Vascular endothelial growth factor induction by prostaglandin E2 in human airway smooth muscle cells is mediated by E prostanoid EP 2/EP4 receptors and SP-1 transcription factor binding sites

Bradbury, Dawn; Clarke, Deborah; Seedhouse, Claire; Corbettt, Lisa; Stocks, Joanne; Knox, Alan

Abstract

Prostaglandin E2 (PGE2) can increase vascular endothelial growth factor A (VEGF-A) production but the mechanisms involved are unclear. Here we characterized the transcriptional mechanisms involved in human airway smooth muscle cells (HASMC). PGE2 increased VEGF-A mRNA and protein but not mRNA stability. PGE2 stimulated the activity of a transiently transfected 2068-bp (-2018 to +50) VEGF-A promoter-driven luciferase construct. Functional 5′ deletional analysis mapped the PGE2 response element to the 135-bp sequence (-85/+50) of the human VEGF-A promoter. PGE2-induced luciferase activity was reduced in cells transfected with a 135-bp VEGF promoter fragment containing mutated Sp-1 binding sites but not in cells transfected with a construct containing mutated EGR-1 binding sites. Electrophoretic mobility shift assay and chromatin immunoprecipitation assay confirmed binding of Sp-1 to the VEGF promoter. PGE2 increased phosphorylation of Sp-1 and luciferase activity of a transfected Sp-1 reporter construct. PGE2 receptor agonists EP2 (ONO-AE1 259) and EP4 (ONO-AE1 329) mimicked the effect of PGE2, and reverse transcription-PCR, Western blotting, and flow cytometry confirmed the presence of EP2 and EP4 receptors. VEGF protein release and Sp-1 reporter activity were increased by forskolin and isoproterenol, which increase cytosolic cAMP, and the cAMP analogue, 8-bromoadenosine-3′,5′-cyclophosphoric acid. These studies suggest that PGE2 increases VEGF transcriptionally and involves the Sp-1 binding site via a cAMP-dependent mechanism involving EP2 and EP4 receptors. © 2005 by The American Society for Biochemistry and Molecular Biology, Inc.

Citation

Bradbury, D., Clarke, D., Seedhouse, C., Corbettt, L., Stocks, J., & Knox, A. (2005). Vascular endothelial growth factor induction by prostaglandin E2 in human airway smooth muscle cells is mediated by E prostanoid EP 2/EP4 receptors and SP-1 transcription factor binding sites. Journal of Biological Chemistry, 280(34), 29993-30000. https://doi.org/10.1074/jbc.M414530200

• Journal Article Type: Article
• Acceptance Date: May 19, 2005
• Online Publication Date: Aug 26, 2005
• Publication Date: Aug 26, 2005
• Deposit Date: Oct 21, 2022
• Publicly Available Date: Oct 24, 2022
• Journal: Journal of Biological Chemistry
• Print ISSN: 0021-9258
• Electronic ISSN: 1083-351X
• Publisher: American Society for Biochemistry and Molecular Biology
• Peer Reviewed: Peer Reviewed
• Volume: 280
• Issue: 34
• Pages: 29993-30000
• DOI: https://doi.org/10.1074/jbc.M414530200
• Public URL: https://nottingham-repository.worktribe.com/output/3128739
• Publisher URL: https://www.jbc.org/article/S0021-9258(20)79419-6/fulltext