phosphorylase activity (glycogenolytic effect) and to inhibit the glucagon-stimulated glycogen-phosphorylase activity (antiglycogenolytic effect) in rat hepatocytes. It was the purpose of this study to resolve this apparent contradiction and to characterize the signalling pathways and receptor subtypes involved in the opposing prostaglandin E2 actions. Prostaglandin E2 (10 microM) increased glucose output, glycogen-phosphorylase activity and inositol trisphosphate formation in hepatocyte cell culture and/or suspension. In the same systems, prostaglandin E2 decreased the glucagon-stimulated (1 nM) glycogen-phosphorylase activity and cAMP formation. The signalling pathway leading to the glycogenolytic effect of PGE2 was interrupted by incubation of the hepatocytes with 4 beta-phorbol 12-myristate 13-acetate (100 nM) for 10 min, while the antiglycogenolytic effect of prostaglandin E2 was not attenuated. The signalling pathway leading to the antiglycogenolytic effect of prostaglandin E2 was interrupted by an incubation of cultured hepatocytes with pertussis toxin (100 ng/ml) for 18 h, whereas the glycogenolytic effect of prostaglandin E2 was enhanced. The EP1/EP3 prostaglandin-E2-receptor-specific prostaglandin E2 analogue Sulproston had a stronger glycogenolytic potency than the EP3 prostaglandin-E2-receptor-specific prostaglandin E2 analogue Misoprostol. The antiglycogenolytic potency of both agonists was equal. It is concluded that the glycogenolytic and the antiglycogenolytic effects of prostaglandin E2 are mediated via different signalling pathways in hepatocytes possibly involving EP1 and EP3 prostaglandin E2 receptors, respectively.
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PMID:Glycogenolytic and antiglycogenolytic prostaglandin E2 actions in rat hepatocytes are mediated via different signalling pathways. 828 25

Functional cDNA clones for two isoforms of the mouse prostaglandin E receptor EP3 subtype derived from alternative RNA splicing were obtained. The two isoforms are only different in the sequence of the putative cytoplasmic carboxyl-terminal tail and their hydrophobicity; one isoform, named EP3 alpha, has a hydrophilic tail, and the other, named EP3 beta, has a hydrophobic tail. When expressed, the two receptors displayed identical ligand binding properties but different responses to guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S). Without a change in the Bmax value, GTP gamma S increased Kd for prostaglandin E2 of EP3 beta and decreased that of EP3 alpha. These effects were abolished by the treatment of membranes with pertussis toxin and restored by the addition of Gi2. Although both isoforms exerted inhibition of forskolin-induced cAMP accumulation, three orders lower concentrations of agonists were required for EP3 alpha than EP3 beta for 50% inhibition of cAMP formation. A similar difference in agonist potency was observed also for agonist-induced stimulation of GTPase activity in membranes. Thus, the two receptors with different carboxyl-terminal tails show different coupling to the Gi protein, leading to the opposite responses to GTP in the ligand binding affinity and to different affinities of the agonist-occupied receptors to the G proteins.
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PMID:Two isoforms of the EP3 receptor with different carboxyl-terminal domains. Identical ligand binding properties and different coupling properties with Gi proteins. 838 13

