Alu elements in ANRIL non-coding RNA at chromosome 9p21 modulate atherogenic cell functions through trans-regulation of gene networks

PLoS Genet. 2013;9(7):e1003588. doi: 10.1371/journal.pgen.1003588. Epub 2013 Jul 4.

Abstract

The chromosome 9p21 (Chr9p21) locus of coronary artery disease has been identified in the first surge of genome-wide association and is the strongest genetic factor of atherosclerosis known today. Chr9p21 encodes the long non-coding RNA (ncRNA) antisense non-coding RNA in the INK4 locus (ANRIL). ANRIL expression is associated with the Chr9p21 genotype and correlated with atherosclerosis severity. Here, we report on the molecular mechanisms through which ANRIL regulates target-genes in trans, leading to increased cell proliferation, increased cell adhesion and decreased apoptosis, which are all essential mechanisms of atherogenesis. Importantly, trans-regulation was dependent on Alu motifs, which marked the promoters of ANRIL target genes and were mirrored in ANRIL RNA transcripts. ANRIL bound Polycomb group proteins that were highly enriched in the proximity of Alu motifs across the genome and were recruited to promoters of target genes upon ANRIL over-expression. The functional relevance of Alu motifs in ANRIL was confirmed by deletion and mutagenesis, reversing trans-regulation and atherogenic cell functions. ANRIL-regulated networks were confirmed in 2280 individuals with and without coronary artery disease and functionally validated in primary cells from patients carrying the Chr9p21 risk allele. Our study provides a molecular mechanism for pro-atherogenic effects of ANRIL at Chr9p21 and suggests a novel role for Alu elements in epigenetic gene regulation by long ncRNAs.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alu Elements / genetics*
  • Apoptosis / genetics
  • Atherosclerosis / genetics*
  • Atherosclerosis / pathology
  • Cell Adhesion / genetics
  • Cell Proliferation
  • Chromosomes, Human, Pair 9 / genetics
  • Coronary Artery Disease / genetics*
  • Coronary Artery Disease / pathology
  • Epigenesis, Genetic
  • Gene Expression Regulation
  • Gene Regulatory Networks
  • Genetic Predisposition to Disease
  • Genome-Wide Association Study
  • HEK293 Cells
  • Humans
  • Polycomb-Group Proteins
  • Polymorphism, Single Nucleotide
  • RNA, Long Noncoding / genetics*

Substances

  • CDKN2B antisense RNA, human
  • Polycomb-Group Proteins
  • RNA, Long Noncoding

Grant support

This study was supported by MSD SHARP & DOHME scholarship (MSD-Stipendium 2010-Arteriosklerose, Wilhelm-Stoffel-Stipendium; LMH), and LIFE – Leipzig Research Center for Civilization Diseases, Universität Leipzig (LMH SH DL MS KK KF FB SG GS PFS JT DT). LIFE is funded by the European Union, the European Regional Development Fund (ERDF) and the Free State of Saxony within its initiative of excellence. Part of the study was funded by a grant from the German Ministry of Education and Research through NGFN-plus (DT). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.