Methyl-CpG binding protein MBD2 is implicated in methylation-mediated suppression of miR-373 in hilar cholangiocarcinoma

Abstract

Abstract. Aberrant expression of miRNAs is associated with particular cancers showing tissue-and clinical-featurespecificity patterns. Some miRNA genes harboring or being embedded in CpG islands undergo methylation mediated silencing. MBP, methyl CpG binding protein, suppresses transcription through binding to methylated CpG dinucleotides. Expression of miR-373 has been reported to be suppressed in malignant bile duct cell lines. Bioinformatic prediction reveals that the transcription start site (TSS) of miR-373 is implanted in a 402 bp canonical CpG island containing 26 CpG dinucleotides. In this study, we aim to determine the epigenetic regulation of miR-373 gene in hilar cholangiocarcinoma. TaqMan microRNA assay shows that downregulation of miR-373 is closely associated with poor cell differentiation, advanced clinical stage and shorter overall and disease-free survival in hilar cholangiocarcinomas. Methylation analysis shows that the promoter-associated CpG island is hypermethylated which is consistent with the inhibition of miR-373. Chromatin immunoprecipitation (ChIP) assay indicates that down-regulation of miR-373 resultes from the selective recruitment of MBD2 to methylated CpG islands. In contrast, MBD2 knock-down by use of a specific siRNA promoted the expression of miR-373. Reactivation of miR-373 by pharmacologic induction of 5-aza-CdR and trichostatin A (TSA) led to decreased enrichment of MBD2 at CpG island regions. Enhanced expression of exogenous MBD2 in stable QBC 939 cells which stably express pGL4-m373-prom induces strengthened recruitment of MBD2. Our findings suggest that miR-373 is a methylationmediated gene and the implication of MBD2 in methylationmediated suppression of miR-373 plays an important role in tumourigenesis and development in hilar cholangiocarcinoma.

Identification of links between small molecules and miRNAs in human cancers based on transcriptional responses

“…The increasing of the miR-129-5p expression was related to antitumor activity of TSA, whereas the inactivation of miR-129-5p significantly blocked the TSA-induced cell death. Chen et al pointed out that miR-373 was reactivated by the pharmacologic induction of TSA26. Zhang et al evaluated the effect of TSA to miRNA expression in BxPC-3 human cancer cell27.…”

MicroRNAs in Brain Tumors

“…All these compounds interact with each other, and miRNAs are regulated by the other parts of this network. One crucial aspect in this context seems to be the interaction of miRNAs with DNA methylation, which has been described to control and "fine tune" various miRNAs by epigenetic inactivation due to aberrant hypermethylation [20][21][22][23][24][25][26][27]. Most interestingly, DNA methylation has been proven to play an important role in drug resistance, and there is effort to assess "DNA methylationtargeted" therapy with drugs such as DNA methyltransferase inhibitors (e.g., decitabine) as potential weapon against cancer [28][29][30][31].…”

Deregulated MicroRNAs in Biliary Tract Cancer: Functional Targets and Potential Biomarkers

“…In this context, Chen et al described MBD2 as a direct target of miRNA-373, a tumor miRNA species downregulated in hilar CC, which was associated with advanced clinical stage [63]. MBD2 is involved in the DNA methylation-dependent repression of gene transcription as a reader of cytosine methylation and was suggested as a marker associated with poor prognosis for patients with hepatocellular carcinoma [64, 65].…”

“…DNA methylation at CpG islands in promoter regions is an epigenetic mechanism that leads to transcriptional repression of the respective gene. Chen et al observed downregulation of miRNA-373 in patients with hilar cholangiocarcinoma [63]. In order to investigate the mechanism of the miRNA-373 deregulation, they analyzed the genomic surrounding of the miRNA-373 gene and identified a region at the 5′ flank that may serve as a promoter.…”