The functional interaction of prostaglandin E (PGE) receptor EP3 subtype with GTP-binding proteins (G proteins) was characterized in the membranes prepared from mouse EP3 receptor cDNA-transfected Chinese hamster ovary cells. PGE2 inhibited forskolin-stimulated adenylate cyclase activity in CHO cells expressing EP3 receptor and this inhibition was abolished by pertussis toxin (PT) treatment. The PGE2 binding to the membranes was increased by GTP gamma S, and PT treatment also increased the binding activity to the same level as that increased by GTP gamma S, but the sensitivity of GTP gamma S was lost. Reconstitution with PT-sensitive G proteins into the ADP-ribosylated membranes reduced the PGE2 binding activity with the following preference: Gi1 = Gi2 > Gi3 > GO, but GTP gamma S completely blocked the reduction by G proteins. The G-protein-induced reduction of the binding was due to the increase in Kd without the change of Bmax, and due to suppression of association rate. [3H]PGE2-bound EP3 receptor solubilized from the ADP-ribosylated membranes in the presence or absence of GTP gamma S was eluted at the position of M(r) = approx. 60 kDa, similar to the relative molecular mass of EP3 receptor deduced from its amino acid sequence. In contrast, [3H]PGE2-bound receptor solubilized from Gi2-reconstituted membranes was eluted at the position of M(r) = approx. 130 kDa, corresponding to the M(r) of the complex of EP3 receptor and Gi2, but GTP gamma S shifted the position of its elution from M(r) = 130 to 60 kDa. Furthermore, addition of PGE2 stimulated the GDP release from G proteins reconstituted into the ADP-ribosylated membranes, and PGE2 inhibited forskolin-stimulated adenylate cyclase activity in G-protein-reconstituted membranes with a selectivity order of Gi1 = Gi2 > Gi3 > GO. These results indicate that EP3 receptor can functionally couple to PT-sensitive G proteins and unusually the complex form with G proteins has low affinity for the ligand but the form not associated with G proteins has high affinity.
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PMID:Functional interaction of prostaglandin E receptor EP3 subtype with guanine nucleotide-binding proteins, showing low-affinity ligand binding. 838 86

PGE2 is a powerful modulator of uterine contractility, but there is uncertainty as to which receptor subtypes (EP1, EP2, EP3, or EP4), G proteins, and second messenger systems are activated by PGE2 in myometrium. Here we show that in cultured human myometrial cells, PGE2 (1-100 microM) activates phospholipase C (PLC) up to 500% over the control level and elevates intracellular calcium ([Ca2+]i) from the resting level of 60-90 nM up to 350 nM in a concentration-dependent manner. Stimulation by the receptor subtype-selective analogs GR63799X (EP3), sulprostone (EP3 > EP1), and misoprostol (EP3 > EP2 > EP1) indicates that these effects are transmitted through EP3 receptors. Both effects are resistant to pertussis toxin (PT). Lower concentrations of PGE2 (1-300 nM) increase [Ca2+]i via a PT-sensitive pathway, without PLC activation. This [Ca2+]i increase occurs after an inverse dose-related delay and is inhibited by the selective EP1 antagonist AH6809 and calcium channel blockers. By comparison, oxytocin stimulates PLC up to 1000% over the control level and elevates [Ca2+]i up to 800 nM in a concentration-dependent manner without any measurable delay; both effects are partly sensitive to PT. These data provide functional evidence for the presence of different stimulatory mechanisms for PGE2 in myometrium: 1) a low affinity receptor (probably EP3D) that activates PLC through a PT-insensitive pathway; and 2) a high affinity receptor (probably EP1), independent from PLC and involving a PT-sensitive G protein (G(i)?). Both pathways lead to elevation of [Ca2+]i.
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PMID:Prostaglandin E2 activates phospholipase C and elevates intracellular calcium in cultured myometrial cells: involvement of EP1 and EP3 receptor subtypes. 864 Dec 11

