title: The carboxy-terminal Neh3 domain of Nrf2 is required for transcriptional activation.
abstract: Nrf2 is a transcription factor critical for the maintenance of cellular redox homeostasis. We have previously found that Nrf2 is a labile protein, and its activation in cells under stress involves mechanisms leading to its stabilization. As a modular protein, Nrf2 possesses distinct transactivation and DNA binding domains essential for its transcriptional activity. In this study, we found that the C-terminal "Neh3" domain of Nrf2 is also important for its activity. Deletion of the last 16 amino acids of the protein completely abolishes its ability to activate both reporter and endogenous gene expression. Using site-directed mutagenesis, we have identified a stretch of amino acids within this region that are essential for its activity and that are found to be conserved across species and among other members of the CNC-bZIP family. Importantly, deletion of the final 16 amino acids of Nrf2 does not influence its dimerizing capability, DNA binding activity, or subcellular localization, although it does increase the half-life of the protein. In addition, this region was found to be important for interaction with CHD6 (a chromo-ATPase/helicase DNA binding protein) in a yeast two-hybrid screen. RNA interference-mediated knockdown of CHD6 reduced both the basal and tert-butylhydroquinone-inducible expression of NQO1, a prototypical Nrf2 target gene. These data suggest that the Neh3 domain may act as a transactivation domain and that it is possibly involved in interaction with components of the transcriptional apparatus to affect its transcriptional activity.
authors.affiliation: Schering-Plough Research Institute, Kenilworth, NJ 07033, USA.
authors.forename: Paul
authors.initials: P
authors.lastname: Nioi
authors.forename: Truyen
authors.initials: T
authors.lastname: Nguyen
authors.forename: Philip J
authors.initials: PJ
authors.lastname: Sherratt
authors.forename: Cecil B
authors.initials: CB
authors.lastname: Pickett
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002467:Cell Nucleus / Q000737:chemistry / Q000378:metabolism; D002478:Cells, Cultured; D017124:Conserved Sequence; D019281:Dimerization; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D000072019:Kelch-Like ECH-Associated Protein 1; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D051267:NF-E2-Related Factor 2 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000378:metabolism; D034622:RNA Interference; D051381:Rats; D020218:Response Elements; D017384:Sequence Deletion; D015533:Transcriptional Activation; D020798:Two-Hybrid System Techniques
chemical_list: D047908:Intracellular Signaling Peptides and Proteins; C431364:KEAP1 protein, human; D000072019:Kelch-Like ECH-Associated Protein 1; D051267:NF-E2-Related Factor 2; D011506:Proteins