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WNT-3A regulates an Axin1/NRF2 complex that regulates antioxidant metabolism in hepatocytes
Patricia Rada1, Ana I Rojo, Anika Offergeld
11 Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), ISCIII , Madrid, Spain .
Aims:
Nuclear factor (erythroid-derived 2)-like 2 (NRF2) is a master regulator of oxidant and xenobiotic metabolism, but it is unknown how it is regulated to provide basal expression of this defense system. Here, we studied the putative connection between NRF2 and the canonical WNT pathway, which modulates hepatocyte metabolism.
Results:
WNT-3A increased the levels of NRF2 and its transcriptional signature in mouse hepatocytes and HEK293T cells. The use of short interfering RNAs in hepatocytes and mouse embryonic fibroblasts which are deficient in the redox sensor Kelch-like ECH-associated protein 1 (KEAP1) indicated that WNT-3A activates NRF2 in a β-Catenin- and KEAP1-independent manner. WNT-3A stabilized NRF2 by preventing its GSK-3-dependent phosphorylation and subsequent SCF/β-TrCP-dependent ubiquitination and proteasomal degradation. Axin1 and NRF2 were physically associated in a protein complex that was regulated by WNT-3A, involving the central region of Axin1 and the Neh4/Neh5 domains of NRF2. Axin1 knockdown increased NRF2 protein levels, while Axin1 stabilization with Tankyrase inhibitors blocked WNT/NRF2 signaling. The relevance of this novel pathway was assessed in mice with a conditional deletion of Axin1 in the liver, which showed upregulation of the NRF2 signature in hepatocytes and disruption of liver zonation of antioxidant metabolism.
Innovation:
NRF2 takes part in a protein complex with Axin1 that is regulated by the canonical WNT pathway. This new WNT-NRF2 axis controls the antioxidant metabolism of hepatocytes.
Conclusion:
These results uncover the participation of NRF2 in a WNT-regulated signalosome that participates in basal maintenance of hepatic antioxidant metabolism.
Insights
The WNT pathway regulates Nuclear factor (erythroid-derived 2)-like 2 (NRF2) levels via Axin1, impacting hepatic antioxidant metabolism. This discovery reveals a novel WNT-NRF2 axis controlling cellular defense mechanisms.
Area of Science:
- Cellular Biology
- Molecular Biology
- Metabolism
Background:
- Nuclear factor (erythroid-derived 2)-like 2 (NRF2) is a key regulator of cellular defense against oxidants and xenobiotics.
- The precise mechanisms regulating basal NRF2 expression remain incompletely understood.
- The canonical WNT pathway is known to modulate hepatocyte metabolism, but its connection to NRF2 is unexplored.
Purpose of the Study:
- To investigate the potential link between the canonical WNT pathway and NRF2.
- To elucidate the regulatory mechanisms controlling basal NRF2 expression.
- To understand the role of this interaction in hepatic antioxidant metabolism.
Main Methods:
- Utilized mouse hepatocytes and HEK293T cells.
- Employed short interfering RNAs (siRNAs) in hepatocytes and mouse embryonic fibroblasts lacking Kelch-like ECH-associated protein 1 (KEAP1).
- Investigated protein-protein interactions using co-immunoprecipitation and assessed protein stability through Western blotting.
- Examined the effects of Axin1 manipulation and Tankyrase inhibitors on WNT/NRF2 signaling.
- Analyzed liver-specific Axin1 conditional knockout mice.
Main Results:
- WNT-3A stimulation increased NRF2 levels and its downstream transcriptional targets in hepatocytes and HEK293T cells.
- WNT-3A activated NRF2 independently of β-Catenin and KEAP1, by preventing NRF2 phosphorylation and subsequent degradation.
- Identified a physical association between Axin1 and NRF2, forming a WNT-regulated complex.
- Axin1 knockdown elevated NRF2 protein levels, while Tankyrase inhibitors stabilizing Axin1 blocked WNT/NRF2 signaling.
- Liver-specific Axin1 deletion led to increased NRF2 signature and disrupted antioxidant metabolism zonation in hepatocytes.
Conclusions:
- NRF2 is part of a novel WNT-regulated signalosome involving Axin1.
- This WNT-NRF2 axis plays a crucial role in the basal maintenance of hepatic antioxidant metabolism.
- The findings uncover a new regulatory pathway for cellular defense mechanisms in hepatocytes.
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