1. Prostaglandin E2 (PGE2) is an autacoid that decreases proteoglycan synthesis, increases metalloprotease production by cultured chondrocytes, and can modulate some of the actions of interleukin-1 on cartilage. The objective of the present study was to characterize the subtype of prostaglandin E2 receptor present in bovine chondrocytes in culture. 2. Primary cultures of articular chondrocytes were prepared from slices of bovine carpal cartilage by sequential digestion with type III hyaluronidase, trypsin, type II collagenase, followed by overnight incubation in Dulbecco's Modified Eagle's Medium (DMEM) with type II collagenase, washing, and seeding at a density of 2 x 10(5) cells cm-2 in DMEM with 10% foetal bovine serum. 3. PGE2 and carbaprostacyclin induced dose-dependent increases in intracellular cyclic AMP in bovine chondrocytes in culture. The potencies of these compounds were different, and maximal doses of PGE2 and carbaprostacyclin had an additive effect. PGD2 induced a small increase in intracellular cyclic AMP only at a high concentration (10(-5) M). 4. PGE2 was more potent that the EP2 agonist 11-deoxy-PGE1 at inducing increases in intracellular cyclic AMP. The EP2 agonist butaprost, however, induced only a small increase at a concentration of 10(-5)M. 17-Phenyl-PGE2 (EP1 agonist), sulprostone and MB 28767 (15S-hydroxy-9-oxo-16-phenoxy-omega-tetranorprost-13E-enoic acid) (EP3 agonists) did not induce an increase in intracellular cyclic AMP at concentrations up to 10(-5)M. 5. The EP4 antagonist AH 23848B ([1 alpha(Z),2 beta, 5 alpha]-(+/-) -7-[5-[[(1,1'-biphenyl)-4-yl]methoxyl-2-(4-morpholinyl) -3-oxocyclopentyl]-5-heptenoic acid) antagonized PGE2 but not carbaprostacyclin effects on intracellular cyclic AMP. The Schild plot slope was different from 1 but this could be due to an interaction of PGE2 with IP receptors in high doses. The exact nature of the antagonism by compound AH 23848B could not be definitely established in these experimental conditions. 6. Neither PGE2 nor any of its analogues inhibited the increase in intracellular cyclic AMP induced by forskolin, and pertussis toxin did not alter the response to PGE2, suggesting that no Gi-coupled PGE2 receptors are present in these cells. Stimulation with PGE2 did not induce significant increases in intracellular inositol-trisphosphate levels nor increases in intracellular free calcium as determined by confocal microscopy, suggesting the absence of phospholipase-C-coupled or of calcium channel-coupled PGE2 receptors in bovine chondrocytes in these experimental conditions. 7. These results show for the first time that bovine chondrocytes in culture present a functional PGE2 receptor that has some pharmacological characteristics of an EP4 subtype, as well as an IP receptor.
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PMID:Characterization of the PGE2 receptor subtype in bovine chondrocytes in culture. 884 20

Prostaglandin E receptor EP3 subtype is widely distributed in the nervous system and is specifically localized to neurons, suggesting that the EP3 receptor plays important roles in the nervous system. We established a PC12 cell line that stably expresses the EP3B receptor isoform isolated from bovine adrenal chromaffin cells and examined the effect of agonist stimulation on the neuronal morphology of the PC12 cells. In the differentiated cells, M&B28767, an EP3 agonist, caused neurite retraction in a pertussis toxin-insensitive manner. 12-O-Tetradecanoylphorbol-13-acetate (TPA) also induced neurite retraction. However, when protein kinase C was down-regulated by long term exposure to TPA, TPA failed to induce neurite retraction, while the EP3B receptor-mediated retraction occurred normally. Clostridium botulinum C3 exoenzyme completely inhibited both EP3 agonist- and TPA-induced neurite retraction when microinjected into the cells, indicating that the morphological effect of the EP3B receptor is dependent on Rho activity. Thus, the activation of the EP3B receptor induced neurite retraction through a protein kinase C-independent Rho-activation pathway.
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PMID:Prostaglandin E receptor EP3 subtype induces neurite retraction via small GTPase Rho. 893 15

