Phosphorylation of glycogen synthase kinase-3 and stimulation of T-cell factor signaling following activation of EP2 and EP4 prostanoid receptors by prostaglandin E2

Hiromichi Fujino, Kimberly A. West, John W Regan

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237 Citations (Scopus)

Abstract

Recently we have shown that the FPB prostanoid receptor, a G-protein-coupled receptor that couples to Gαq, activates T-cell factor (Tcf)/lymphoid enhancer factor (Lef)-mediated transcriptional activation (Fujino, H., and Regan, J. W. (2001) J. Biol. Chem. 276, 12489-12492). We now report that the EP2 and EP4 prostanoid receptors, which couple to Gαs, also activate Tcf/Lef signaling. By using a Tcf/Lef-responsive luciferase reporter gene, transcriptional activity was stimulated ∼10-fold over basal by 1 h of treatment with prostaglandin E2 (PGE2) in HEK cells that were stably transfected with the human EP2 and EP4 receptors. This stimulation of reporter gene activity was accompanied by a PGE2-dependent increase in the phosphorylation of both glycogen synthase kinase-3 (GSK-3) and Akt kinase. H-89, an inhibitor of protein kinase A (PKA), completely blocked the agonist-dependent phosphorylation of GSK-3 in both EP2- and EP4-expressing cells. However, H-89 pretreatment only blocked PGE2-stimulated Lef/Tcf reporter gene activity by 20% in EP4-expressing cells compared with 65% inhibition in EP2-expressing cells. On the other hand wortmannin, an inhibitor of phosphatidylinositol 3-kinase, had the opposite effect and inhibited PGE2-stimulated reporter gene activity to a much greater extent in EP4-expressing cells as compared with EP2-expressing cells. These findings indicate that the activation of Tcf/Lef signaling by EP2 receptors occurs primarily through a PKA-dependent pathway, whereas EP4 receptors activate Tcf/Lef signaling mainly through a phosphatidylinositol 3-kinase-dependent pathway. This is the first indication of a fundamental difference in the signaling potential of EP2 and EP4 prostanoid receptors.

Original languageEnglish (US)
Pages (from-to)2614-2619
Number of pages6
JournalJournal of Biological Chemistry
Volume277
Issue number4
DOIs
StatePublished - Jan 25 2002

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Receptors, Prostaglandin E, EP2 Subtype
Receptors, Prostaglandin E, EP4 Subtype
TCF Transcription Factors
Glycogen Synthase Kinase 3
Phosphorylation
Dinoprostone
Chemical activation
Reporter Genes
Genes
Phosphatidylinositol 3-Kinase
Cyclic AMP-Dependent Protein Kinases
G-Protein-Coupled Receptors

ASJC Scopus subject areas

  • Biochemistry

Cite this

@article{a304f4c0e7934e7d9e33f89c3f65feed,
title = "Phosphorylation of glycogen synthase kinase-3 and stimulation of T-cell factor signaling following activation of EP2 and EP4 prostanoid receptors by prostaglandin E2",
abstract = "Recently we have shown that the FPB prostanoid receptor, a G-protein-coupled receptor that couples to Gαq, activates T-cell factor (Tcf)/lymphoid enhancer factor (Lef)-mediated transcriptional activation (Fujino, H., and Regan, J. W. (2001) J. Biol. Chem. 276, 12489-12492). We now report that the EP2 and EP4 prostanoid receptors, which couple to Gαs, also activate Tcf/Lef signaling. By using a Tcf/Lef-responsive luciferase reporter gene, transcriptional activity was stimulated ∼10-fold over basal by 1 h of treatment with prostaglandin E2 (PGE2) in HEK cells that were stably transfected with the human EP2 and EP4 receptors. This stimulation of reporter gene activity was accompanied by a PGE2-dependent increase in the phosphorylation of both glycogen synthase kinase-3 (GSK-3) and Akt kinase. H-89, an inhibitor of protein kinase A (PKA), completely blocked the agonist-dependent phosphorylation of GSK-3 in both EP2- and EP4-expressing cells. However, H-89 pretreatment only blocked PGE2-stimulated Lef/Tcf reporter gene activity by 20{\%} in EP4-expressing cells compared with 65{\%} inhibition in EP2-expressing cells. On the other hand wortmannin, an inhibitor of phosphatidylinositol 3-kinase, had the opposite effect and inhibited PGE2-stimulated reporter gene activity to a much greater extent in EP4-expressing cells as compared with EP2-expressing cells. These findings indicate that the activation of Tcf/Lef signaling by EP2 receptors occurs primarily through a PKA-dependent pathway, whereas EP4 receptors activate Tcf/Lef signaling mainly through a phosphatidylinositol 3-kinase-dependent pathway. This is the first indication of a fundamental difference in the signaling potential of EP2 and EP4 prostanoid receptors.",
author = "Hiromichi Fujino and West, {Kimberly A.} and Regan, {John W}",
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T1 - Phosphorylation of glycogen synthase kinase-3 and stimulation of T-cell factor signaling following activation of EP2 and EP4 prostanoid receptors by prostaglandin E2

