Functional Genomics Identify a Regulatory Risk Variation rs4420550 in the 16p11.2 Schizophrenia-Associated Locus
Functional Genomics Identify a Regulatory Risk Variation rs4420550 in the 16p11.2 Schizophrenia-Associated Locus
复制标题
功能基因组学鉴定 16p11.2 精神分裂症相关基因座中的监管风险变异 rs4420550
DOI：
10.1016/j.biopsych.2020.09.016
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发表时间：
2021-02-01
影响因子：
10.6
通讯作者：
Xiao, Xiao
中科院分区：
文献类型：
--
作者：
Chang, Hong;Cai, Xin;Xiao, Xiao
背景：全基因组关联研究（GWASs）已经报道了数百个与精神分裂症相关的基因组位点，然而识别功能风险变异是阐明潜在机制的关键一步。方法：我们应用了多种生物信息学和分子方法，包括表达数量性状位点分析、表观基因组特征鉴定、荧光素酶报告基因测定、染色质构象捕获、同源定向基因组编辑CRISPR/Cas9（聚集规律间隔短回文重复序列/Cas9）、RNA测序和ATAC-Seq（利用测序方法检测转座酶可及的染色质）。结果：我们发现16p11.2的精神分裂症GWAS风险变异与人脑多个基因的信使RNA水平显著相关，其中一个主要表达数量性状位点基因MAPK3位于基因组中距离这些风险变异约200kb的位置。基于人脑和细胞系表观基因组标记的进一步分析表明，非编码单核苷酸多态性rs4420550（在精神分裂症GWAS中p = 2.36 × 10(-9)）位于DNA增强子区域内，通过体外荧光素酶报告基因试验验证了这一点。染色质构象捕获实验表明，rs4420550区域与MAPK3启动子和TAOK2启动子发生物理相互作用。对细胞中单个碱基对进行精确的CRISPR/Cas9编辑，然后进行RNA测序，进一步证实了rs4420550对16p11.2基因转录的调控作用，ATAC-Seq证实rs4420550影响16p11.2区域染色质的可及性。rs4420550-[A/A]细胞的增殖率明显高于rs4420550-[G/G]细胞。结论：这些结果表明rs4420550是一个功能风险变异，本研究为精神分裂症GWAS风险位点的综合功能表征提供了一个例子。
BACKGROUND: Genome-wide association studies (GWASs) have reported hundreds of genomic loci associated with schizophrenia, yet identifying the functional risk variations is a key step in elucidating the underlying mechanisms.METHODS: We applied multiple bioinformatics and molecular approaches, including expression quantitative trait loci analyses, epigenome signature identification, luciferase reporter assay, chromatin conformation capture, homology-directed genome editing by CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/Cas9), RNA sequencing, and ATAC-Seq (assay for transposase-accessible chromatin using sequencing).RESULTS: We found that the schizophrenia GWAS risk variations at 16p11.2 were significantly associated with messenger RNA levels of multiple genes in human brain, and one of the leading expression quantitative trait loci genes, MAPK3, is located similar to 200 kb away from these risk variations in the genome. Further analyses based on the epigenome marks in human brain and cell lines suggested that a noncoding single nucleotide polymorphism, rs4420550 (p = 2.36 x 10(-9) in schizophrenia GWAS), was within a DNA enhancer region, which was validated via in vitro luciferase reporter assays. The chromatin conformation capture experiment showed that the rs4420550 region physically interacted with the MAPK3 promoter and TAOK2 promoter. Precise CRISPR/Cas9 editing of a single base pair in cells followed by RNA sequencing further confirmed the regulatory effects of rs4420550 on the transcription of 16p11.2 genes, and ATAC-Seq demonstrated that rs4420550 affected chromatin accessibility at the 16p11.2 region. The rs4420550-[A/A] cells showed significantly higher proliferation rates compared with rs4420550-[G/G] cells.CONCLUSIONS: These results together suggest that rs4420550 is a functional risk variation, and this study illustrates an example of comprehensive functional characterization of schizophrenia GWAS risk loci.