Novel n-3 Fatty Acid Oxidation Products Activate Nrf2 by Destabilizing the Association between Keap1 and Cullin3

Abstract

Consumption of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) can mitigate the progression of diseases in which oxidative stress represents a common underlying biochemical process. Nrf2-regulated gene expression regulates detoxification of reactive oxygen species. EPA and DHA were subjected to an in vitro free radical oxidation process that models in vivo conditions. Oxidized n-3 fatty acids reacted directly with the negative regulator of Nrf2, Keap1, initiating Keap1 dissociation with Cullin3, thereby inducing Nrf2-directed gene expression. Liquid chromatography-tandem mass spectrometry analyses of oxidized EPA demonstrated the presence of novel cyclopentenone-containing molecules termed J 3 -isoprostanes in vitro and in vivo and were shown to induce Nrf2-directed gene expression. These experiments provide a biochemical basis for the hypothesis that formation of J-ring compounds generated from oxidation of EPA and DHA in vivo can reach concentrations high enough to induce Nrf2-based cellular defense systems.Eicosapentaenoic acid (EPA, C20:5n-3) 5 and docosahexaenoic acid (DHA, C22:6n-3) are two major components of fish oil. Epidemiological studies and randomized controlled trials have demonstrated that n-3 fatty acid supplementation can reduce sudden cardiac death, nonfatal stroke, and decrease progression of arteriosclerosis (1-3). Dietary supplementation with fish oil also shows promise for suppressing progression of neurodegenerative diseases (4), neuropsychiatric disorders (5), human immunodeficiency virus (6), and diseases of the retina (7). Thus, identifying the molecular mechanism(s) responsible for n-3 fatty acid mitigation of these diverse diseases represents an important and intriguing question.Oxidative stress is a common factor in the etiology of the diseases impacted by DHA and EPA (1, 8 -17). These n-3 fatty acids are very susceptible to free radical oxidation, exceeding that of arachidonic acid (AA) (18 -20). Yet, recent studies have shown that EPA and DHA supplementation reduced urinary F 2 -isoprotane levels, a marker for oxidative stress, as well as enhanced cellular antioxidant defense systems (21-24). Although a reduction of F 2 -isoprostane levels can be attributed, in part, to a decrease in membrane AA content (20,25), the relationship between EPA/ DHA oxidation versus EPA/DHA-mediated induction of antioxidant responses is not well understood.Non-enzymatic free-radical peroxidation of AA results in the formation of multiple stereoisomers of prostaglandin H 2 -like bicyclic endoperoxides that can undergo thiol-mediated reduction to form prostaglandin-like compounds termed F 2 -isoprostanes (F 2 -IsoPs) or rearrangement and reduction to form E 2 -IsoPs, D 2 -IsoPs, and isothromboxanes. E 2 /D 2 -IsoPs can then dehydrate to highly reactive A 2 /J 2 -isoprostanes containing cyclopentenone rings. These compounds readily adduct thiol groups on proteins and have been shown to induce Nrf2/ ARE-directed gene expression (26).NF-E2-related factor 2, Nrf2, is a master transcription factor ...

The marine n-3 PUFA DHA evokes cytoprotection against oxidative stress and protein misfolding by inducing autophagy and NFE2L2 in human retinal pigment epithelial cells

“…These data are consistent with previous reports demonstrating that DHA directly or indirectly induces NFE2L2. 74,75 Also, we found that DHA pretreatment protected the ARPE-19 cells from a cytostatic effect upon a subsequent challenge with hydrogen peroxide. Together, these results indicate that DHA induces activation of NFE2L2 and an increased buffer capacity for oxidative stress in retinal pigment epithelial cells.…”

“…However, these lipids serve as precursors for different types of lipid-derived signaling compounds resulting from both nonenzymatic and enzymatic reactions. 74 The ARPE-19 cells did not display any changes in cell growth or survival after adding any of the lipids (70 mM). Nevertheless, in cells where NFE2L2 was downregulated, both the basal and DHA-induced levels of ROS were increased consistent with a central role of NFE2L2 in redox balance.…”

Zerumbone Induces Heme Oxygenase-1 Expression in Mouse Skin and Cultured Murine Epidermal Cells through Activation of Nrf2

“…have shown that modification of Keap1 cysteines is insufficient to hamper the interaction between Keap1 and Nrf2 (36). Although disruption of Keap1-Nrf2 interaction does not occur on cysteine modification of Keap1, it has been suggested that Keap1 modification by an ARE inducer results in Nrf2 activation through the disruption of Keap1-Cul3 interaction, alternative to the Keap1-Nrf2 complex (35,37). Therefore, cysteine residues of Keap1 are considered to be potential targets for Nrf2 activation by zerumbone that retains electrophilic property.…”

Nrf2 activation in the treatment of neurodegenerative diseases: a focus on its role in mitochondrial bioenergetics and function

“…It has also been described that some electrophilic lipids, such as the cyclopentenone prostaglandin, 15-deoxy-Δ 12,14 -prostaglandin J 2 (which serves as a model to study the effects of lipid peroxidation products derived by the reaction of reactive oxygen/ nitrogen species with unsaturated fatty acids), are able to induce Nrf2 by modifying thiol groups of Keap1 (Levonen et al, 2004). Another mechanism by which modification of certain cysteines in Keap1 leads to diminished degradation of Nrf2 is by decreasing the affinity of Keap1 for Cul3, and subsequently diminishing Nrf2 ubiquitination (Gao et al, 2007;Eggler et al, 2009).…”