COMT expression protein level and activity are down
COMT expression, protein level and activity are down regulated by E2. Using estrogen receptor (ER) positive MCF-7 cells, Xie et al. demonstrated an E2 dose dependent (1, 10 and 100 nM) decrease in S-COMT 1.3 kb mRNA that was maximum after 48 h (Xie et al., 1999). A similar decrease was not observed in ER negative HeLa cells. Their further analysis revealed that the E2-induced down regulation was related to the presence of two half palindromic estrogen response elements (EREs) in the promoter for S-COMT and two putative CAAT/enhancer binding protein (C/EBP) sites in the promoter for MB-COMT.
Although inhibition of COMT activity has been suggested to increase oxidative DNA damage and DNA adducts in MCF-7 Fmoc-Glu(OtBu)-OPfp and could be a risk factor for estrogen-associated cancers, mechanisms underlying regulation of COMT gene expression in human breast cancer cells were not fully understood. It has been suggested that DNA methylation at COMT promoters associate with the decrease in their transcript levels. The role of COMT gene methylation in disease pathogenesis has been investigated with mixed results in relation to various diseases (Norrholm et al., 2013; Teroganova et al., 2016 and van der Knaap et al., 2014). Especially, Swift-Scanlan and Smith et al. reported that in unspecified human breast cells lines, COMT gene expression was inversely correlated with CpG methylation within exon 3, which contains the translational start sites (ATG) for both MB- and S-COMT (Swift-Scanlan et al., 2014). On the other hand, COMT methylation was changed in respond to xenoestrogens. Nahar et al., 2014 demonstrated an association of increased site-specific methylation within the 5’ MB-COMT regulatory region with higher levels of a xenoestrogen, bispenol A (BPA), in fetal human liver specimens. Genistein (GEN) is another xenoestrogen commonly found in large quantities in soybeans. It has been well documented that GEN induced changes in DNA methylation in tissues of reproductive organs (Tang et al., 2008; Rietjens et al., 2013; Guerrero-Bosagna and Skinner, 2014). GEN was shown to influence COMT activity in MCF-7 cells (Lehmann et al., 2008). However, in this case, the relationship between inhibition of COMT activity and COMT promoter methylation has not been studied.
In this present study, we aimed to determine the mechanisms of which E2 and GEN regulate COMT gene expression, via epigenetic modifications at the 5′ regulatory region of COMT in MCF-7 cells. Sulforaphane (SFN) is an inducer of phase II protective enzymes including glutathione-S-transferases and quinone reductase. Previously, it was reported that SFN (Nrf2 activator) may alter estrogen metabolism in breast MCF-10A cells and thus protect against estrogen-mediated DNA damage through alteration of E2 metabolizing enzymes such as COMT, NQO1 and GSTA1 (Yang et al., 2013). In this study, we investigated whether SFN would modulate the epigenetic regulation of COMT transcription. Data would provide new insights in regulating E2/E1 metabolism in human breast cancer cells, which may ultimately modulate breast cancer risk.
Materials and methods
Discussion The goal of this study was to investigate the mechanisms underlying regulation of COMT transcription in MCF-7 cells given the fact that inhibition of COMT activity may result in the formation of DNA adducts and oxidative DNA damage which contributes to the initiation, promotion and progression of BCa (Yager et al., 2016). Estradiol or xenoestrogen has been shown to decrease COMT expression in breast epithelial cells (Lavigne et al., 2001; Zahid et al., 2007; Lehmann et al., 2008; Yue et al., 2013). In this study, we performed a mechanistic study at the COMT promoter and illustrated how E2 and GEN modulated the COMT transcription via epigenetic modifications at specific COMT promoter. E2 and GEN suppressed COMT gene expression in association with alterations in CpG site-specific DNA methylation and binding of DNA methylation-related proteins (5′ cytosine methylation) and histone modifying enzymes (N′ terminal deacetylation) at the MB-COMT distal promoter but not at other COMT promoters (MB-COMT and S-COMT promoters). It is supported by evidences that cancer cells may use multiple COMT promoters to selectively inactive COMT during development of cancer (Sasaki et al., 2003). On the other hand, Xie et al. reported that ER and C/EBP contributed to transcriptional activity of S-COMT promoter and MB-COMT promoter respectively (Xie et al., 1999). However, it is unclear if these transcription factors co-operate with enzymes modifying DNA methylation and histone acetylation at these promoters and results in selectively gene silencing of COMT. It requires future experiment to prove this hypothesis.