Allele-specific epigenetic regulation ofFURINexpression at a coronary artery disease susceptibility locus

Abstract

AbstractBackgroundGenome-wide association studies have revealed an association between the genetic variant rs17514846 on chromosome 15q26.1 and coronary artery disease susceptibility. Studies have shown that rs17514846 influences the expression of the FES Upstream Region (FURIN) gene located at this locus in monocytes. We investigated the mechanism through which rs17514846 modulatesFURINexpression.Methods and ResultsAn analysis of isogenic monocytic cell lines with either the rs17514846 C/C or A/A genotype showed that the cells of the A/A genotype expressed higher levels ofFURINthan cells of the C/C genotype. A pyrosequencing methylation analysis showed that the cytosine (in a CpG motif) at the rs17514846 position on the C allele was methylated. Treatment with the DNA methylation inhibitor 5-aza-2’-deoxycytidine increasedFURINexpression. A bioinformatic analysis indicated that the rs17514846 site might interact with the transcription factor MeCP2 that often functions as a gene repressor by binding to methylated CpG sites. An electrophoretic mobility super-shift assay with a probe corresponding to the DNA sequence at and around the rs17514846 position of the C allele detected two DNA-protein complex bands, which were altered by the addition of an anti-MeCP2 antibody in the assay, whilst these DNA-protein complexes were barely detectable with a probe for the A allele. A chromatin immunoprecipitation assay showed an enrichment of the DNA sequence containing the rs17514846 site in chromatin precipitates pulled down by an anti-MeCP2 antibody. siRNA-mediated knockdown of MeCP2 caused an increase inFURINexpression. Furthermore, MeCP2 knockdown increased monocyte migration and proliferation, and this effect was diminished by a FURIN inhibitor.ConclusionsThe results of our study suggest that DNA methylation inhibitsFURINexpression and that the coronary artery disease predisposing variant rs17514846 modulatesFURINexpression and monocyte migration via an allele-specific effect on DNA methylation.