The Keap1-Nrf2 system in cancers: stress response and anabolic metabolism

Abstract

The Keap1-Nrf2 [Kelch-like ECH-associated protein 1-nuclear factor (erythroid-derived 2)-like 2] pathway plays a central role in the protection of cells against oxidative and xenobiotic stresses. Nrf2 is a potent transcription activator that recognizes a unique DNA sequence known as the antioxidant response element (ARE). Under normal conditions, Nrf2 binds to Keap1 in the cytoplasm, resulting in proteasomal degradation. Following exposure to electrophiles or reactive oxygen species, Nrf2 becomes stabilized, translocates into the nucleus, and activates the transcription of various cytoprotective genes. Increasing attention has been paid to the role of Nrf2 in cancer cells because the constitutive stabilization of Nrf2 has been observed in many human cancers with poor prognosis. Recent studies have shown that the antioxidant and detoxification activities of Nrf2 confer chemo- and radio-resistance to cancer cells. In this review, we provide an overview of the Keap1-Nrf2 system and discuss its role under physiological and pathological conditions, including cancers. We also introduce the results of our recent study describing Nrf2 function in the metabolism of cancer cells. Nrf2 likely confers a growth advantage to cancer cells through enhancing cytoprotection and anabolism. Finally, we discuss the possible impact of Nrf2 inhibitors on cancer therapy.

Keywords: glutathione; purine nucleotide; redox homeostasis; stress response; transcription.

FIGURE 1

The Keap1–Nrf2 system. Under normal conditions, Nrf2 is constantly ubiquitinated through Keap1 and degraded in the proteasome. Following exposure to electrophiles or oxidative stress, Keap1 is inactivated. Stabilized Nrf2 accumulates in the nucleus and activates many cytoprotective genes. Ub, ubiquitin.

FIGURE 2

Keap1 is a thiol-rich protein that is sensitive to electrophilic covalent modification. Domain structure of Keap1 is shown, and reactive cysteine residues are indicated with brown circles. Direct modification of cysteine residues was demonstrated using various electrophiles. Each electrophile attacks a unique set of cysteines.

FIGURE 3

Molecular mechanism of stress sensing in the Keap1–Nrf2 system. (A) The Keap1 homodimer binds one molecule of Nrf2. The ETGE and DLG motifs of Nrf2 represent high and low affinity binding sites, respectively. The lysine residues (K) are clustered between the two motifs and represent ubiquitination targets. (B) The modification of cysteine residues in Keap1 with electrophiles is expected to modify the overall conformation of the Keap1 homodimer, resulting in the termination of Nrf2 ubiquitination.

FIGURE 4

Domain structures of Keap1 (A) and Nrf2 (B).(A) The N-terminal region of Keap1 mediates homodimerization and association with Cul3, and the C-terminal region of Keap1 mediates binding with Nrf2. (B) The N-terminal region of Nrf2 is designated Neh2 domain, which contains two motifs, DLG and ETGE, responsible for the interaction with Keap1. Neh4, Neh5, and Neh3 domains are important for the transactivation activity of Nrf2 (Katoh et al., 2001; Nioi et al., 2005). Neh6 domain contains the phosphodegron that is recognized by β-TrCP. Neh1 domain is a basic-region leucine zipper motif for DNA binding and dimerization with small Maf.

FIGURE 5

Increased activity of NRF2 in cancer cells. The degradation of NRF2 is inhibited in some cases, and the production of NRF2 is increased in other cases.

FIGURE 6

Somatic mutations in KEAP1 and NRF2 genes identified in human cancers. (A) More than half of the KEAP1 gene mutations were identified in the DC domain. (B) All NRF2 gene mutations were restricted to the DLG and ETGE motifs.

FIGURE 7

Dominant negative effect of the Keap1 gene mutation. The intact Keap1 homodimer ubiquitinates Nrf2, while the Keap1 dimer, containing one or two mutant Keap1 proteins, cannot ubiquitinate Nrf2. A single allele mutation in the Keap1 gene results in the production of an equal molar ratio of the wild-type Keap1 (W) to the mutant Keap1 (M). Keap1 dimerization generates three kinds of dimers, W–W, W–M, and M–M, by 1:2:1 ratio. Since W–M and M–M dimers are incapable of ubiquitinating Nrf2, the overall Keap1 activity is reduced by 75%, and consequently, Nrf2 is stabilized.

FIGURE 8

Contribution of Nrf2 to cellular metabolism. The enzymes regulated through Nrf2 are indicated with double-framed boxes (G6PD, glucose-6-phosphate dehydrogenase; PGD, phosphogluconate dehydrogenase; TKT, transketolase; TALDO1, transaldolase 1; PPAT, phosphoribosyl pyrophosphate amidotransferase; MTHFD2, methylenetetrahydrofolate dehydrogenase 2; ME1, malic enzyme 1; γGCL, γ-glutamylcysteinyl ligase). Abbreviations of metabolites; 1,3-BPG, 1,3-bisphosphoglycerate; 2-PG, 2-phosphoglycerate; 3-PG, 3-phosphoglycerate; 6-PG, 6-phosphogluconate; 5-PRA, β-5-phosphorybosylamine; F1,6P, fructose 1,6-bis-phosphate; F6P, fructose 6-phosphate; G6P, glucose 6-phosphate; GAP, glyceraldehyde 3-phosphate; PEP, phosphoenolpyruvate; PRPP, phosphoribosyl phosphate; R5P, ribose 5-phosphate; Ru5P, ribulose 5-phosphate; S7P, sedoheptulose 7-phosphate.

FIGURE 9

Functional expansion of Nrf2 in proliferating cells. In quiescent cells, Nrf2 is activated in response to oxidative stress and induces the expression of cytoprotective genes encoding antioxidant proteins and detoxification enzymes, which maintains the cellular redox homeostasis. In proliferating cells, Nrf2 activity is augmented especially under the sustained activation of PI3K–Akt pathway. Nrf2 activates metabolic genes in addition to cytoprotective genes, resulting in the redirection of glucose and glutamine into anabolic pathway, which is advantageous for cell proliferation.