Abstract 4487: An INDEL variant confers melanoma risk through PARP1 expression regulation
Authors: Choi Jiyeon , Makowski Matthew , Zhang Tongwu , Law Matthew , Kim Wendy , Kovacs Michael , Parikh Hemang , Aoude Lauren , Gartside Michael , Trent Jeffrey , Vermeulen Michiel , Macgregor Stuart , Hayward Nicholas , Xu Mai , Brown Kevin 
Published: July 2016

Abstract
Recent genome wide association studies identified several new loci for melanoma susceptibility including chr1q42.1 locus encompassing Poly [ADP-ribose] polymerase 1 (PARP1). As an effort to identify effecter genes and functional risk variants from this locus we performed expression quantitative trait loci (eQTL) analysis in 62 melanoma cell lines. Among the genes in 1Mb area eQTL was identified for PARP1 and subsequently validated by qPCR, where increased PARP1 levels are significantly correlated with the risk allele (p = 0.03, copy number adjusted). Further allele discrimination qPCR of PARP1 transcripts in 14 melanoma cell lines heterozygous and of neutral copy for the lead SNP indicated significantly higher proportion for the risk allele (p = .0001). Same allelic imbalance was also observed in 51 heterozygous melanomas with neutral copy numbers from The Cancer Genome Atlas (p = .028). To identify functional risk variants mediating these effects we annotated this locus using six melanoma relevant cell types available from ENCODE and Roadmap database. Based on recent fine mapping data suggesting single-SNP model for this locus we prioritized high LD variants for nomination. Among 65 SNPs of high LD with the lead SNP or imputed best SNP (r2&gt;0.6 using 1000 Genomes phase3), four exhibited strong evidence as potential transcriptional enhancers. Electro Mobility Shift Assays and luciferase assays on these four variants demonstrated that one of them, a six-base pair INDEL (-/GGGCCC) in the first intron, displayed strong allelic functionality. Consistent with the expression data melanoma-associated deletion allele results in higher luciferase activity while protective insertion allele binds a group of proteins with higher affinity. Subsequent comparative mass-spectrometry for these insertion-binding proteins identified a striking collection of Guanine-quadruplex binding proteins including RECQ1 as potential inhibitors of PARP1 expression. Consistent with this hypothesis over-expression of RECQL results in more pronounced allelic difference in luciferase activities indicating that RECQ1 contributes to allelic PARP1 expression. Further interrogation of RECQ1 and PARP1 expression correlation analysis suggested that RECQ1 levels are inversely correlated with PARP1 in melanomas carrying insertion alleles. These data demonstrate that increased PARP1 expression is correlated with melanoma risk and an INDEL variant mediates differential PARP1 expression possibly through secondary DNA structure binding proteins including RECQ1.
Citation Format: Jiyeon Choi, Matthew Makowski, Tongwu Zhang, Matthew Law, Wendy Kim, Michael Kovacs, Hemang Parikh, Lauren Aoude, Michael Gartside, Jeffrey Trent, Michiel Vermeulen, Stuart Macgregor, Nicholas Hayward, Mai Xu, Kevin Brown. An INDEL variant confers melanoma risk through PARP1 expression regulation. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4487.