Action of Nrf2 and Keap1 in ARE-mediated NQO1 expression by quercetin

Polyphenols are characterized by the presence of more than one phenolic group and are widely distributed in many fi-uits and vegetables. They possess antioxidant properties and interact with cellular defense systems through the antioxidant-responsive element/electrophile-responsive element (ARE/EpRE) although the precise mechanism by which polyphenols influence transcription factor complexes to target ARE is poorly understood. In the present study, we chose a typical polyphenol, quercetin, to investigate the mechanism in human HepG2 cells. Quercetin enhanced the ARE binding activity and Ntf2-mediated transcription activity. Molecular evidence revealed that quercetin not only up-regulated the expression of Nrf2 mRNA and protein, but also stabilized Nrf2 protein by inhibiting the ubiquitination and proteasomal turnover of Nrf2. At the same time, quercetin markedly reduced the level of Keap1 protein in posuranslational levels through the formation of modified Keap1 protein, rather than 26S proteasome-dependent degradation mechanisms, without affecting the dissociation of Keap1-Nrf2. Silencing Keap1 using Keap1 siRNA significantly increased the NrIF2-dependent ARE activity, whereas silencing Nlrf2 using Nrf2 siRNA markedly reduced the ARE activity under both baseline and quercetin-induced conditions. Thus, we conclude that the pathway of quercetin-induced ARE activity involves up-regulation of Nrf2 through the regulation of both transcription and posttranscription sites and repression of Keap1 by affecting the posttranscription site, revealing some substantial differences between oxidative inducers. Thus, the findings provide an insight into the mechanisms underlying polyphenolic compounds in cytoprotection and cancer chemoprevention. (c) 2007 Elsevier Inc. All rights reserved.