Nrf2 is controlled by two distinct β-TrCP recognition motifs in its Neh6 domain, one of which can be modulated by GSK-3 activity.

By S. Chowdhry, Yiguo Zhang, Michael T. McMahon, C. Sutherland, A. Cuadrado, J. Hayes. Oncogene, 2012

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Identification of regulatable mechanisms by which transcription factor NF-E2 p45-related factor 2 (Nrf2) is repressed will allow strategies to be designed that counter drug resistance associated with its upregulation in tumours that harbour somatic mutations in Kelch-like ECH-associated protein-1 (Keap1), a gene that encodes a joint adaptor and substrate receptor for the Cul3–Rbx1/Roc1 ubiquitin ligase. We now show that mouse Nrf2 contains two binding sites for β-transducin repeat-containing protein (β-TrCP), which acts as a substrate receptor for the Skp1–Cul1–Rbx1/Roc1 ubiquitin ligase complex. Deletion of either binding site in Nrf2 decreased β-TrCP-mediated ubiquitylation of the transcription factor. The ability of one of the two β-TrCP-binding sites to serve as a degron could be both increased and decreased by manipulation of glycogen synthase kinase-3 (GSK-3) activity. Biotinylated-peptide pull-down assays identified DSGIS338 and DSAPGS378 as the two β-TrCP-binding motifs in Nrf2. Significantly, our pull-down assays indicated that β-TrCP binds a phosphorylated version of DSGIS more tightly than its non-phosphorylated counterpart, whereas this was not the case for DSAPGS. These data suggest that DSGIS, but not DSAPGS, contains a functional GSK-3 phosphorylation site. Activation of GSK-3 in Keap1-null mouse embryonic fibroblasts (MEFs), or in human lung A549 cells that contain mutant Keap1, by inhibition of the phosphoinositide 3-kinase (PI3K)–protein kinase B (PKB)/Akt pathway markedly reduced endogenous Nrf2 protein and decreased to 10–50% of normal the levels of mRNA for prototypic Nrf2-regulated enzymes, including the glutamate-cysteine ligase catalytic and modifier subunits, glutathione S-transferases Alpha-1 and Mu-1, haem oxygenase-1 and NAD(P)H:quinone oxidoreductase-1. Pre-treatment of Keap1−/− MEFs or A549 cells with the LY294002 PI3K inhibitor or the MK-2206 PKB/Akt inhibitor increased their sensitivity to acrolein, chlorambucil and cisplatin between 1.9-fold and 3.1-fold, and this was substantially attenuated by simultaneous pre-treatment with the GSK-3 inhibitor CT99021.

Subfields: Molecular Biology, Public Health, Environmental and Occupational Health

View Paper: https://www.semanticscholar.org/paper/9507448eacc5af1461b126c7da6fcb2cc92ebcd5

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Citations in the corpus

Mechanisms of activation of the transcription factor Nrf2 by redox stressors, nutrient cues, and energy status and the pathways through which it attenuates degenerative disease (2015, neutral): “10B), whereas the DSAPGS motif does not appear to be phosphorylated by GSK-3 [74].”
The emerging role of the Nrf2–Keap1 signaling pathway in cancer (2013, background): “Phosphorylation of the DSGIS motif by GSK-3 increases the ability of β-TrCP to ubiquitinate Nrf2 and promote its rapid turnover (McMahon et al. 2004; Rada et al. 2011; Chowdhry et al. 2013).”
Hydrogen peroxide sensing, signaling and regulation of transcription factors (2014, background): “Other kinases that inactivate GSK-3 are ERK, p38 MAPK and PKC [216], and it has been suggested that these kinases may regulate NRF2 indirectly by inhibiting GSK-3 [217].”
Transcriptional Regulation by Nrf2 (2017, background): “It contains two conserved peptide motifs, DSGIS and DSAPGS, which are recognized by b-transducing repeat-containing protein (b-TrCP) (24).”
The Keap1-Nrf2 pathway: Mechanisms of activation and dysregulation in cancer (2013, background): “In addition, PI3K inhibitors [11,13] and Nrf2 siRNA [14] have been used to inhibit Nrf2 in cancer cells.”
Nrf2 and Oxidative Stress: A General Overview of Mechanisms and Implications in Human Disease (2022, background): “Similar to Neh2, Neh6 contains two highly conserved 334DSGIS338 and 373DSAPGS378 motifs to which the β-transducin repeat-containing protein (βTrCP) binds, and within the DSGIS motif, a phosphorylation site for glycogen synthase kinase-3 (GSK3) that enhances βTrCP activity upon GSK3-mediated phosphor”