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. 2017 Mar 13;127(4):1505–1516. doi: 10.1172/JCI88574

Figure 4. SIRT2 deacetylates NRF2 and promotes its ubiquitination.

Figure 4

(A) Co-IP of SIRT2 and either NRF2 or KEAP1 in HeLa cells transfected with Flag-SIRT2. The label “GFP” refers to transfection with a control GFP plasmid, while Flag-SIRT2 was transfected with a Flag-SIRT2 construct. Endogenous NRF2 was detected in the input lane transfected with GFP. (B) Co-IP experiments of HEK293T cells transfected with V5-NRF2 and either GFP (control) or different HATs including EP300, KAT2B, KAT2A, KAT5, and CREBBP, using an anti-V5 antibody for IP and anti–acetyl-lysine (α-Ac-K) or anti-V5 (α-V5) antibodies for immunoblotting. WB, Western blot. (C) HEK293T cells cotransfected with p300 and V5-NRF2 were harvested, and NRF2 was purified by IP. Immunoprecipitated acetylated NRF2 was mixed with NAD+ and WT purified SIRT2 in the presence and absence of NAM, which was used as a deacetylase inhibitor. rSIRT2, recombinant SIRT2. (D) Time course of NRF2 protein stability in Sirt2+/+ and Sirt2–/– MEFs after treatment with 100 μg/ml CHX. All panels are representative blots from 2 independent experiments.