The Role of Nrf2 in Cardiovascular Function and Disease

Figure 1

Nrf2 and KEAP1 structure. Nrf2 is a cap‘n’collar-basic region leucine zipper (CNC-bZIP), and its human sequence contains 605 amino acids, divided into seven domains: Neh1 to Neh7. Neh1 contains a CNC-bZIP motif, allowing heterodimerization with Maf proteins and DNA binding [54]. The Neh2 domain contains the Keap1 binding site (DLG and ETGE motifs), necessary for its cytoplasmic retention and degradation [55]. The Neh3 domain is fundamental for Nrf2 transcriptional activation through binding with chromo-ATPase/helicase DNA-binding protein 6 (CHD6) [56]. Neh4 and Neh5 provide an interaction site for nuclear cofactor RAC3/AIB1/SRC-3 [57] and CREB-binding protein (CBP) [58] which enhances the Nrf2/ARE activation pathways, partially by promoting acetylation of Nrf2 [59]. Additionally, Nrf2 possesses a redox-sensitive nuclear exporting signal within the Neh5 transactivation domain able to regulate its cellular localization [60]. The serine-rich Neh6 domain contains two motifs (DSGIS and DSAPGS) involved in the negative regulation of Nrf2. Glycogen synthase kinase 3 (GSK-3) phosphorylates serine residues within Neh6 enabling the interaction with the β-transducin repeat-containing protein (β-TrCP) which acts as a substrate receptor for Skp1–Cul1–Rbx1/Roc1 ubiquitin ligase complex, leading to KEAP1-independent degradation [41]. Neh7 domain interacts with retinoid X receptor alpha (RXRα), responsible for Nrf2/ARE signaling inhibition [61]. Human Kelch-like ECH-associated protein 1 (KEAP1) is a 69 kD protein, containing 27 cysteine residues. It is a substrate adaptor for cullin (Cul3) which contains E3 ubiquitin ligase (E3). KEAP1 is composed of five domains starting from the N-terminal region, a BTB dimerization domain (Broad-Complex, Tramtrack, and Bric-à-brac) which contains the Cys151 residue, a cysteine-rich intervening region (IVR) domain with two cysteine domain residues Cys273 and Cys288, critical for stress sensing. A Kelch domain/double glycine repeat (DGR) domain possessing 6 Kelch repeats and ending with a C-terminal region [62]. KEAP1 needs a domain capable to homodimerize and interact with Cul3, forming the Nrf2 inhibitor complex (iNrf2), and this is the BTB domain [63]. The Cys151 in the same domain plays an important role on Nrf2 activation in response to oxidative stress [64]. Furthermore, the IVR domain is highly sensitive to oxidation and contains three cysteines, 273, 288, and 297 which regulate Nrf2 activation and repression [16, 65]. The DGR domain acts as an Nrf2 repressor; it contains six repetitive Kelch structures that specifically bind to the Neh2 domain on Nrf2 [15].