Upregulation of orphan nuclear receptor Nur77 following PGF_2α, Bimatoprost, and Butaprost treatments. Essential role of a protein kinase C pathway involved in EP₂ receptor activated Nur77 gene transcription

Abstract

Using gene chip technology, we first identified that PGF2α (FP agonist) and Butaprost (EP2 agonist) induced about a five‐fold upregulation of Nur77 mRNA expression in hFP‐HEK 293/EBNA and hEP2‐HEK293/EBNA cells. Northern Blot analysis revealed that PGF2α‐ and Butaprost‐induced upregulation of Nur77 expression are dose‐ and time‐dependent. Both PGF2α and Butaprost upregulated Nur77 gene expression through the protein kinase C (PKC) pathway. These data are the first showing a link between EP2 receptor stimulation and protein kinase C activation. Calcineurin was found to be involved downstream of the PKC pathway in PGF2α‐induced Nur77 expression, but not in Butaprost‐induced Nur77 expression. We also used Nur77 as a marker gene to compare the effects of PGF2α, Butaprost, and Bimatoprost (a prostamide) on Nur77 expression in human primary trabecular meshwork and ciliary smooth muscle (SM) cells, which are target cells for antiglaucoma drugs. The results showed that PGF2α and Butaprost, but not Bimatoprost, induced upregulation of Nur77 expression in human TM cells. PGF2α, but not Bimatoprost, dramatically induced upregulation of Nur77 mRNA expression in human ciliary SM cells, whereas Butaprost slightly upregulated Nur77 mRNA expression in SM cells. Nur77 promoter deletion analysis indicated that PGF2α, but not Bimatoprost, activated Nur77 promoter‐luciferase reporter in hFP‐HEK 293/EBNA cells. Butaprost was less efficacious in inducing Nur77 promoter‐luciferase reporter activity in hEP2‐HEK293/EBNA cells relative to PGF2α in the comparable assay. The data for Nur77 promoter functional analysis were matched to the Northern blot analysis. It appears that PGF2α and Butaprost activate Nur77 transcription mechanisms through the activation of FP and EP2 receptor‐coupled signaling pathways, whereas Bimatoprost stimulates neither FP nor EP2 receptors. British Journal of Pharmacology (2004) 142, 737–748. doi:10.1038/sj.bjp.0705829

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