Abstract
The EP1 prostanoid receptor is one of four subtypes whose cognate physiological ligand is prostaglandin-E2 (PGE2). It is in the family of G-protein-coupled receptors and is known to activate Ca2+ signaling, although relatively little is known about other aspects of E-type prostanoid receptor (EP) 1 receptor signaling. In human embryonic kidney (HEK) cells expressing human EP1 receptors, we now show that PGE2 stimulation of the EP1 receptor up-regulates the expression of hypoxia-inducible factor-1α (HIF-1α), which can be completely blocked by pertussis toxin, indicating coupling to Gi/o. This up-regulation of HIF-1α occurs under normoxic conditions and could be inhibited with wortmannin, Akt inhibitor, and rapamycin, consistent with the activation of a phosphoinositide-3 kinase/Akt/mammalian target of rapamycin (mTOR) signaling pathway, respectively. In contrast to the hypoxia-induced up-regulation of HIF-1α, which involves decreased protein degradation, the up-regulation of HIF-1α by the EP1 receptor was associated with the phosphorylation of ribosomal protein S6 (rpS6), suggesting activation of the ribosomal S6 kinases and increased translation. Stimulation of endogenous EP1 receptors in human HepG2 hepatocellular carcinoma cells recapitulated the normoxic up-regulation of HIF-1α observed in HEK cells, was sensitive to pertussis toxin, and involved the activation of mTOR signaling and phosphorylation of rpS6. In addition, treatment of HepG2 cells with sulprostone, an EP1-selective agonist, up-regulated the mRNA expression of vascular endothelial growth factor-C, a HIF-regulated gene. HIF-1α is known to promote tumor growth and metastasis and is often up-regulated in cancer. Our findings provide a potential mechanism by which increased PGE2 biosynthesis could up-regulate the expression of HIF-1α and promote tumorigenesis.
Footnotes
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This work was supported by the National Institutes of Health National Eye Institute [Grant EY11291] (to J.W.R.); by Allergan Inc.; and by the University of Arizona College of Pharmacy. Portions of this work were also made possible by grants to the Arizona Cancer Center Genomics Facility Core from the National Institutes of Health National Institute of Environmental Health Sciences [Grant 5P30-ES06694-14900209].
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Article, publication date, and citation information can be found at http://molpharm.aspetjournals.org.
doi:10.1124/mol.110.063933.
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ABBREVIATIONS:
- EP
- E-type prostanoid receptor
- PGE2
- prostaglandin E2
- SC-19220
- 8-chloro-dibenz[b,f][1,4]oxazepine-10(11H)-carboxy-(2-acetyl)hydrazide
- PI3K
- phosphatidylinositol 3-kinase
- PLC
- phospholipase C
- U73122
- 1-[6-[[(17β)-3-methoxyestra-1,3,5(10)-trien-17-yl] amino]hexyl]-1H-pyrrole-2,5-dione
- HIF-1α
- hypoxia-inducible factor-1α
- HEK
- human embryonic kidney
- CMV
- cytomegalovirus
- GAPDH
- glyceraldehyde-3-phosphate dehydrogenase
- PKG1
- phosphoglycerate kinase 1
- CTGF
- connective tissue growth factor
- GLUT1
- facilitated glucose transporter member 1
- VEGF-A
- vascular endothelial growth factor A
- VEGF-C
- lymphatic vascular endothelial growth factor C
- EPO
- erythropoietin.
- EBNA
- Epstein Barr nuclear antigen
- PCR
- polymerase chain reaction
- IP
- inositol phosphate
- HEK-hEP1
- HEK cells expressing the human EP1 receptor
- HRE
- HIF response element
- MG132
- N-benzoyloxycarbonyl (Z)-Leu-Leu-leucinal
- AKT
- protein kinase B
- mTOR
- mammalian target of rapamycin
- BAPTA
- 1,2-bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid
- AM
- acetoxymethyl ester
- PKC
- protein kinase C
- BIM
- bisindolylmaleimide I
- rpS6
- ribosomal protein S6
- EGR-1
- early growth response factor-1
- COX-2
- cyclooxygenase-2
- HCC
- hepatocellular carcinoma
- ANOVA
- analysis of variance.
- Received February 4, 2010.
- Accepted March 15, 2010.
- Copyright © 2010 The American Society for Pharmacology and Experimental Therapeutics
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