Abstract
Abstract Curcumin, the yellow pigment of tumeric (Curcuma longa L., Zingiberaceae), is one of the most widely used spices in the East Asia. Curcumin has been shown to possess anti-inflammatory, antioxidative and chemopreventive effects. Although a growing body of evidence suggests that the chemopreventive potential of curcumin depends partly on its ability to induce cytoprotective proteins through the activation of nuclear factor erythroid-related factor-2 (Nrf2), the molecular mechanisms remain elusive. The present study was aimed to elucidate the mechanisms underlying the activation of Nrf2 and induction of cytoprotective gene expression by curcumin. Incubation of mouse epidermal (JB6) cells with curcumin resulted in the induction of heme oxygenase-1 (HO-1) and NAD(P)H oxidoreductase 1 (NQO1) at both mRNA and protein levels. Curcumin-induced expression of HO-1 and NQO1 was abrogated in cells transiently transfected with Nrf2 siRNA. Furthermore, embryo fibroblasts from Nrf2 knock-out mice were not responsive to curcumin, compared with those from wild-type animals, in terms of inducing HO-1 and NQO1 expression. While curcumin treatment increased protein expression of Nrf2, it failed to alter the steady-state level of the Nrf2 mRNA transcript, suggesting that protein stabilization might be involved in curcumin-induced Nrf2 accumulation. Treatment of JB6 cells with curcumin did stabilize Nrf2 by inhibiting ubiquitination and subsequent 26S proteasomal degradation of Nrf2. Kelch-like ECH-associated protein-1 (Keap1), a substrate adaptor of Cullin3-Rbx1 E3 ubiquitin ligase complex, has several reactive cysteine residues, and modification of these cysteine residues has been proposed as a molecular mechanism underlying oxidative and electrophilic stress-induced activation of Nrf2. The thiol-reducing agent dithiothreitol abrogated curcumin-induced accumulation of Nrf2 and expression of cytoprotective proteins. In addition, tetrahydrocurcumin, a non-electrophilic analogue of curcumin that lacks the α,β-unsaturated carbonyl group, failed to induce cytoprotective protein expression as well as Nrf2 nuclear translocation, indicative of a pivotal role of cysteine residues of Keap1 in curcumin-induced Nrf2 activation. Cells transfected with a mutant Keap1 protein in which cysteine 151 is replaced by serine exhibited reduction in curcumin-induced Nrf2 activation. Although curcumin did not cause dissociation of Cullin3-Rbx1 E3 ubiquitin ligase complex components, it increased interaction of Nrf2 with the Keap1. Thus, it is likely that curcumin inhibits the ability of the Cullin3-Rbx1 E3 ubiquitin ligase to target Nrf2 for ubiquitination by modifying Keap1 Cys 151 residue. This may change the conformation of the complex and saturates the binding capacity of Keap1 to Nrf2, facilitating nuclear translocation of Nrf2. Citation Information: Cancer Prev Res 2011;4(10 Suppl):B67.
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