Gene expression and targeted RNA sequencing in genomic regions associated with increased endometriosis risk (#20)
Genome-wide association studies have identified genomic regions strongly associated with risk for developing endometriosis (1,2). Candidate genes exist within some regions including wingless-type MMTV integration site family, member 4 (WNT4) on chromosome 1 (2). However, the signals associated with increased risk are located within introns or intergenic regions and the causal variants are most likely to be regulatory. The next step towards developing better diagnostic methods and treatments is to use bioinformatics, gene expression and functional studies to identify the gene or genes in each region regulated by these key variants. Illumina HT-12 v4.0 gene expression arrays have been run on RNA samples from endometrium in 31 women with and without a diagnosis of endometriosis. DNA samples available for most cases will be genotyped to identify any allele specific differences in gene expression. We also used a novel technique, RNA-Capture-Seq (3) to identify all transcripts expressed in RNA samples from endometrium located within the associated regions. The method uses oligonucleotide “tiling” arrays designed to cover the regions of interest. RNA samples are hybridised to the tiling arrays to enrich for coverage of transcripts in each region. Sequencing of the captured products greatly increases the power to identify novel and rare transcripts. We designed tiling arrays to capture transcripts from approximately 1 Mb around key variants in each region and conducted RNA-Capture-Seq on total RNA from the endometrium of eight women with and without a diagnosis of endometriosis. The RNA-Capture-Seq procedure successfully enriched for transcripts within the target regions. Transcripts have been assembled and we have identified a number of novel transcripts within regions previously considered to be intergenic.
- Uno S, Zembutsu H, Hirasawa A, et al. (2010) A genome-wide association study identifies genetic variants in the CDKN2BAS locus associated with endometriosis in Japanese. Nature Genetics 42: 707-710.
- Painter JN, Anderson CA, Nyholt DR, et al. (2011) Genome-wide association study identifies a locus at 7p15.2 associated with endometriosis. Nature Genetics 43 :51-54.
- Mercer TR, Gerhardt DJ, Dinger ME, et al. (2012) Targeted RNA sequencing reveals the deep complexity of the human transcriptome. Nature Biotechnology 30: 99-104.