AU - Fujino, Hiromichi

AU - West, Kimberly A.

AU - Regan, John W

PY - 2002/1/25

Y1 - 2002/1/25

N2 - Recently we have shown that the FPB prostanoid receptor, a G-protein-coupled receptor that couples to Gαq, activates T-cell factor (Tcf)/lymphoid enhancer factor (Lef)-mediated transcriptional activation (Fujino, H., and Regan, J. W. (2001) J. Biol. Chem. 276, 12489-12492). We now report that the EP2 and EP4 prostanoid receptors, which couple to Gαs, also activate Tcf/Lef signaling. By using a Tcf/Lef-responsive luciferase reporter gene, transcriptional activity was stimulated ∼10-fold over basal by 1 h of treatment with prostaglandin E2 (PGE2) in HEK cells that were stably transfected with the human EP2 and EP4 receptors. This stimulation of reporter gene activity was accompanied by a PGE2-dependent increase in the phosphorylation of both glycogen synthase kinase-3 (GSK-3) and Akt kinase. H-89, an inhibitor of protein kinase A (PKA), completely blocked the agonist-dependent phosphorylation of GSK-3 in both EP2- and EP4-expressing cells. However, H-89 pretreatment only blocked PGE2-stimulated Lef/Tcf reporter gene activity by 20% in EP4-expressing cells compared with 65% inhibition in EP2-expressing cells. On the other hand wortmannin, an inhibitor of phosphatidylinositol 3-kinase, had the opposite effect and inhibited PGE2-stimulated reporter gene activity to a much greater extent in EP4-expressing cells as compared with EP2-expressing cells. These findings indicate that the activation of Tcf/Lef signaling by EP2 receptors occurs primarily through a PKA-dependent pathway, whereas EP4 receptors activate Tcf/Lef signaling mainly through a phosphatidylinositol 3-kinase-dependent pathway. This is the first indication of a fundamental difference in the signaling potential of EP2 and EP4 prostanoid receptors.

AB - Recently we have shown that the FPB prostanoid receptor, a G-protein-coupled receptor that couples to Gαq, activates T-cell factor (Tcf)/lymphoid enhancer factor (Lef)-mediated transcriptional activation (Fujino, H., and Regan, J. W. (2001) J. Biol. Chem. 276, 12489-12492). We now report that the EP2 and EP4 prostanoid receptors, which couple to Gαs, also activate Tcf/Lef signaling. By using a Tcf/Lef-responsive luciferase reporter gene, transcriptional activity was stimulated ∼10-fold over basal by 1 h of treatment with prostaglandin E2 (PGE2) in HEK cells that were stably transfected with the human EP2 and EP4 receptors. This stimulation of reporter gene activity was accompanied by a PGE2-dependent increase in the phosphorylation of both glycogen synthase kinase-3 (GSK-3) and Akt kinase. H-89, an inhibitor of protein kinase A (PKA), completely blocked the agonist-dependent phosphorylation of GSK-3 in both EP2- and EP4-expressing cells. However, H-89 pretreatment only blocked PGE2-stimulated Lef/Tcf reporter gene activity by 20% in EP4-expressing cells compared with 65% inhibition in EP2-expressing cells. On the other hand wortmannin, an inhibitor of phosphatidylinositol 3-kinase, had the opposite effect and inhibited PGE2-stimulated reporter gene activity to a much greater extent in EP4-expressing cells as compared with EP2-expressing cells. These findings indicate that the activation of Tcf/Lef signaling by EP2 receptors occurs primarily through a PKA-dependent pathway, whereas EP4 receptors activate Tcf/Lef signaling mainly through a phosphatidylinositol 3-kinase-dependent pathway. This is the first indication of a fundamental difference in the signaling potential of EP2 and EP4 prostanoid receptors.

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