DNA methylation of claudin-6 promotes breast cancer cell migration and invasion by recruiting MeCP2 and deacetylating H3Ac and H4Ac

Abstract

BackgroundClaudin-6 (CLDN6), a member of claudin transmembrane protein family, has recently been reported to be undetectable or at low levels in human breast cancer cell lines and tissues and plays a role in suppression of migration and invasion in breast cancer cells. In addition, it is reported that CLDN6 expression is regulated by DNA methylation in various human cancers and cell lines. However, it is unclear how DNA methylation regulates CLDN6 expression. Here we show the mechanism by which DNA methylation regulates CLDN6 expression in human breast cancer cell line MCF-7.MethodsRT-PCR, Western blot and immunofluorescent staining were utilized to investigate CLDN6 expression in breast cancer tissues and MCF-7 cells. Methylation-Specific PCR (MSP) was applied to determine DNA methylation status in CLDN6 gene promoter region. Wound-healing assay and invasion assay were utilized to test mobility of MCF-7 cells treated with 5-aza-dC (DNA methyltransferase inhibitor). MeCP2 binding, H3Ac and H4Ac in CLDN6 promoter region were analyzed by ChIP assay. Nuclease accessibility assay was performed for analysis of the chromatin conformation of CLDN6 gene. To study the role of CLDN6 in malignant progression, we used RNAi to knockdown CLDN6 expression in MCF-7 cells treated with 5-aza-dC, and examined the mobility of MCF-7 cells by wound-healing assay and invasion assay.Results5-aza-dC and TSA (histone deacetylase inhibitor) application induced CLDN6 expression in MCF-7 cells respectively and synergistically. 5-aza-dC treatment induced CLDN6 demethylation, inhibited MeCP2 binding to CLDN6 promoter and increased H3Ac and H4Ac in the promoter. In addition, TSA increased H4Ac, not H3Ac in the promoter. The chromatin structure of CLDN6 gene became looser than the control group after treating with 5-aza-dC in MCF-7 cells. 5-aza-dC up-regulated CLDN6 expression and suppressed migration and invasion in MCF-7 cells, whereas CLDN6 silence restored tumor malignance in MCF-7 cells.ConclusionsDNA methylation down-regulates CLDN6 expression through MeCP2 binding to the CLDN6 promoter, deacetylating H3 and H4, and altering chromatin structure, consequently promoting migratory and invasive phenotype in MCF-7 cells.

Global methylation profiles in buccal cells of long-term smokers and moist snuff consumers

“…DNA methylation is a key regulatory mechanism in normal development and disease pathways for several diseases, including cancer (Tsaprouni et al 2014, Zhang et al 2016. Alterations in DNA methylation status have been reported to regulate chromatin structure, and potentially influence transcription (Sneller and Gunter 1987, Kang et al 2015, Liu et al 2016b.…”

Expression of claudin 1, claudin 4, and claudin 7 in colorectal cancer and its relation with CLDN DNA methylation patterns

“…1,2 So far, there are limited data available on claudin (CLDN), epigenetics, and the relation to claudin protein expression as well as its role in the development of colorectal carcinoma (CRC). While DNA methylation of claudin 6 has been shown to be involved in the invasion of breast cancer cells 3 Abstract Altered claudin expression has been described in colon, prostatic, ovarian, and breast carcinoma. However, the role of epigenetic modifications in these genes and their role in colorectal cancer is unknown.…”

MECP2 promotes migration and invasion of gastric cancer cells via modulating the Notch1/c-Myc/mTOR signaling pathways by suppressing FBXW7 transcription

“…Silencing MECP2 reduces human prostate transformed cell proliferation [31]. Recent research has found that MECP2 regulates cancer cell migration and invasion in glioma and breast cancer [32,33]. Our previous studies have proved that MECP2 promotes cell proliferation and cell cycle G1-S transition, and restrains cell apoptosis in liver cancer, gastric cancer and breast cancer [19][20][21][22].…”