Dynamics of Nrf2 and Keap1 in ARE-mediated NQO1 expression by wasabi 6-(methylsulfinyl)hexyl isothiocyanate.
6-(Methylsulfinyl)hexyl isothiocyanate (6-MSITC) is a bioactive ingredient present in wasabi, a popular pungent spice in Japan. Previous studies have revealed the cytoprotective and cancer chemopreventive effects of 6-MSITC. This study aims to clarify the molecular mechanisms by investigating the action of 6-MSITC on the Nrf2/Keap1 system. 6-MSITC up-regulated the expression of NAD(P)H:quinone oxidoreductase 1 (NQO1) by increasing the Nrf2 level. Treatment with 6-MSITC extended the half-life (t(1/2)) of Nrf2 protein from 11.5 to 35.2 min, approximately three times longer. Moreover, 6-MSITC suppressed the ubiquitination of Nrf2 but not Keap1. Alternatively, a modified Keap1 was observed in 6-MSITC-treated cells accompanying reduction of normal Keap1 protein. The results from cellular fractionation and immunocytochemistry assay revealed that Nrf2 was primarily accumulated in nucleus. EMSA and the reporter gene assay further demonstrated that 6-MSITC augmented Nrf2-ARE binding and transcription activity. Silencing Nrf2 using Nrf2 siRNA markedly reduced the Nrf2 level and ARE-driven activity under both baseline and 6-MSITC-induced conditions. Our data revealed that 6-MSITC enhanced Nrf2/ARE-driven NQO1 expression by stabilizing Nrf2 that was accomplished by modifying Keap1 with consequent inhibition of the ubiquitination and proteasomal turnover of Nrf2. The findings provide an insight into the mechanisms underlying 6-MSITC in cytoprotection and cancer chemoprevention.
The Engineering Research Center of Feed Safety and Efficient Utilization, Education Ministry, Hunan Agriculture University, Changsha, Hunan 410128, China. hou@chem.agri.kagoshima-u.ac.jp
De-Xing
DX
Hou
Yoshimi
Y
Korenori
Shunsuke
S
Tanigawa
Tomoe
T
Yamada-Kato
Masashi
M
Nagai
Xi
X
He
Jianhua
J
He
D002460:Cell Line; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D017930:Genes, Reporter; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D017879:Isothiocyanates / Q000494:pharmacology*; D000072019:Kelch-Like ECH-Associated Protein 1; D016660:NAD(P)H Dehydrogenase (Quinone) / Q000235:genetics* / Q000378:metabolism; D051267:NF-E2-Related Factor 2 / Q000235:genetics / Q000378:metabolism*; D010936:Plant Extracts / Q000494:pharmacology*; D011485:Protein Binding; D020218:Response Elements; D015533:Transcriptional Activation / Q000187:drug effects; D031228:Wasabia / Q000737:chemistry*
C505386:6-(Methylsulfinyl)hexyl isothiocyanate; D047908:Intracellular Signaling Peptides and Proteins; D017879:Isothiocyanates; C431364:KEAP1 protein, human; D000072019:Kelch-Like ECH-Associated Protein 1; D051267:NF-E2-Related Factor 2; D010936:Plant Extracts; D016660:NAD(P)H Dehydrogenase (Quinone); C447479:NQO1 protein, human