Allelic Imbalance in Regulation of ANRIL through Chromatin Interaction at 9p21 Endometriosis Risk Locus

PLoS Genet. 2016 Apr 7;12(4):e1005893. doi: 10.1371/journal.pgen.1005893. eCollection 2016 Apr.

Abstract

Genome-wide association studies (GWASs) have discovered numerous single nucleotide polymorphisms (SNPs) associated with human complex disorders. However, functional characterization of the disease-associated SNPs remains a formidable challenge. Here we explored regulatory mechanism of a SNP on chromosome 9p21 associated with endometriosis by leveraging "allele-specific" functional genomic approaches. By re-sequencing 1.29 Mb of 9p21 region and scrutinizing DNase-seq data from the ENCODE project, we prioritized rs17761446 as a candidate functional variant that was in perfect linkage disequilibrium with the original GWAS SNP (rs10965235) and located on DNase I hypersensitive site. Chromosome conformation capture followed by high-throughput sequencing revealed that the protective G allele of rs17761446 exerted stronger chromatin interaction with ANRIL promoter. We demonstrated that the protective allele exhibited preferential binding affinities to TCF7L2 and EP300 by bioinformatics and chromatin immunoprecipitation (ChIP) analyses. ChIP assays for histone H3 lysine 27 acetylation and RNA polymerase II reinforced the enhancer activity of the SNP site. The allele specific expression analysis for eutopic endometrial tissues and endometrial carcinoma cell lines showed that rs17761446 was a cis-regulatory variant where G allele was associated with increased ANRIL expression. Our work illuminates the allelic imbalances in a series of transcriptional regulation from factor binding to gene expression mediated by chromatin interaction underlie the molecular mechanism of 9p21 endometriosis risk locus. Functional genomics on common disease will unlock functional aspect of genotype-phenotype correlations in the post-GWAS stage.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Allelic Imbalance*
  • Cell Line, Tumor
  • Chromatin / genetics*
  • Chromosomes, Human, Pair 9*
  • E1A-Associated p300 Protein / metabolism
  • Endometriosis / genetics*
  • Female
  • Genome-Wide Association Study
  • Haplotypes
  • Humans
  • Polymorphism, Single Nucleotide
  • Promoter Regions, Genetic
  • RNA, Long Noncoding / genetics*
  • RNA, Long Noncoding / metabolism
  • Transcription Factor 7-Like 2 Protein / metabolism

Substances

  • CDKN2B antisense RNA, human
  • Chromatin
  • RNA, Long Noncoding
  • TCF7L2 protein, human
  • Transcription Factor 7-Like 2 Protein
  • E1A-Associated p300 Protein
  • EP300 protein, human

Grants and funding

This work was supported by Japan Society for the Promotion of Science (JSPS) Grant-in-Aid for Scientific Research (A) Grant Number 15H02373 (II) and The Tailor-made Medical Treatment Program by the Ministry of Education, Culture, Sports, Science and Technology (MEXT) Grant Number 10100324 (II, and SA). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.