Liu Xing-yuan, Yang Yi-qing, Yang Ying, Lin Xiao-ping, Chen Yi-han
Department of Paediatrics, Tongji University Affiliated Tongji Hospital, Shanghai 200065, China.
Zhonghua Er Ke Za Zhi. 2009 Sep;47(9):696-700.
The purpose of this investigation was to identify the novel genetic mutations in patients with a congenital atrial septal defect (ASD).
Clinical data and blood specimens from a total of 12 unrelated ASD pedigrees and a cohort of 168 unrelated subjects with sporadic ASD were collected and evaluated in contrast to 200 healthy individuals. The whole exons and partial flanking introns of NKX2-5 gene were amplified by polymerase chain reaction and sequenced using the di-deoxynucleotide chain termination approach. The acquired sequences were aligned with those publicized in GenBank by the aid of programme BLAST to identify the sequence variations. The software ClustalW was applied for analysis of the conservative of the altered amino acids.
A novel heterozygous mutation of NKX2-5 gene, i.e., a substitution of thymine for cytosine at nucleotide 536, predicting the conversion of serine into phenylalanine at amino acid residue 179, was identified initially in a proband. The same missense mutation was thereafter detected in other 3 affected members of the identical family but neither in the healthy members of the kindred nor in the 200 normal controls. A cross-species alignment of the protein sequences encoded by NKX2-5 gene displayed that the mutated amino acid was highly conserved evolutionarily. No mutation of NKX2-5 gene was observed in the other 11 ASD pedigrees or in 168 patients with sporadic ASD. Additionally, a common synonymous single nucleotide polymorphism, a transition of adenine (A) into guanine (G) at nucleotide 63, was found in NKX2-5 gene. However, there were no significant differences in the prevalence of alleles A and G between ASD patients and healthy controls (chi(2) = 2.8641, P = 0.0906).
A novel mutation of NKX2-5 gene identified in an ASD family suggests that mutated NKX2-5 gene is responsible for familial ASD.
本研究旨在鉴定先天性房间隔缺损（ASD）患者中的新型基因突变。
收集了来自12个无亲缘关系的ASD家系的临床资料和血液标本，以及168名散发ASD的无亲缘关系个体，并与200名健康个体进行对比评估。采用聚合酶链反应扩增NKX2-5基因的全部外显子和部分侧翼内含子，并使用双脱氧核苷酸链终止法进行测序。借助BLAST程序将获得的序列与GenBank中公布的序列进行比对，以鉴定序列变异。应用ClustalW软件分析氨基酸改变后的保守性。
在一名先证者中首次鉴定出NKX2-5基因的一种新型杂合突变，即核苷酸536处胸腺嘧啶取代胞嘧啶，预测氨基酸残基179处丝氨酸转变为苯丙氨酸。随后在同一家族的其他3名患病成员中检测到相同的错义突变，但在该家族的健康成员以及200名正常对照中均未检测到。NKX2-5基因编码的蛋白质序列的跨物种比对显示，突变的氨基酸在进化上高度保守。在其他11个ASD家系或168例散发ASD患者中未观察到NKX2-5基因的突变。此外，在NKX2-5基因中发现了一种常见的同义单核苷酸多态性，即核苷酸63处腺嘌呤（A）向鸟嘌呤（G）的转变。然而，ASD患者和健康对照之间等位基因A和G的患病率无显著差异（χ² = 2.8641，P = 0.0906）。
在一个ASD家族中鉴定出的NKX2-5基因新型突变表明，突变的NKX2-5基因是家族性ASD的病因。
