The Association Exploration Of Functional Variants In Transposase Accessible Chromatin Regions And Susceptibility Of Non-small Cell Lung Cancer In Chinese Han Population

Objective:To identify functional polymorphisms in accessible open chromatin regions within genome-wide association study（GWAS）loci which contributes to non-small cell lung cancer（NSCLC）susceptibility,and to explore the biological functions of casual variants in the etiology of NSCLC.Methods:We selected candidate variants by performing an integrative analysis of assay for transposase accessible chromatin with high-throughput sequencing（ATAC-seq）data and acetyl-histone H3 lysine 27（H3K27ac）marks in chromatin immunoprecipitation followed by sequencing（Ch IP-seq）data,with a combination of lung cancer GWAS signal.A two-stage case-control study with 1830 cases and 2001controls was conducted to test their associations with NSCLC risk.Based on bioinformatics function prediction,dual-luciferase reporter assays and electrophoretic mobility shift assays were performed to explore the biological functions of promising variants in NSCLC tumorigenesis.Results:Sixteen potential functional single-nucleotide polymorphisms（SNPs）were selected as candidates from the integrative analysis of ATAC-seq,Ch IP-seq and GWAS signals.Three marginally significant variants survived in stage 1 case-control study containing 348 cases and 479 controls,including rs13064999（OR=1.21,95%CI:0.98-1.48,P=0.072）,rs12752（OR=0.83,95%CI:0.67-1.03,P=0.097）and rs62085661（OR=0.82,95%CI:0.66-1.02,P=0.072）.Only the positive results of rs13064999（OR=1.16,95%CI:1.04-1.29,P=0.006）were successfully validated in the analyses of stage 2 case-control study containing 1482 cases and 1522controls.In combined analyses,the associations between rs13064999 and NSCLC risk remained significant（OR=1.17,95%CI:1.07-1.29,P=0.001）.Dual-luciferase reporter assays revealed a significant increase of luciferase expression in rs13064999 A allele,when comparing to G allele(P_A549<0.001,P_SK-MES-1=0.004).Further electrophoretic mobility shift assays（EMSA）confirmed a stronger affinity of HP1γto the motif containing SNP rs13064999 A allele than the other allele.Conclusion:Based on the pointcut of chromatin accessibility,through integrative analyses with ATAC-seq and Ch IP-seq data,two-stage association studies and biological functional assays,we firstly found that the functional variant rs13064999contributes to the susceptibility of NSCLC by affecting HP1γbinding,and regulating the transcription of target gene.