SCF/β-TrCP Promotes Glycogen Synthase Kinase 3-Dependent Degradation of the Nrf2 Transcription Factor in a Keap1-Independent Manner

Regulation of transcription factor Nrf2 (NF-E2-related factor 2) involves redox-sensitive proteasomal degradation via the E3 ubiquitin ligase Keap1/Cul3. However, Nrf2 is controlled by other mechanisms that have not yet been elucidated. We now show that glycogen synthase kinase 3 (GSK-3) phosphorylates a group of Ser residues in the Neh6 domain of mouse Nrf2 that overlap with an SCF/beta-TrCP destruction motif (DSGIS, residues 334 to 338) and promotes its degradation in a Keap1-independent manner. Nrf2 was stabilized by GSK-3 inhibitors in Keap1-null mouse embryo fibroblasts. Similarly, an Nrf2(Delta ETGE) mutant, which cannot be degraded via Keap1, accumulated when GSK-3 activity was blocked. Phosphorylation of a Ser cluster in the Neh6 domain of Nrf2 stimulated its degradation because a mutant Nrf2(Delta ETGE 6S/6A) protein, lacking these Ser residues, exhibited a longer half-life than Nrf2(Delta ETGE). Moreover, Nrf2(Delta ETGE 6S/6A) was insensitive to beta-TrCP regulation and exhibited lower levels of ubiquitination than Nrf2(Delta ETGE). GSK-3 beta enhanced ubiquitination of Nrf2(Delta ETGE) but not that of Nrf2(Delta ETGE 6S/6A). The Nrf2(Delta ETGE) protein but not Nrf2(Delta ETGE 6S/6A) coimmunoprecipitated with beta-TrCP, and this association was enhanced by GSK-3 beta. Our results show for the first time that Nrf2 is targeted by GSK-3 for SCF/beta-TrCP-dependent degradation. We propose a dual degradation model to describe the regulation of Nrf2 under different pathophysiological conditions.