Chen Chong, Sun Qiao, Gu Mingmin, Liu Kun, Sun Yong, Xu Xun
Department of Ophthalmology, Shanghai First People's Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China.
Graefes Arch Clin Exp Ophthalmol. 2015 Jun;253(6):915-24. doi: 10.1007/s00417-015-3019-x. Epub 2015 May 7.
To unravel the molecular genetic background responsible for autosomal dominant congenital pulverulent nuclear cataracts in a four-generation Chinese family.
Family history data were collected, ophthalmological examinations were performed, and genomic DNA was extracted from peripheral blood of the family members. The candidate genes were captured and sequenced by targeted next-generation sequencing, and the results were confirmed by Sanger sequencing. The structure modelling of the protein was displayed based on Swiss-Model Server, and its possible changes in the secondary structure were predicted using Antheprot 2000 software. The chemical dissimilarity and possible functional impact of an amino acid substitution were performed with Grantham score, PolyPhen-2, and SIFT predictions. Protein distributions were assessed by confocal microscopy.
A novel heterozygous c.829C > T transition that led to the substitution of a highly conserved histidine by tyrosine at codon 277 (p.H277Y) in the coding region of connexin50 (Cx50, GJA8) was identified. Bioinformatics analysis showed that the mutation likely altered the secondary structure of the protein by replacing the helix of the COOH-terminal portion with a turn. The mutation was predicted to be moderately conservative by Grantham score and to be deleterious by both PolyPhen-2 and SIFT with consistent results. In addition, when expressed in COS1 cells, the mutation led to protein accumulation and caused changes in Cx 50 protein localization pattern.
This is a novel missense mutation [c.829C > T, (p.H277Y)] identified in exon 2 of Cx50. Our findings expand the spectrum of Cx50 mutations that are associated with autosomal dominant congenital pulverulent nuclear cataract.
为了阐明一个四代中国家系中常染色体显性遗传性先天性粉末状核性白内障的分子遗传背景。
收集家族病史数据，进行眼科检查，并从家族成员的外周血中提取基因组DNA。通过靶向二代测序捕获并测序候选基因，结果用桑格测序法进行确认。基于瑞士模型服务器展示蛋白质的结构模型，并使用Antheprot 2000软件预测其二级结构可能发生的变化。用格兰瑟姆评分、PolyPhen-2和SIFT预测对氨基酸替换的化学差异和可能的功能影响进行分析。通过共聚焦显微镜评估蛋白质分布。
在连接蛋白50（Cx50，GJA8）编码区发现一个新的杂合c.829C>T转换，该转换导致第277密码子处一个高度保守的组氨酸被酪氨酸取代（p.H277Y）。生物信息学分析表明，该突变可能通过用一个转角取代COOH末端部分的螺旋来改变蛋白质的二级结构。格兰瑟姆评分预测该突变具有中等保守性，PolyPhen-2和SIFT均预测该突变有害，结果一致。此外，当在COS1细胞中表达时，该突变导致蛋白质积累并引起Cx 50蛋白定位模式的改变。
这是在Cx50外显子2中鉴定出的一个新的错义突变[c.829C>T，（p.H277Y）]。我们的发现扩展了与常染色体显性遗传性先天性粉末状核性白内障相关的Cx50突变谱。
