Nrf2-mediated Induction of Cytoprotective Enzymes by 15-Deoxy-Δ12,14-Prostaglandin J2 Is Attenuated by Alkenal/one Oxidoreductase

Abstract

NADPH-dependent alkenal/one oxidoreductase (Aor) was discovered to be highly inducible in rat liver following treatment with the cancer chemopreventive agent 3H-1, 2-dithiole-3-thione. Aor was further characterized as an Nrf2-regulated antioxidative enzyme that reduces carbon-carbon double bonds in a variety of ␣, ␤-unsaturated aldehydes and ketones. 15-Deoxy-⌬ 12,14 -prostaglandin J 2 (15d-PGJ 2 ) is a reactive membrane lipid metabolite that activates multiple pathways, including Nrf2-mediated induction of cytoprotective enzymes. Physiologically, it is postulated that 15d-PGJ 2 alkylates key regulatory proteins via the electrophilic carbon centers found in two ␣, ␤-unsaturated ketone moieties. This current study addresses the metabolism of 15d-PGJ 2 by rat Aor (rAor) and subsequent deactivation of the Nrf2 signaling pathway by both rat and human AOR. We demonstrate that induction of NADPHdependent quinone oxidoreductase activity by 15d-PGJ 2 is markedly attenuated in mouse embryonic fibroblasts that overexpress rAor. Luciferase reporter assay and quantitative real-time PCR confirmed these findings. Concentrations required for doubling the NADPH-dependent quinone oxidoreductase response are increased from 1.8 M in wild-type to >10 M in rat Aor transgenic fibroblasts. 15d-PGJ 2 is metabolized by recombinant rAor with a K m of 9.6 M and k cat of 18.5 min ؊1. The major product is 12,13-dihydro-15-deoxy-⌬ 12,14 -prostaglandin J 2 (dihydro-15d-PGJ 2 ). The reduction of C‫؍‬C by Aor yielding dihydro-15d-PGJ 2 abolishes the inducibility in an antioxidant response element-driven luciferase assay. Collectively, these results demonstrate that 15d-PGJ 2 can be catabolized by Aor, thereby attenuating subsequent Nrf2 signaling and possibly inflammatory and apoptotic processes also influenced by 15d-PGJ 2 . Prostaglandins (PG)2 are a subfamily of eicosanoids shown to regulate a variety of physiological processes, including growth, differentiation, vascular constriction, inflammation, and homeostasis. Prostaglandins are primarily derived from arachidonic acid following phospholipase-catalyzed release from membrane lipids. Arachidonic acid is converted by cyclooxygenases into PGH 2 , which is further metabolized into PGD 2 , PGE 2 , prostacyclin, or thromboxane via various prostaglandin synthases. PGD 2 can undergo dehydration reactions to yield the J 2 series of prostaglandins, including PGJ 2 , ⌬ 12,14 -PGJ 2 , and 15d-PGJ 2 (1). The J 2 series of prostaglandins influences multiple signaling pathways by covalently binding with key signaling molecules (2-6). Among them, 15d-PGJ 2 has displayed highest potency as an inducer of gene expression. Discovered as an agonist for peroxisome-proliferator-activated receptor ␥ (PPAR␥), 15d-PGJ 2 was shown to modulate the expression of genes containing a peroxisome-proliferator response element in the promoter region, resulting in adipocyte differentiation (7). In addition, 15d-PGJ 2 contains two ␣,␤-unsaturated ketone moieties in tandem within its structure. These moieties covalently modi...

Modulation of Nitro-fatty Acid Signaling

“…PtGR-1 is the product of the DIG-1 (dithiolethione-inducible gene-1) gene and is identical to the leukotriene-B 4 dehydrogenase that catalyzes the oxidation of leukotriene B 4 (LTB 4 ) to 12-oxo-LTB 4 in humans and rats (37). PtGR-1 is broadly distributed in mammalian tissues and also functions as a NADP(H)-dependent oxidoreductase in the inactivation of 15-oxo-prostaglandin-E 2 and other ␣,␤-unsaturated carbon-containing molecules, motivating the more inclusive name alkenal/one oxidoreductase (40,44,45,63). PtGR-1 reduces fatty acid nitroalkenes to nonelectrophilic nitroalkanes, as evidenced by MS analysis and a lack of reactivity of products with ␤-mercaptoethanol.…”

“…Cells were incubated at 37°C with 95% air and 5% carbon dioxide. To generate the pCMV-3Tag-hPtGR-1, the hPtGR-1 cDNA was inserted into pCMV-3Tag-1 (Stratagene) such that it was tagged in-frame with a 3ϫ FLAG epitope (40). HEK293T cells were plated in 100-mm dishes at 60 -70% cell density and allowed to attach overnight.…”

Up-Regulation of Human Prostaglandin Reductase 1 Improves the Efficacy of Hydroxymethylacylfulvene, an Antitumor Chemotherapeutic Agent

“…PTGR1 was originally identified as leukotriene B4 dehydrogenase (Yokomizo et al, 1993), and it transforms prostaglandins by enone reduction (Tai et al, 2002;Yu et al, 2006). PTGR1 was later discovered to be strongly induced by 1,2-dithiol-3-thione (D3T) in rat liver by nuclear factor E2-related factor 2 (Nrf2)-mediated gene activation (Primiano et al, 1996;Dick et al, 2001) and further characterized as alkenal/one oxidoreductase for its capacity to reduce endogenous and exogenous ␣,␤-unsaturated aldehydes and ketones (Dick et al, 2001;Gong et al, 2006Gong et al, , 2007Yu et al, 2006;Neels et al, 2007).…”

Proteomic studies on protein modification by cyclopentenone prostaglandins: Expanding our view on electrophile actions

“…In addition, the cyPG 15d-PGJ 2 has been reported to be a substrate for alkenal reductase, and this attenuates its potential to induce a cytoprotective response [15]. Therefore, the levels of GSH and the expression of detoxifying enzymes will be important determinants of the biological effects of cyPG in cells.…”