FIG. 2.

Rapid turnover of Nrf2 requires its association with Keap1. (A) Schematic presentation of Nrf2 deletion mutants. ΔC lacks the C terminus including the NLS of wild-type Nrf2. ΔC/ETGE lacks both this C terminus and the ETGE motif, which is crucial for association with Keap1. (B) Cytoplasmic localization of ΔC and ΔC/ETGE mutants in Cos7 cells (b and c). These mutant proteins were stained by an immunohistochemical method with anti-Nrf2 (C4) antibody. Nuclei were stained with DAPI (d to f). (C and D) Deletion of the ETGE motif abolished the degradation of Nrf2 by Keap1. This suggests that Keap1 positively regulates the degradation of Nrf2 through its association. The experimental procedure was described in the legend to Fig. 1. The averages of the band intensities of ΔC and ΔC/ETGE mutants represent two independent experiments done in duplicate.