title: Cap-independent Nrf2 translation is part of a lipoic acid-stimulated detoxification stress response.
abstract: Little is known about either the basal or stimulated homeostatic mechanisms regulating nuclear tenure of Nf-e2-related factor 2 (Nrf2), a transcription factor that mediates expression of over 200 detoxification genes. Our data show that stress-induced nuclear Nrf2 accumulation is largely from de novo protein synthesis, rather than translocation from a pre-existing cytoplasmic pool. HepG2 cells were used to monitor nuclear Nrf2 24h following treatment with the dithiol micronutrient (R)-α-lipoic acid (LA; 50μM), or vehicle. LA caused a ≥2.5-fold increase in nuclear Nrf2 within 1h. However, pretreating cells with cycloheximide (50μg/ml) inhibited LA-induced Nrf2 nuclear accumulation by 94%. Providing cells with the mTOR inhibitor, rapamycin, decreased basal Nrf2 levels by 84% after 4h, but LA overcame this inhibition. LA-mediated de novo protein translation was confirmed using HepG2 cells transfected with a bicistronic construct containing an internal ribosome entry sequence (IRES) for Nrf2, with significant (P<0.05) increase in IRES use under LA treatment. These results suggest that a dithiol stimulus mediates Nrf2 nuclear tenure via cap-independent protein translation. Thus, translational control of Nrf2 synthesis, rather than reliance solely on pre-existing protein, may mediate the rapid burst of Nrf2 nuclear accumulation following stress stimuli.
authors.affiliation: Linus Pauling Institute, Oregon State University, Corvallis, OR 97331, USA.
authors.forename: Kate Petersen
authors.initials: KP
authors.lastname: Shay
authors.forename: Alexander J
authors.initials: AJ
authors.lastname: Michels
authors.forename: Wenge
authors.initials: W
authors.lastname: Li
authors.forename: Ah-Ng Tony
authors.initials: AN
authors.lastname: Kong
authors.forename: Tory M
authors.initials: TM
authors.lastname: Hagen
mesh_terms: D001483:Base Sequence; D002451:Cell Compartmentation / Q000187:drug effects; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D056945:Hep G2 Cells; D006801:Humans; D008658:Inactivation, Metabolic; D051267:NF-E2-Related Factor 2 / Q000096:biosynthesis*; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D061988:Proteasome Inhibitors; D014176:Protein Biosynthesis / Q000187:drug effects*; D012315:RNA Caps / Q000187:drug effects / Q000378:metabolism*; D020123:Sirolimus / Q000494:pharmacology; D013312:Stress, Physiological / Q000187:drug effects*; D008063:Thioctic Acid / Q000494:pharmacology*
chemical_list: D051267:NF-E2-Related Factor 2; C495635:NFE2L2 protein, human; D061988:Proteasome Inhibitors; D012315:RNA Caps; D008063:Thioctic Acid; D046988:Proteasome Endopeptidase Complex; D020123:Sirolimus