We characterized the proliferative action of prostaglandins (PGs) in relation to their membrane receptors on rat hepatocytes in primary culture. PGs in the order 16,16-dimethyl PGE2 > PGE2 > PGF2alpha >> PGD2 augmented epidermal growth factor (EGF)/insulin-induced DNA synthesis, assessed by [(3)H]thymidine incorporation, in a concentration-dependent manner, whereas PGs alone did not stimulate basal DNA synthesis without EGF and insulin. The cells exhibited [(3)H]PGE2 binding sites that were displaced by unlabeled PGs in the order PGE1 = PGE2 > PGF2alpha > PGD2. PGE2 inhibited glucagon-stimulated adenosine 3',5'-cyclic monophosphate (cAMP) accumulation concentration dependently. The mean effective concentration for DNA synthesis, median inhibitory concentration for cAMP accumulation, and dissociation constant for [(3)H]PGE2 binding at 25 degrees C were almost identical (approximately 70 nM). Treatment of the cells with pertussis toxin (100 ng/ml), which ADP-ribosylated most of the 41-kDa substrate, abolished the proliferative effects of PGs. We detected the expression of mRNA of the EP3 subtype PGE2 receptor using reverse transcription-polymerase chain reaction. Moreover, an EP3 agonist, enprostil, but not the EP1 agonist 17-phenyl-trinor-PGE2 or the EP2/EP4 agonist 11-deoxy-PGE1, stimulated EGF/insulin-induced DNA synthesis. These results indicate that PGs act as comitogenic growth factors through the EP3 subtype PGE2 receptor coupled with G(i) protein in cultured rat hepatocytes.
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PMID:Prostaglandins induce proliferation of rat hepatocytes through a prostaglandin E2 receptor EP3 subtype. 912 80

1. The human EP3 prostaglandin receptor is a seven transmembrane, G protein-coupled receptor that couples to inhibition of adenylyl cyclase. The receptor occurs as at least six isoforms which result from alternative splicing. The isoforms are identical over the first 359 amino acids, comprising the seven transmembrane helices, but differ in the carboxyl terminal tail which ranges in length from 6 to 65 amino acids beyond the common region. 2. We have stably expressed in CHO-K1 cells four of the isoforms (EP3I-EP3IV) and a form of the EP3 receptor (T-359) truncated at the carboxyl-terminal region defined by the alternative splicing site at amino acid number 359. 3. Isoforms EP3I and EP3II showed concentration-dependent inhibition of forskolin-stimulated adenylyl cyclase in CHO-K1 cells by the EP3 receptor agonist, sulprostone. The IC50 calculated for sulprostone inhibition was 0.2 nM for EP3I and 0.15 nM for EP3II. The maximum extent of inhibition was 80% for both isoforms. 4. Isoforms EP3III and EP3IV showed marked constitutive activity, inhibiting forskolin-stimulated adenylyl cyclase in the absence of agonist. EP3IV also displayed some agonist-dependent inhibition whereas EP3III was fully constitutively active. 5. The truncated receptor T-359 was fully constitutively active, inhibiting forskolin-stimulated adenylyl cyclase by about 70% in the absence of agonist, and showed no agonist-dependent inhibition, in agreement with a similar truncation of the mouse EP3 receptor. 6. To confirm that differences in cyclic AMP level between isoforms represent constitutive activity, we treated cells with pertussis toxin for 6 h to abolish Gi function. Pertussis toxin reversed sulprostone-mediated inhibition of cyclic AMP formation in EP3I and EP3II and abolished constitutive activity of EP3III, EP3IV and T-359 so that the level of forskolin-stimulated cyclic AMP produced was the same in all cells and similar to that obtained in mock-transfected cells. In mock-transfected cells, sulprostone had no effect on forskolin-stimulated cyclic AMP formation. 7. For these experiments we chose clones that showed similar expression levels of each isoform, as determined by binding of [3H]-prostaglandin E2 (PGE2) (EP3I, 0.71; EP3II, 1.47; EP3IV, 1.59 pmol mg-1 protein). Mock-transfected cells showed no detectable binding of [3H]-PGE2. In addition, we performed a detailed study of the effects of expression level on constitutive activity. Over a six fold range of expression there was no change in the properties of each isoform with regard to whether it was constitutively active or not. 8. The degree of constitutive activity correlated with the inverse of the length of the C-terminal tail of the isoforms. However, no correlation was found between isoforms from human and mouse: whereas EP3II shows no constitutive activity, its mouse homologue, EP3 gamma, shows almost complete constitutive activity, even though the C-terminal domains of the receptors following the splice site differ in only 7 of 29 amino acids.
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PMID:Constitutive activity of human prostaglandin E receptor EP3 isoforms. 915 43

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