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

BMC Pediatrics

Research

Jiangwei Zhen¹  Xizhou An²  Jie Yu³  Jiacheng Li³  Lan Huang³  Yao Zhu³  Yang Wang³ 

[1] Department of Endocrinology, Shenzhen Samii International Medical Center, 518000, Shenzhen, China;Department of Hematology and Oncology, Children’s Hospital of Chongqing Medical University, National Clinical Research Center for Child Health and Disorders, Ministry of Education Key Laboratory of Child Development and Disorders, Children’s Hospital of Chongqing Medical University, 136 Zhong shan er lu, Yu zhong district, 400014, Chongqing, China;Department of Hematology and Oncology, Children’s Hospital of Chongqing Medical University, National Clinical Research Center for Child Health and Disorders, Ministry of Education Key Laboratory of Child Development and Disorders, Children’s Hospital of Chongqing Medical University, 136 Zhong shan er lu, Yu zhong district, 400014, Chongqing, China;Chongqing Key Laboratory of Pediatrics, Chongqing, China;

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

DOI  :  10.1186/s12887-022-03795-0

 received in 2022-05-16, accepted in 2022-12-09,  发布年份 2022

来源: Springer

【 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.

【 授权许可】

CC BY   
© The Author(s) 2023

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