Mutual regulation between microRNA-373 and methyl-CpG-binding domain protein 2 in hilar cholangiocarcinoma

World J Gastroenterol. 2012 Aug 7;18(29):3849-61. doi: 10.3748/wjg.v18.i29.3849.


Aim: To investigate the reciprocal modulation between microRNA (miRNA) and DNA methylation via exploring the correlation between miR-373 and methyl-CpG-binding domain protein (MBD)2.

Methods: MiR-373 expression was examined using the TaqMan miRNA assay. Methylation of miR-373 was investigated using methylation-specific polymerase chain reaction, and recruitment of methyl binding proteins was studied using the chromatin immunoprecipitation assay. Mutation analysis was conducted using the QuikChange™ Site-Directed Mutagenesis kit. The activity of miR-373 gene promoter constructs and targeting at MBD2-three prime untranslated region (3'UTR) by miR-373 were evaluated by a dual-luciferase reporter gene assay.

Results: In hilar cholangiocarcinoma, miR-373 decreased and was closely associated with poor cell differentiation, advanced clinical stage, and shorter survival. The promoter-associated CpG island of miR-373 gene was hypermethylated and inhibited expression of miR-373. MBD2 was up-regulated and enriched at the promoter-associated CpG island of miR-373. Methylation-mediated suppression of miR-373 required MBD2 enrichment at the promoter-associated CpG island, and miR-373 negatively regulated MBD2 expression through targeting the 3'UTR.

Conclusion: MiR-373 behaves as a direct transcriptional target and negative regulator of MBD2 activity through a feedback loop of CpG island methylation.

Keywords: Hilar cholangiocarcinoma; Methyl-CpG binding domain proteins 2; Methylation; MicroRNA-373; Three prime untranslated region.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 3' Untranslated Regions
  • Base Sequence
  • Bile Duct Neoplasms / genetics*
  • Bile Duct Neoplasms / metabolism*
  • Bile Duct Neoplasms / pathology
  • Cell Line, Tumor
  • Cholangiocarcinoma / genetics*
  • Cholangiocarcinoma / metabolism*
  • Cholangiocarcinoma / pathology
  • CpG Islands
  • DNA Methylation / genetics
  • DNA-Binding Proteins / metabolism*
  • Female
  • Gene Expression Regulation, Neoplastic
  • Hepatic Duct, Common*
  • Humans
  • Klatskin Tumor / genetics*
  • Klatskin Tumor / metabolism*
  • Klatskin Tumor / pathology
  • Male
  • MicroRNAs / genetics*
  • MicroRNAs / metabolism*
  • Middle Aged
  • Models, Biological
  • Molecular Sequence Data
  • Promoter Regions, Genetic
  • RNA, Neoplasm / genetics*
  • RNA, Neoplasm / metabolism*


  • 3' Untranslated Regions
  • DNA-Binding Proteins
  • MBD2 protein, human
  • MIRN373 microRNA, human
  • MicroRNAs
  • RNA, Neoplasm