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.