title: G-Quadruplex in the NRF2 mRNA 5' Untranslated Region Regulates De Novo NRF2 Protein Translation under Oxidative Stress.
abstract: Inhibition of protein synthesis serves as a general measure of cellular consequences of chemical stress. A few proteins are translated selectively and influence cell fate. How these proteins can bypass the general control of translation remains unknown. We found that low to mild doses of oxidants induce de novo translation of the NRF2 protein. Here we demonstrate the presence of a G-quadruplex structure in the 5' untranslated region (UTR) of NRF2 mRNA, as measured by circular dichroism, nuclear magnetic resonance, and dimethylsulfate footprinting analyses. Such a structure is important for 5'-UTR activity, since its removal by sequence mutation eliminated H2O2-induced activation of the NRF2 5' UTR. Liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteomics revealed elongation factor 1 alpha (EF1a) as a protein binding to the G-quadruplex sequence. Cells responded to H2O2treatment by increasing the EF1a protein association with NRF2 mRNA, as measured by RNA-protein interaction assays. The EF1a interaction with small and large subunits of ribosomes did not appear to change due to H2O2treatment, nor did posttranslational modifications, as measured by two-dimensional (2-D) Western blot analysis. Since NRF2 encodes a transcription factor essential for protection against tissue injury, our data have revealed a novel mechanism of cellular defense involving de novo NRF2 protein translation governed by the EF1a interaction with the G-quadruplex in the NRF2 5' UTR during oxidative stress.
authors.affiliation: Department of Pharmacology, College of Medicine, University of Arizona, Tucson, Arizona, USA.
authors.forename: Sang C
authors.initials: SC
authors.lastname: Lee
authors.affiliation: Department of Pharmacology, College of Medicine, University of Arizona, Tucson, Arizona, USA.
authors.forename: Jack
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Department of Pharmacology, College of Medicine, University of Arizona, Tucson, Arizona, USA.
authors.forename: Josh
authors.initials: J
authors.lastname: Strom
authors.affiliation: Department of Pharmacology, College of Medicine, University of Arizona, Tucson, Arizona, USA.
authors.forename: Danzhou
authors.initials: D
authors.lastname: Yang
authors.affiliation: Department of Pharmacology, College of Medicine, University of Arizona, Tucson, Arizona, USA.
authors.forename: Thai Nho
authors.initials: TN
authors.lastname: Dinh
authors.affiliation: Department of Pharmacology, College of Medicine, University of Arizona, Tucson, Arizona, USA.
authors.forename: Kyle
authors.initials: K
authors.lastname: Kappeler
authors.affiliation: Department of Pharmacology, College of Medicine, University of Arizona, Tucson, Arizona, USA qchen@email.arizona.edu.
authors.forename: Qin M
authors.initials: QM
authors.lastname: Chen
mesh_terms: D020121:5' Untranslated Regions / Q000187:drug effects; D002942:Circular Dichroism; D054856:G-Quadruplexes; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D009682:Magnetic Resonance Spectroscopy; D051267:NF-E2-Related Factor 2 / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D018384:Oxidative Stress; D020648:Peptide Elongation Factor 1 / Q000378:metabolism*; D014176:Protein Biosynthesis / Q000187:drug effects; D012333:RNA, Messenger / Q000737:chemistry
chemical_list: D020121:5' Untranslated Regions; D051267:NF-E2-Related Factor 2; C495635:NFE2L2 protein, human; D020648:Peptide Elongation Factor 1; D012333:RNA, Messenger; D006861:Hydrogen Peroxide
keywords: RNA binding proteins; RNA structure; antioxidant genes; protein translation; proteomics