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Sulforaphane-induced epigenetic regulation of Nrf2 expression by DNA methyltransferase in human Caco-2 cells
Jia-Wei Zhou1, Min Wang2,3, Nuan-Xin Sun4
1Medical School, Qi-Lu Hospital of Shandong University, Jinan, Shandong 250012, P.R. China.
Abstract:
The present study aimed to investigate the mechanism underlying sulforaphane-mediated epigenetic regulation of nuclear factor-erythroid derived 2-like 2 (Nrf2) expression in human colon cancer. Proteins were extracted from normal Caco-2 cells using sulforaphane and 5-aza-2'-deoxycytidine (5-Aza) combined with trichostatin A (TSA). The mRNA and protein expression levels and activity of DNA methyltransferase 1 (DNMT1) were determined. Methylation-specific polymerase chain reaction and bisulfite genomic sequencing were also used to measure the methylation levels of CpG sites in the Nrf2 promoter region. Nrf2 expression was measured using reverse transcription-quantitative PCR and western blot analysis. The results demonstrated that sulforaphane did not affect DNMT1 mRNA expression levels. DNMT1 protein expression was inhibited by sulforaphane and 5-Aza co-treatment with TSA. Nrf2 promoter methylation decreased significantly in the sulforaphane group compared with the control group. Nrf2 promoter methylation level in the 5-Aza+TSA group was the lowest among all groups. Nrf2 mRNA levels exhibited significant differences between the sulforaphane-treated and control groups, as well as between the 5-Aza+TSA and control groups, and the sulforaphane-treated and 5-Aza+TSA groups. Nrf2 protein expression was also inhibited by sulforaphane, as well as 5-Aza co-treatment with TSA. The results revealed that sulforaphane may promote demethylation of the Nrf2 promoter region to increase activation of Nrf2, which induces chemoprevention of colon cancer.
Insights
Sulforaphane enhances colon cancer chemoprevention by demethylating the Nrf2 promoter, increasing Nrf2 activation. This epigenetic regulation boosts protective mechanisms against cancer development.
Area of Science:
- Epigenetics
- Cancer Biology
- Nutritional Biochemistry
Background:
- Colon cancer is a significant health concern.
- Nuclear factor-erythroid derived 2-like 2 (Nrf2) is a key regulator of cellular defense against oxidative stress and inflammation, often suppressed in cancer.
- Epigenetic modifications, such as DNA methylation, play a crucial role in regulating gene expression in cancer.
Purpose of the Study:
- To investigate the epigenetic mechanism by which sulforaphane regulates Nrf2 expression in human colon cancer cells.
- To determine the effect of sulforaphane on DNA methyltransferase 1 (DNMT1) activity and Nrf2 promoter methylation.
- To elucidate the role of Nrf2 activation in sulforaphane-induced chemoprevention of colon cancer.
Main Methods:
- Caco-2 colon cancer cells were treated with sulforaphane, 5-aza-2'-deoxycytidine (5-Aza), and trichostatin A (TSA).
- DNA methyltransferase 1 (DNMT1) mRNA and protein levels, and activity were assessed.
- Methylation-specific PCR and bisulfite sequencing were used to analyze Nrf2 promoter methylation.
- Nrf2 mRNA and protein expression were quantified using RT-qPCR and Western blot.
Main Results:
- Sulforaphane did not alter DNMT1 mRNA levels but inhibited DNMT1 protein expression when co-administered with 5-Aza and TSA.
- Sulforaphane significantly decreased Nrf2 promoter methylation compared to controls.
- Nrf2 mRNA and protein levels were significantly increased by sulforaphane treatment, indicating enhanced Nrf2 activation.
Conclusions:
- Sulforaphane promotes the demethylation of the Nrf2 promoter region in colon cancer cells.
- This epigenetic modification leads to increased Nrf2 activation.
- Sulforaphane-induced Nrf2 activation contributes to the chemopreventive effects against colon cancer.
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