Shidal Chris, Shu Xiang, Wu Jie, Wang Jifeng, Huang Shuya, Long Jirong, Bauer Joshua A, Ping Jie, Guo Xingyi, Zheng Wei, Shu Xiao-Ou, Cai Qiuyin
Vanderbilt Epidemiology Center, Vanderbilt-Ingram Cancer Center, Department of Medicine, Division of Epidemiology, Vanderbilt University School of Medicine, Nashville, TN 37203, USA.
Memorial Sloan Kettering Cancer Center, Department of Epidemiology & Biostatistics, New York, NY 10075, USA.
Cancers (Basel). 2021 Apr 23;13(9):2037. doi: 10.3390/cancers13092037.
We previously identified a locus at 21q22.3, tagged by the single nucleotide polymorphism (SNP) rs35418111, being associated with breast cancer risk at a genome-wide significance level; however, the underlying causal functional variants and gene(s) responsible for this association are unknown. We performed functional genomic analyses to identify potential functional variants and target genes that may mediate this association. Functional annotation for SNPs in high linkage disequilibrium (LD, r > 0.8) with rs35418111 in Asians showed evidence of promoter and/or enhancer activities, including rs35418111, rs2078203, rs8134832, rs57385578, and rs8126917. These five variants were assessed for interactions with nuclear proteins by electrophoretic mobility shift assays. Our results showed that the risk alleles for rs2078203 and rs35418111 altered DNA-protein interaction patterns. Cis-expression quantitative loci (cis-eQTL) analysis, using data from the Genotype-Tissue Expression database (GTEx) European-ancestry female normal breast tissue, indicated that the risk allele of rs35418111 was associated with a decreased expression of the gene, a relatively uncharacterized endoribonuclease in humans. We investigated the biological effects of on breast cancer cell lines by transient knock-down of expression in MCF-7, T47D, and MDA-MB-231 cell lines. Knockdown of mRNA in breast cancer cell lines consistently decreased cell proliferation, colony formation, and migration/invasion, regardless of estrogen receptor status. We performed RNA sequencing in MDA-MB-231 cells transfected with siRNA targeting and subsequent gene set enrichment analysis to identify gene networks associated with knockdown. These data indicated was involved in networks associated with inflammation and metabolism. Finally, we showed trends in expression patterns in breast tissues from The Cancer Genome Atlas (TCGA); early-stage breast cancers had elevated expression compared with normal tissue, but significantly decreased expression in late-stage disease. Our study provides evidence of a significant role for the human gene in breast cancer pathogenesis and the association between the rs35418111/21q22.3 locus and breast cancer risk, which may be mediated through functional SNPs, rs35418111 and rs2078203, that regulate expression of .
我们之前在21q22.3区域鉴定出一个位点，由单核苷酸多态性（SNP）rs35418111标记，该位点在全基因组显著水平上与乳腺癌风险相关；然而，导致这种关联的潜在因果功能变异和基因尚不清楚。我们进行了功能基因组分析，以鉴定可能介导这种关联的潜在功能变异和靶基因。对亚洲人群中与rs35418111处于高连锁不平衡（LD，r>0.8）的SNP进行功能注释，结果显示包括rs35418111、rs2078203、rs8134832、rs57385578和rs8126917在内的这些SNP具有启动子和/或增强子活性的证据。通过电泳迁移率变动分析评估了这五个变异与核蛋白的相互作用。我们的结果表明，rs2078203和rs35418111的风险等位基因改变了DNA-蛋白质相互作用模式。使用来自基因型-组织表达数据库（GTEx）欧洲血统女性正常乳腺组织的数据进行顺式表达定量位点（cis-eQTL）分析表明，rs35418111的风险等位基因与一个基因的表达降低相关，该基因是人类中一个相对未被充分表征的核糖核酸内切酶。我们通过在MCF-7、T47D和MDA-MB-231细胞系中瞬时敲低该基因的表达，研究了其对乳腺癌细胞系的生物学效应。在乳腺癌细胞系中敲低该基因的mRNA，无论雌激素受体状态如何，均能持续降低细胞增殖、集落形成以及迁移/侵袭能力。我们在转染了靶向该基因的siRNA的MDA-MB-231细胞中进行了RNA测序，并随后进行基因集富集分析，以鉴定与该基因敲低相关的基因网络。这些数据表明该基因参与了与炎症和代谢相关的网络。最后，我们展示了来自癌症基因组图谱（TCGA）的乳腺组织中该基因的表达模式趋势；早期乳腺癌与正常组织相比，该基因表达升高，但在晚期疾病中表达显著降低。我们的研究提供了证据，证明人类该基因在乳腺癌发病机制中具有重要作用，以及rs35418111/21q22.3位点与乳腺癌风险之间的关联，这可能是通过调控该基因表达的功能性SNP，即rs35418111和rs2078203介导的。
