De novo variations of ANK1 gene caused hereditary spherocytosis in two Chinese children by affecting pre-mRNA splicing

作者全名："Wang, Yang; Huang, Lan; Zhu, Yao; An, Xizhou; Li, Jiacheng; Zhen, Jiangwei; Yu, Jie"

作者地址："[Wang, Yang; Huang, Lan; Zhu, Yao; An, Xizhou; Li, Jiacheng; Yu, Jie] Chongqing Med Univ, Childrens Hosp, Natl Clin Res Ctr Child Hlth & Disorders, Dept Hematol & Oncol,Minist Educ,Key Lab Child Dev, 136 Zhong Shan Er Lu, Chongqing 400014, Peoples R China; [Wang, Yang; Huang, Lan; Zhu, Yao; Li, Jiacheng; Yu, Jie] Chongqing Key Lab Pediat, Chongqing, Peoples R China; [Zhen, Jiangwei] Shenzhen Samii Int Med Ctr, Dept Endocrinol, Shenzhen 518000, Peoples R China"

通信作者："Yu, J (通讯作者)，Chongqing Med Univ, Childrens Hosp, Natl Clin Res Ctr Child Hlth & Disorders, Dept Hematol & Oncol,Minist Educ,Key Lab Child Dev, 136 Zhong Shan Er Lu, Chongqing 400014, Peoples R China.; Yu, J (通讯作者)，Chongqing Key Lab Pediat, Chongqing, Peoples R China."

来源：BMC PEDIATRICS

ESI学科分类：CLINICAL MEDICINE

WOS号：WOS:000913183600003

JCR分区：Q2

影响因子：2

年份：2023

卷号：23

期号：1

开始页：　

结束页：　

文献类型：Article

关键词：De novo variation; Hereditary spherocytosis; ANK1; Minigene splicing assay

摘要："Background and aimsHereditary spherocytosis (HS) is one of the most common hereditary haemolytic disorders. Here, two unrelated families with the probands displaying typical manifestations of HS were enrolled. Our study aimed to characterize the effect of two novel variants in HS patients on gene splicing to help minimize the rate of misdiagnosis of HS and enhance clinicians' understanding of the disease.Participants and methodsA retrospective review was conducted. Peripheral blood samples were collected from all the family members, and genomic DNA was extracted for genetic diagnostics. First, high-throughput sequencing technology was used for the preliminary screening of candidate causative variants. Thereafter, the variants were verified via Sanger sequencing. Furthermore, a pathogenicity analysis of the detected variants was performed including in silico prediction and in vitro experiments. We constructed matched wild-type and mutant-type minigene plasmid of ANK1 based on HEK293T cells to address the effects of variants on mRNA splicing.ResultsThe c.1305 + 2 T > A (family1) and c.1305 + 2del (family2) variants were detected in the ANK1 gene. These two de novo mutations described by us which have not been reported prior to this study. Moreover, the validation results of splicing reporter systems revealed that the intronic mutations resulted in abnormal pre-mRNA splicing. Specifically, the minigene plasmid expressing the c.1305 + 2 T > A variant transcribed the two aberrant transcripts: r.1305_1306ins1305 + 1_1305 + 229 and r.1305_1306ins1305 + 1_1305 + 552. The minigene plasmid expressing c.1305 + 2del transcribed the two aberrant transcripts: r.1305_1306ins1305 + 1_1305 + 228 and r.1305_1306ins1305 + 1_1305 + 551.ConclusionThe two de novo variants identified in the ANK1 gene were the genetic etiology of the probands with HS in our study. Our findings further enrich the HS genotype database and provide a basis for genetic counselling and molecular diagnosis."

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