Functional analysis of a new splicing mutation in the MYBPC3 gene in hypertrophic cardiomyopathy

R. R. Salakhov, M. Golubenko, M. Skoblov, R. R. Savchenko, N. Valiakhmetov, E. N. Pavlyukova, M. S. Nazarenko
{"title":"Functional analysis of a new splicing mutation in the MYBPC3 gene in hypertrophic cardiomyopathy","authors":"R. R. Salakhov, M. Golubenko, M. Skoblov, R. R. Savchenko, N. Valiakhmetov, E. N. Pavlyukova, M. S. Nazarenko","doi":"10.20538/1682-0363-2024-2-183-189","DOIUrl":null,"url":null,"abstract":"Aim. To study the pathogenic effect in the MYBPC3 splice-site variant in the patient with hypertrophic cardiomyopathy. Materials and methods. The study was conducted using a DNA sample obtained from a patient with hypertrophic cardiomyopathy, in whom a previously undescribed variant was identified in the splice donor site of intron 21. The methods used included constructing and cloning of minigenes (vector pSpl3-Flu2-TKdel) and transfection of a human cell culture (HEK293T), followed by isolation of mRNA, production of cDNA, PCR of the minigene region containing the analyzed fragment, agarose gel electrophoresis, and Sanger sequencing. Results. The chr11:47339649-A-C (hg38) variant, disrupting the splice donor site in intron 21 (NM_000256.3: c.2067+2T>G), was identified in the 23-year-old patient with obstructive hypertrophic cardiomyopathy. To directly analyze the effect of this variant on splicing, a vector containing exon 21, intron 21, exon 22, and partially introns 20 and 22 of the MYBPC3 gene was obtained. A comparison of mRNAs from the minigenes containing / not containing the variant showed that the chr11:47339649-A-C substitution led to exon 21 and exon 22 skipping during splicing. Conclusion. The study established the functional significance of the previously undescribed variant c.2067+2T>G in the MYBPC3 gene, resulting in disruption of the mRNA splicing mechanism in the patient with hypertrophic cardiomyopathy. This variant can be classified as pathogenic.","PeriodicalId":256912,"journal":{"name":"Bulletin of Siberian Medicine","volume":"142 32","pages":""},"PeriodicalIF":0.0000,"publicationDate":"2024-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Bulletin of Siberian Medicine","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.20538/1682-0363-2024-2-183-189","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
引用次数: 0

Abstract

Aim. To study the pathogenic effect in the MYBPC3 splice-site variant in the patient with hypertrophic cardiomyopathy. Materials and methods. The study was conducted using a DNA sample obtained from a patient with hypertrophic cardiomyopathy, in whom a previously undescribed variant was identified in the splice donor site of intron 21. The methods used included constructing and cloning of minigenes (vector pSpl3-Flu2-TKdel) and transfection of a human cell culture (HEK293T), followed by isolation of mRNA, production of cDNA, PCR of the minigene region containing the analyzed fragment, agarose gel electrophoresis, and Sanger sequencing. Results. The chr11:47339649-A-C (hg38) variant, disrupting the splice donor site in intron 21 (NM_000256.3: c.2067+2T>G), was identified in the 23-year-old patient with obstructive hypertrophic cardiomyopathy. To directly analyze the effect of this variant on splicing, a vector containing exon 21, intron 21, exon 22, and partially introns 20 and 22 of the MYBPC3 gene was obtained. A comparison of mRNAs from the minigenes containing / not containing the variant showed that the chr11:47339649-A-C substitution led to exon 21 and exon 22 skipping during splicing. Conclusion. The study established the functional significance of the previously undescribed variant c.2067+2T>G in the MYBPC3 gene, resulting in disruption of the mRNA splicing mechanism in the patient with hypertrophic cardiomyopathy. This variant can be classified as pathogenic.
肥厚型心肌病 MYBPC3 基因新剪接突变的功能分析
研究目的研究MYBPC3剪接位点变异对肥厚型心肌病患者的致病作用。材料和方法。研究使用了从一名肥厚型心肌病患者身上获得的 DNA 样本,在该患者的内含子 21 的剪接供体位点上发现了一个以前未曾描述过的变异。所用方法包括构建和克隆迷你基因(载体 pSpl3-Flu2-TKdel),转染人细胞培养物(HEK293T),然后分离 mRNA、制作 cDNA、对包含分析片段的迷你基因区域进行 PCR、琼脂糖凝胶电泳和 Sanger 测序。结果。在一名 23 岁的阻塞性肥厚型心肌病患者体内发现了 chr11:47339649-A-C (hg38) 变体,该变体破坏了内含子 21 的剪接供体位点(NM_000256.3: c.2067+2T>G)。为了直接分析该变异对剪接的影响,研究人员获得了包含 MYBPC3 基因第 21 号外显子、第 21 号内含子、第 22 号外显子以及部分第 20 号和第 22 号内含子的载体。对含有/不含有该变异的迷你基因的 mRNA 进行比较后发现,chr11:47339649-A-C 的置换导致剪接过程中 21 号外显子和 22 号外显子的跳过。结论该研究确定了 MYBPC3 基因中以前未曾描述过的 c.2067+2T>G 变异的功能意义,它导致肥厚型心肌病患者的 mRNA 剪接机制紊乱。该变异可归类为致病性。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
求助全文
约1分钟内获得全文 求助全文
来源期刊
自引率
0.00%
发文量
0
×
引用
GB/T 7714-2015
复制
MLA
复制
APA
复制
导出至
BibTeX EndNote RefMan NoteFirst NoteExpress
×
提示
您的信息不完整,为了账户安全,请先补充。
现在去补充
×
提示
您因"违规操作"
具体请查看互助需知
我知道了
×
提示
确定
请完成安全验证×
copy
已复制链接
快去分享给好友吧!
我知道了
右上角分享
点击右上角分享
0
联系我们:info@booksci.cn Book学术提供免费学术资源搜索服务,方便国内外学者检索中英文文献。致力于提供最便捷和优质的服务体验。 Copyright © 2023 布克学术 All rights reserved.
京ICP备2023020795号-1
ghs 京公网安备 11010802042870号
Book学术文献互助
Book学术文献互助群
群 号:481959085
Book学术官方微信