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在microRNA结合部位中搜索和功能分析基因变异,使用大规模并行记者试验

E Yu Rykova¹, N I Ershov², A O Degtyareva²

¹Federal Research Center Institute of Cytology and Genetics, Siberian Branch of the Russian Academy of Sciences, Novosibirsk, Russia. rykova.elena.2014@gmail.com.

Bulletin of experimental biology and medicine

|May 9, 2024

概括

这项研究确定了影响等位基因特异性表达 (AE SNPs) 的遗传变异及其对微RNA结合部位的影响. 使用大规模并行记者试验 (MPRA) 的功能验证证实了它们在基因调节中的重要性.

科学领域:

基因组学就是基因组学.
分子生物学分子生物学
生物信息学是一种生物信息学.

背景情况:

遗传变异可以影响基因表达和调节.
微RNAs (miRNAs) 在转录后基因沉默中发挥着至关重要的作用,通过与3'-未翻译区域 (3'-UTRs) 结合.
了解基因变异,miRNA结合和基因表达之间的相互作用对于破译复杂的生物过程至关重要.

研究的目的:

确定与等位基因特异性表达 (AE SNPs) 相关的遗传变异,这些变异也会影响3'-UTRs中的miRNA结合位.
通过大规模并行记者测试 (MPRA) 功能性地评估这些AE SNPs对miRNA结合和基因调节的意义.

主要方法:

利用GTEx分析V8数据在人类肝脏组织中识别表达定量特征位置 (eQTL).
利用TargetScanHuman 7.0和PolymiRTS数据库来预测受eQTL-SNPs影响的miRNA结合部位.
根据Ago2-CLIP-seq,CLEAR-CLIP和eCLIP-seq数据选择的功能性miRNA结合位.
为选定的AE SNP构建了一个具有主要和替代基因的等离子体库 (102个构造).
通过转移HepG2细胞和测序向DNA和RNA进行MPRA,以检测6个SNP的等位基因特异性表达.

主要成果:

在3'-UTR中确定了4394个多态位置,代表629,559个SNP-基因对的eQTL.
选择了51个受eQTL-SNP影响的miRNA结合部位,具有强有力的功能证据.
使用MPRA在HepG2细胞中成功检测了6个AE SNPs的等位基特异性表达.

结论:

影响miRNA结合部位的AE SNPs是普遍存在的,并且具有功能意义.
MPRA是验证遗传变异对基因调节的功能影响的强大工具.
这些发现有助于理解基因表达调节的遗传基础及其对人类疾病的影响.

关键词:

大规模并行报告测试 (MPRA)微RNA结合位点是微RNA的结合位点.单个核酸的替代.

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