ATP7B基因同义变异通过影响外显子剪接增强子导致mRNA剪接异常  

Synonymous variants of the ATP7B gene may cause abnormal splicing of mRNA by affecting the exonic splicing enhancers

作　　者：周霄颖 郑必霞[1] 刘志峰[1] 金玉[1] Zhou Xiaoying;Zheng Bixia;Liu Zhifeng;Jin Yu(Department of Gastroenterology,the Affiliated Children’s Hospital of Nanjing Medical University,Nanjiang,Jiangsu 210008,China)

机构地区：[1]南京医科大学附属儿童医院消化科,210008

出　　处：《中华医学遗传学杂志》2020年第11期1236-1240,共5页Chinese Journal of Medical Genetics

基　　金：国家自然科学基金(81873542,81570470)。

摘　　要：目的探讨ATP7B基因罕见同义变异对其前体mRNA剪接的影响。方法从ExAc数据库中筛选出人群等位基因频率<0.005的248种罕见同义变异,用Human Splicing Finder(HSF)软件分析其对于前体mRNA剪接的影响,进一步用ESE Finder 3.0软件预测其对于反式作用因子SR蛋白家族结合能力的影响。筛选出同时影响两种或以上SR蛋白结合的罕见同义变异,用体外迷你基因剪接报告系统进行验证。结果HSF分析提示有136种罕见同义变异可能破坏外显子剪接增强子(exonic splicing enhancer,ESE)的基序;ESE Finder 3.0分析提示其中19种会同时影响两种或以上SR蛋白的结合。体外迷你基因实验证实其中c.1620C>T(p.L540L)和c.3888C>T(p.A1296A)变异可导致相应外显子的剪接异常,分别造成第4外显子的完全跳跃以及使第18外显子的跳跃增加25%。结论同义变异可能通过各种途径影响前体mRNA的剪接,其中以破坏ESEs基序最为常见。本研究结果证实c.1620C>T(p.L540L)和c.3888C>T(p.A1296A)变异会对ATP7B基因的mRNA剪接产生影响,使相应的外显子发生跳跃,从而为携带者的基因诊断和遗传咨询提供了依据。Objective To explore the effect of rare synonymous variants of the ATP7B gene on the splicing of its precursor mRNA.Methods A total of 248 rare synonymous variants with allelic frequency of<0.005 were retrieved from the ExAc database.Human Splicing Finder(HSF)was used to predict their effect on the splicing of precursor mRNA.And ESE Finder 3.0 was used to predict the effect of such variants on the binding ability of SR protein family.Rare synonymous variants affecting the binding of two or more SR proteins were selected and verified with an in vitro mini gene splicing report system.Results HSF analysis indicated that 136 of the 248 rare synonymous variants may destroy the exonic splicing enhancer(ESE)motif.Analysis using ESE Finder 3.0 indicated that 19 of them may affect the binding of two or more SR proteins at the same time.In vitro mini gene experiment confirmed that the c.1620C>T(p.L540L)and c.3888C>T(p.A1296A)variants could lead to abnormal splicing of the corresponding exons,resulting in complete skipping of exon 4 and 25%increase in the skipping of exon 18,respectively.Conclusion Synonymous variants may affect the splicing of precursor mRNA in various ways,particularly the destruction of ESE motif.This study confirmed that the c.1620C>T(p.L540L)and c.3888C>T(p.A1296A)variants can affect the mRNA splicing of the ATP7B gene,resulting in skipping of corresponding exons,which may provide a basis for genetic diagnosis and consultation of carriers.

关 键 词：ATP7B基因 同义变异 外显子剪接增强子 

分 类 号：R714.5[医药卫生—妇产科学]