Abstract 2555: Tissue-specific regulation of alternative splicing by breast cancer associated genetic variants identified in recent GWAS.
Authors: Wagner Erin K. , Shevate Swapnil Kanifnath , Liu Yunlong , He Chunyan 
Published: August 2014

Abstract
Recent genome-wide association studies (GWAS) have identified more than 30 novel loci associated with breast cancer. However, the biological functions of these genetic variants have not been characterized yet. Alternative splicing is one mechanism through which genetic variants may act on gene expression. We hypothesize that the associated genetic variants influence tissue-specific regulation of alternative splicing leading to misregulation of specific isoforms involved in breast tumorigenesis. We performed an integrated analysis of genomic variation and transcriptome sequencing to elucidate alternatively spliced isoform expression quantitative trait loci (asQTL) and to identify breast cancer susceptibility genes whose isoform-specific expression were influenced by the candidate genetic variants. The current study utilized a unique source of paired blood and normal breast tissue from 20 healthy women free of diagnosed breast cancer. We genotyped 42 SNPs that were associated with breast cancer in previous GWAS reports, and assessed expression of approximately 35,000 known splicing variants in more than 24,000 gene transcripts in normal breast tissue using RNA sequencing data. Analysis of variance (ANOVA) was performed to assess the differences of isoform-specific expression across genotypes. After account of age, race and batch effects, we found 384 unique genes whose isoform-specific expressions in normal breast tissue were correlated with the 42 breast cancer candidate SNPs (Bonferroni corrected p &lt; 0.05). The majority of the identified gene isoforms (&gt;90%) were associated with the candidate SNPs in trans, i.e. isoform-specific expression were influenced by SNPs on a different chromosome. In addition, 27 out of the 384 identified genes also showed differential isoform-specific expression in breast tumor vs. normal tissue in our separate analyses. We have identified known cancer genes including SOD1, NPM1, FAU, GABARAP, and SMAD5, as well as novel putative breast cancer genes including PCBP2, SAP18, ATPF7IP, and N4BP2L2. An ingenuity pathway analysis of the 27 genes suggested three functional networks. Network 1, related to “Cell Death and Survival, Cell Cycle, and Gene Expression," includes genes EIF4A1, GABARAP, HARS, HNRNPC and USP33; network 2, related to “Organ Morphology, Reproductive
System Development of Function, and Embryonic Development," included genes ATF7IP, NPM1, RAB2A, SMAD5, and SOD1; network 3, related to “Cancer, Neurological Disease, Cell Death and Survival," includes gene FAM45A. Our results identify known and novel putative breast cancer susceptibility genes whose isoform-specific expression were influenced by breast cancer candidate SNPs from previous GWAS, showing the impact of germline variants on the regulation of alternative splicing in transcriptome.
Citation Format: Erin K. Wagner, Swapnil Kanifnath Shevate, Yunlong Liu, Chunyan He. Tissue-specific regulation of alternative splicing by breast cancer associated genetic variants identified in recent GWAS. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2555. doi:10.1158/1538-7445.AM2013-2555