表皮松解性掌跖角化病一家系的致病基因突变分析  

Analysis for mutation of pathogenic gene in a pedigree with epidermolytic palmoplantar keratoderma

作　　者：葛海军 刘旭[2] 陈晨[1] 王述森[1] 王谦[1] 曹丽华[1] 罗阳[1] 张学[1] GE Hai-jun;LIU Xu;CHEN Chen;WANG Shu-sen;WANG Qian;CAO Li-hua;LUO Yang;ZHANG Xue(Department of Medical Genomics,College of Life Sciences,China Medical University,Key Laboratory of Medical Cell Biology,Ministry of Education,Key Laboratory of Cell Biology,Ministry of Health,Shenyang 110122;Department of Neurology,First Affiliated Hospital of China Medical University,Shenyang 110001,China)

机构地区：[1]中国医科大学生命科学学院医学基因组学教研室教育部医学细胞生物学重点实验室卫生部细胞生物学重点实验室,辽宁沈阳110122 [2]中国医科大学附属第一医院神经内科,辽宁沈阳110001

出　　处：《解剖科学进展》2020年第5期492-495,499,共5页Progress of Anatomical Sciences

基　　金：国家自然科学基金青年基金(81502176)。

摘　　要：目的对一个表皮松解性掌跖角化症(EPPK)家系进行基因突变检测,为该家系成员提供临床遗传咨询,并初步探讨该家系患者特殊表型指节垫、指关节弯曲与KRT9基因突变的相关性。方法收集该家系中的先证者、先证者的父亲及舅舅三人的外周血并提取基因组DNA。对以上三位家系成员进行全外显子组测序(WES),通过突变位点频率、突变类型、是否改变剪接或编码氨基酸进行筛选,并按照常染色体显性遗传方式筛选杂合突变,然后在家系内进行共分离分析,针对候选基因表达及已知功能分析,得到候选基因突变位点,并进行Sanger测序验证。应用生物信息学软件对KRT9基因结构域和功能域进行分析,针对161位氨基酸位点改变,进行文献综述和基因型-表型相关性探讨。结果本研究报道了一个中国EPPK家系,伴有罕见的指关节屈曲表型;WES测序检测到KRT9基因第1外显子存在杂合突变c.482A>G(p.N161S),该突变在家系内与疾病表型共分离,Sanger测序验证与WES结果一致。利用生物信息学软件预测到N161突变成S161后,角蛋白9的无规则卷曲发生空间位置改变,α螺旋结构有明显缺少,突变位点附近氢键数量增加,键长变长。对角蛋白9的161位点突变所致EPPK进行文献综述,发现N161S突变与指节垫表型高度相关。结论本研究报道了国际上第三例伴有罕见指关节屈曲表型的EPPK家系,初步认为KRT9基因c.482A>G(p.N161S)为该EPPK家系致病突变,对患者明确诊断及遗传咨询有重要意义。Objective To detect gene mutations in a pedigree with epidermolytic palmoplantar keratoderma(EPPK),provide clinical genetic counseling for members of the family and explore the relationship between KRT9 mutations and the special phenotype of knuckle pads and camptodactyly.Methods The peripheral blood of the proband,his father and uncle was collected and genomic DNA was extracted.Whole Exome Sequencing(WES)was performed on the above three family members,the variants were filtered by frequency,type of mutations,alteration of splicing or coding amino acids,and heterozygous mutations were selected according to autosomal dominant inheritance.The candidate mutations were obtained by co-separation analysis within families,and gene expression and known functions were analyzed.Candidate mutation sites were amplified by PCR and verified by Sanger sequencing.Analysis of KRT9 gene domains and functional domains was performed using bioinformatics softwares,and genotype-phenotype correlations of 161 amino acid were reviewed.Results Chinese EPPK family with a rare phenotype of camptodactyly was reported firstly,and a heterozygous mutation c.482 A>G(p.N161 S)was detected in exon 1 of KRT9 gene by WES sequencing.The mutation was co-separated with the phenotypes in the family.The results of Sanger sequencing were consistent with WES results.Bioinformatics software predicted that the random coils of keratin 9 altered in spatial position in the S161 mutant with decreased alpha helixes.In addition,the number of hydrogen bonds surrounding the mutation site increased and the bond length became longer,N161 S mutation is highly correlated to Knuckle pads.Conclusion The third EPPK family with phenotype of camptodactyly was reported in the world,and c.482 A>G(p.N161 S)of KRT9 gene is the pathogenic mutation of this EPPK family,with a great significance for diagnosis and genetic counseling of patients.

关 键 词：EPPK KRT9基因 突变 指节垫 

分 类 号：R394.1[医药卫生—医学遗传学]