The prostaglandin E2 receptor, EP2, stimulates keratinocyte proliferation in mouse skin by G protein-dependent and {beta}-arrestin1-dependent signaling pathways.
The prostaglandin E(2) (PGE(2)) G protein-coupled receptor (GPCR), EP2, plays important roles in mouse skin tumor development (Chun, K. S., Lao, H. C., Trempus, C. S., Okada, M., and Langenbach, R. (2009) Carcinogenesis 30, 1620-1627). Because keratinocyte proliferation is essential for skin tumor development, EP2-mediated signaling pathways that contribute to keratinocyte proliferation were investigated. A single topical application of the EP2 agonist, butaprost, dose-dependently increased keratinocyte replication via activation of epidermal growth factor receptor (EGFR) and PKA signaling. Because GPCR-mediated activation of EGFR can involve the formation of a GPCR-β-arrestin-Src signaling complex, the possibility of a β-arrestin1-Src complex contributing to EP2-mediated signaling in keratinocytes was investigated. Butaprost induced β-arrestin1-Src complex formation and increased both Src and EGFR activation. A role for β-arrestin1 in EP2-mediated Src and EGFR activation was demonstrated by the observation that β-arrestin1 deficiency significantly reduced Src and EGFR activation. In agreement with a β-arrestin1-Src complex contributing to EGFR activation, Src and EGFR inhibition (PP2 and AG1478, respectively) indicated that Src was upstream of EGFR. Butaprost also induced the activation of Akt, ERK1/2, and STAT3, and both β-arrestin1 deficiency and EGFR inhibition (AG1478 or gefitinib) decreased their activation. In addition to β-arrestin1-dependent EGFR activation, butaprost increased PKA activation, as measured by phospho-GSK3β (p-GSK3β) and p-cAMP-response element-binding protein formation. PKA inhibition (H89 or R(P)-adenosine-3',5'-cyclic monophosphorothioate (R(P)-cAMPS)) decreased butaprost-induced cAMP-response element-binding protein and ERK activation but did not affect EGFR activation, whereas β-arrestin1 deficiency decreased EGFR activation but did not affect butaprost-induced PKA activation, thus indicating that they were independent EP2-mediated pathways. Therefore, the results indicate that EP2 contributed to mouse keratinocyte proliferation by G protein-independent, β-arrestin1-dependent activation of EGFR and G protein-dependent activation of PKA.
Laboratory of Toxicology and Pharmacology, NIEHS, National Institutes of Health, Research Triangle Park, North Carolina 27709, USA.
Kyung-Soo
KS
Chun
Huei-Chen
HC
Lao
Robert
R
Langenbach
D000527:Alprostadil / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D019390:Arrestins / Q000235:genetics / Q000378:metabolism*; D049109:Cell Proliferation; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects / Q000235:genetics; D066246:ErbB Receptors / Q000235:genetics / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D000077156:Gefitinib; D038362:Glycogen Synthase Kinase 3 / Q000235:genetics / Q000378:metabolism; D000071679:Glycogen Synthase Kinase 3 beta; D015603:Keratinocytes / Q000166:cytology / Q000378:metabolism*; D051379:Mice; D048052:Mitogen-Activated Protein Kinase 3 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D046912:Multiprotein Complexes / Q000235:genetics / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D051057:Proto-Oncogene Proteins c-akt / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D011799:Quinazolines / Q000494:pharmacology; D058307:Receptors, Prostaglandin E, EP2 Subtype / Q000235:genetics / Q000378:metabolism*; D050796:STAT3 Transcription Factor / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D012867:Skin / Q000378:metabolism*; D020032:Tyrphostins / Q000494:pharmacology; D000071557:beta-Arrestins; D019061:src-Family Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism
D019390:Arrestins; D046912:Multiprotein Complexes; D047428:Protein Kinase Inhibitors; D011799:Quinazolines; D058307:Receptors, Prostaglandin E, EP2 Subtype; D050796:STAT3 Transcription Factor; C494087:Stat3 protein, mouse; D020032:Tyrphostins; D000071557:beta-Arrestins; C101044:RTKI cpd; C541485:EGFR protein, mouse; D066246:ErbB Receptors; D019061:src-Family Kinases; D000071679:Glycogen Synthase Kinase 3 beta; C000605507:Gsk3b protein, mouse; D051057:Proto-Oncogene Proteins c-akt; D017868:Cyclic AMP-Dependent Protein Kinases; D048052:Mitogen-Activated Protein Kinase 3; D038362:Glycogen Synthase Kinase 3; D019204:GTP-Binding Proteins; D000527:Alprostadil; C048491:butaprost; D000077156:Gefitinib