Relationship between CAD Risk Genotype in the Chromosome 9p21 Locus and Gene Expression. Identification of Eight New ANRIL Splice Variants

Abstract

BackgroundSeveral genome-wide association studies have recently linked a group of single nucleotide polymorphisms in the 9p21 region with cardiovascular disease. The molecular mechanisms of this link are not fully understood. We investigated five different expression microarray datasets in order to determine if the genotype had effect on expression of any gene transcript in aorta, mammary artery, carotid plaque and lymphoblastoid cells.Methodology/Principal FindingsAfter multiple testing correction, no genes were found to have relation to the rs2891168 risk genotype, either on a genome-wide scale or on a regional (8 MB) scale. The neighbouring ANRIL gene was found to have eight novel transcript variants not previously known from literature and these varied by tissue type. We therefore performed a detailed probe-level analysis and found small stretches of significant relation to genotype but no consistent associations. In all investigated tissues we found an inverse correlation between ANRIL and the MTAP gene and a positive correlation between ANRIL and CDKN2A and CDKN2B.Conclusions/SignificanceInvestigation of relation of the risk genotype to gene expression is complicated by the transcript complexity of the locus. With our investigation of a range of relevant tissue we wish to underscore the need for careful attention to the complexity of the alternative splicing issues in the region and its implications to the design of future gene expression studies.

Genome-wide association study identifies susceptibility loci for open angle glaucoma at TMCO1 and CDKN2B-AS1

“…3B). This is consistent with expression of more than one CDKN2B-AS1 splice variants in a tissue or cell line 11,12 . We utilized well characterized antibodies directed against CDKN2A, CDKN2B, MTAP and TMCO1 to explore the distribution of these proteins in rat retina.…”

“…The expected size of each PCR product is indicated in Supplementary Table 5B 11,12 . The full-length variant (DQ485453) and alternatively spliced variants (DQ485454 and GQ495924) 11,12 were undetectable in human retina (data not shown). M, molecular weight markers in basepairs; RT -, reverse transcription negative control; -ve C, PCR negative control.…”

Long non-coding RNA ANRIL regulates inflammatory responses as a novel component of NF-κB pathway

“…11,12 ANRIL gene is transcribed and spliced in a complex pattern. Multiple isoforms of ANRIL have been identified 13,14 and they are mainly grouped into 2 categories including short forms terminating with exon 13 (short ANRIL isoforms, SANRIL) and long forms which lack exon 13 and terminate with exon 19 and exon 20 (long ANRIL isoforms, LANRIL). The chromosome 9p21 CAD risk haplotype containing multiple single nucleotide polymorphisms (SNP) in tight linkage disequilibrium resides at the 3 0 end of ANRIL gene and exhibits a clear association with ANRIL expression.…”

Macro lncRNAs

“…99 The resulting altered regenerative capacity could explain some ANRIL-mediated age-associated diseases such as type II diabetes, atherosclerosis and cancer. 100 More than eight ANRIL splice variants are known 101 and it was proposed that SNPs in the ANRIL gene body could regulate alternative splicing of ANRIL and thus the expression of p16 INK4A and p15 INK4B in diseases that rely on a certain replication potential. 99 The SNPs linked to coronary artery disease are located in a 58 kb interval on human chromosome 9p21 (Fig.…”

“…3C) and the deletion of the orthologous 70 kb region on mouse chromosome 7 showed that expression levels of p16 INK4A and p15 INK4B are markedly reduced in cis. 102 Despite the existence of at least eight non-coding splice variants, 101 the functions of unspliced ANRIL in regulating p15 INK4B in cis cannot be dismissed. 91 …”