SCOPE: 6-Methylthiohexyl isothiocyanate (6-MTITC), one of the major bioactive ingredients in Japanese Wasabi, has revealed cytoprotective and cancer chemopreventive effects. This study aims to clarify the molecular mechanisms how 6-MTITC modulates nuclear factor E2-related factor 2 (Nrf2)/Kelchlike ECH-associating protein 1 (Keap1) system in antioxidant-responsive element (ARE)-mediated nicotinamide adenine dinucleotide phosphate (NADP): quinone oxidoreductase 1 (NQO1) expression. METHODS AND RESULTS: HepG2 cells were treated with 6-MTITC with varying time and dose. NQO1, Nrf2, and Keap1 proteins were detected by Western blotting. ARE transactivation was detected by electrophilic mobility shift assay and reporter gene assay. Nuclear localization of Nrf2 was determined by immunocytochemistry assay. Ubiquitination of Nrf2 and Keap1 was detected using immunoprecipitation after treatment with MG132. Small interfering RNA was used to knockdown Nrf2 or Keap1. The results revealed that 6-MTITC modulated Nrf2/ARE pathway by stimulating Keap1 modification, and inhibiting Nrf2 ubiquitination and protein turnover. These actions finally increased nuclear Nrf2 accumulation and ARE-binding activity. Moreover, silencing Nrf2 markedly reduced ARE-driven activity induced by 6-MTITC. CONCLUSION: 6-MTITC modulated ARE-driven NQO1 expression by stabilizing Nrf2 with enhanced Keap1 modification and decreased Nrf2 degradation.
SCOPE: 6-Methylthiohexyl isothiocyanate (6-MTITC), one of the major bioactive ingredients in Japanese Wasabi, has revealed cytoprotective and cancer chemopreventive effects. This study aims to clarify the molecular mechanisms how 6-MTITC modulates nuclear factor E2-related factor 2 (Nrf2)/Kelchlike ECH-associating protein 1 (Keap1) system in antioxidant-responsive element (ARE)-mediated nicotinamide adenine dinucleotide phosphate (NADP): quinone oxidoreductase 1 (NQO1) expression. METHODS AND RESULTS: HepG2 cells were treated with 6-MTITC with varying time and dose. NQO1, Nrf2, and Keap1 proteins were detected by Western blotting. ARE transactivation was detected by electrophilic mobility shift assay and reporter gene assay. Nuclear localization of Nrf2 was determined by immunocytochemistry assay. Ubiquitination of Nrf2 and Keap1 was detected using immunoprecipitation after treatment with MG132. Small interfering RNA was used to knockdown Nrf2 or Keap1. The results revealed that 6-MTITC modulated Nrf2/ARE pathway by stimulating Keap1 modification, and inhibiting Nrf2 ubiquitination and protein turnover. These actions finally increased nuclear Nrf2 accumulation and ARE-binding activity. Moreover, silencing Nrf2 markedly reduced ARE-driven activity induced by 6-MTITC. CONCLUSION:6-MTITC modulated ARE-driven NQO1 expression by stabilizing Nrf2 with enhanced Keap1 modification and decreased Nrf2 degradation.
Authors: Jean Bousquet; Wienczyslawa Czarlewski; Torsten Zuberbier; Joaquim Mullol; Hubert Blain; Jean-Paul Cristol; Rafael De La Torre; Nieves Pizarro Lozano; Vincent Le Moing; Anna Bedbrook; Ioana Agache; Cezmi A Akdis; G Walter Canonica; Alvaro A Cruz; Alessandro Fiocchi; Joao A Fonseca; Susana Fonseca; Bilun Gemicioğlu; Tari Haahtela; Guido Iaccarino; Juan Carlos Ivancevich; Marek Jutel; Ludger Klimek; Helga Kraxner; Piotr Kuna; Désirée E Larenas-Linnemann; Adrian Martineau; Erik Melén; Yoshitaka Okamoto; Nikolaos G Papadopoulos; Oliver Pfaar; Frederico S Regateiro; Jacques Reynes; Yves Rolland; Philip W Rouadi; Boleslaw Samolinski; Aziz Sheikh; Sanna Toppila-Salmi; Arunas Valiulis; Hak-Jong Choi; Hyun Ju Kim; Josep M Anto Journal: Int Arch Allergy Immunol Date: 2021-02-10 Impact factor: 2.749