MUC2 gene promoter methylation in mucinous and non-mucinous colorectal cancer tissues

Abstract

Abstract. Abundant mucin production and MUC2 expression is the key feature of mucinous colorectal cancer (CRC). Although MUC2 gene methylation has been thought to play an important role in loss of MUC2 expression, the tissues are difficult to analyze because of the cellular heterogeneity of tissue samples. In the present study, we determined the role of region-specific methylation in the MUC2 promoter in MUC2 expression in CRC. Additionally, we optimized the conditions for quantification of methylation analysis in mucinous and non-mucinous CRC tissues. We identified two regions in MUC2 promoter, region A (-289 and -274) and region C (-193 and -160), that correlated with loss of MUC2 expression by comparing the methylation status in 13 CRC cell lines with no or low MUC2 expression and those in 4 cell lines with high MUC2 expression. To prove the correlation of MUC2 methylation status and loss of expression in CRC tissues, MUC2 methylation status in tumors needs to be determined. Since the critical CpG sites have been identified in cell lines by sequencing, a more rapid and sensitive methylation specific PCR (MSP) was used. We conducted MSP at 3 CpG sites (-289, -274, -193) in 19 mucinous and 34 non-mucinous CRC tissues because this analysis worked at only these sites in the preliminary cell line experiments. Our results showed that methylation status of mucinous CRC was significantly lower than that of non-mucinous CRC at 3 sites (-289; p=0.001, -274; p=0.013, -193; p=0.001), and correlated with high level of MUC2 expression as determined by immunohistochemistry. Besides, these results indicated that MUC2 expression and mucin contents decreased in accordance with the increase of methylation status. We concluded that low methylation status of MUC2 gene plays a predominant role in high level MUC2 expression in mucinous CRC.

Low Expression of Mucin 2, High Expression of Mucin 13, and High Expression of Nuclear Factor Kappa-Light-Enhancer of Activated B Cells were Significant Pathways in Colorectal Cancer Development

“…In cancer cells, the cytokines IL-4, IL-3, and TNF-alpha increased, resulting in the downregulation of MUC2. In addition, the carcinogenesis process directly triggers direct methylation of MUC2 in colorectal cancer cells in promoter area A (-289 and -274) and area C (-193 and -160), which in turn downregulate MUC2 and triggers transcription factors (Okudaira et al, 2010). In this study, it was shown that there was a significantly lower MUC2 expression in the cancerous colonic mucosa, which was consistent with other studies.…”

Resistance to Cell Death in Mucinous Colorectal Cancer—A Review

“…Expression of MUC2 is regulated by a number of factors in the colonic epithelium. Analysis of mucin-producing cell lines by Okudaira et al demonstrated that methylation of the MUC2 gene promotor was significantly lower in mucinous CRC lines compared to non-mucinous and correlated with mucin protein expression [24]. Loss of functional p53 is responsible for downregulation of MUC2 expression [25].…”

Mucin in cancer: a stealth cloak for cancer cells

“…These demonstrate that MCC thoroughly exploits the superior abilities of mucins. For example, mucins are tightly controlled by a regulatory mechanism such as promoter methylation and transcription factor ( 10 , 11 ). The expression patterns of MUC2 and MUC5 are similar, but the function appears to be independent, which could be utilized by MCC.…”

“…Given that the Muc2 -deficient mouse developments colitis ( 11 ), methylation of the MUC2 promoter might contribute to CRC initiation. However, the results of MUC2 gene promoter methylation analysis of MCC and non-MCC by narrowing the scope in the entire CRC analysis shows demethylation in the MUC2 promoter of MCC ( 10 ). MCC (10-15% of human CRC) is metastatic and therapeutically resistant ( 12 ) and shows accumulated genetic mutations including K-Ras , TOP-1 , MAPK , and BRAF ( 12 - 18 ).…”

“…MUCIN2 (MUC2): Secreted protein MUC2 is a major com-ponent of the mucus layer in the small and large intestines. Decreased MUC2 is highly involved in the early stage of colorectal cancer (CRC) development, which is caused by methylation of the MUC2 promoter ( 10 ). Given that the Muc2 -deficient mouse developments colitis ( 11 ), methylation of the MUC2 promoter might contribute to CRC initiation.…”