β-Catenin gene promoter hypermethylation by reactive oxygen species correlates with the migratory and invasive potentials of colon cancer cells
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Over half of the colon cancer patients suffer from cancer-related events, mainly metastasis. Loss of β-catenin activity has previously been found to facilitate cancer cell dissociation and migration. Here, we aimed to investigate whether epigenetic silencing of β-catenin induces human colon cancer cell migration and/or invasion.
HCT-116, Caco-2, HT-29 and SW620 cell migration and invasion capacities were assessed using scratch wound healing and Matrigel invasion assays, respectively. Confocal microscopy, qRT-PCR and Western blotting were performed to determine gene expression levels, whereas methylation-specific quantitative real-time PCR was used to assess the extent of β-catenin gene (CTNNB1) promoter methylation after treatment of the cells with TPA, hydrogen peroxide, 5-aza-2′-deoxycytidine and/or VAS2870.
We found that treatment of HT-29 and Caco-2 cells (differentiated and low metastatic) with 12-O-tetradecanoyl phorbol-13-acetate (TPA; a tumor promoter) suppressed E-cadherin and β-catenin expression at both the mRNA and protein levels and, in addition, enhanced cell migration. Furthermore, we found that the CTNNB1 gene promoter methylation levels were higher in the more invasive HCT-116 and SW620 colon cancer cells than in HT-29 and CCD-841 (normal colon epithelial) cells. We also found that TPA or hydrogen peroxide induced CTNNB1 gene promoter methylation to a higher extent in HT-29 and CCD-841 cells than in HCT-116 and SW620 cells, and that the degree of CTNNB1 gene promoter methylation positively correlated with cell dissociation and migration. In addition, we found that co-treatment with 5-aza-2′-deoxycytidine (decitabine, a DNA methyl transferase inhibitor) and VAS2870 (a NADPH oxidase inhibitor) almost completely blocked the invasion of TPA-treated HT-29 and TPA-untreated HCT-116 and SW620 cells, and that these inhibitions surpassed those of the cells treated with decitabine or VAS2870 alone.
From our data we conclude that the extent of CTNNB1 gene promoter methylation by reactive oxygen species correlates with the migratory and invasive abilities of colon cancer cells. Our results suggest that epigenetic regulation of CTNNB1 may serve as a novel avenue to block colon cancer cell migration and invasion.
KeywordsColon cancer CTNNB1 gene promoter hypermethylation Reactive oxygen species NADPH oxidase (NOX) 2 Migration Invasion
This work was supported by the National Research Foundation of Korea (NRF) funded by the Korean Ministry of Science and ICT (grant no. NRF-2017R1E1A1A01073590), and by a Yeungnam University research grant (2017).
Compliance with ethical standards
Conflict of interest
The authors have no conflict of interest to declare.
- 12.X. Tian, Z. Liu, B. Niu, J. Zhang, T.K. Tan, S.R. Lee, Y. Zhao, D.C.H. Harris and G. Zheng, E-cadherin/β-catenin complex and the epithelial barrier. Biomed Res Int 2011, 567305 (2011)Google Scholar
- 23.S.H. Lin, J. Wang, P. Saintigny, C.-C. Wu, U. Giri, J. Zhang, T. Menju, L. Diao, L. Byers and J.N. Weinstein, Genes suppressed by DNA methylation in non-small cell lung cancer reveal the epigenetics of epithelial–mesenchymal transition. BMC Genomics 15, 1079 (2014)CrossRefPubMedPubMedCentralGoogle Scholar
- 33.T. Ikenoue, H. Ijichi, N. Kato, F. Kanai, T. Masaki, W. Rengifo, M. Okamoto, M. Matsumura, T. Kawabe, Y. Shiratori, Analysis of the β-catenin/T cell factor signaling pathway in 36 gastrointestinal and liver Cancer cells. Cancer Sci 93, 1213–1220 (2002)Google Scholar
- 36.B.C. Park, D. Thapa, Y.S. Lee, M.K. Kwak, E.S. Lee, H.G. Choi, C.S. Yong, J.A. Kim, 1-furan-2-yl-3-pyridin-2-yl-propenone inhibits the invasion and migration of HT1080 human fibrosarcoma cells through the inhibition of proMMP-2 activation and down regulation of MMP-9 and MT1-MMP. Eur J Pharmacol 567, 193–197 (2007)CrossRefPubMedGoogle Scholar