复合杂合性F11基因突变导致的遗传性凝血因子Ⅺ缺陷症家系分析  

Identification of compound heterozygous mutations of F11 gene in a pedigree affected with heriditary coagulation factor Ⅺ deficiency

作　　者：刘媚娜[1] 李小龙[1] 周星星[1] 金艳慧[1] 杨丽红[1] 潘景业[1] 苏看看 王明山[1] Liu Meina;Li Xiaolong;Zhou Xingxing;Jin Yanhui;Yang Lihong;Pan Jingye;Su Kankan;Wang Mingshan(Center of Laboratory Medicine, the First Affiliated Hospital of Wenzhou Medical University, Wenzhou, Zhejiang 325015, China)

机构地区：[1]温州医科大学附属第一医院医学检验中心,浙江325015

出　　处：《中华医学遗传学杂志》2019年第4期363-367,共5页Chinese Journal of Medical Genetics

基　　金：浙江省科技厅公益技术研究计划基金(LGF18H080003).

摘　　要：目的对一个遗传性凝血因子Ⅺ(coagulation factor Ⅺ,FⅪ)缺陷症患者家系进行表型和基因检测,寻找致病基因并初步探讨其分子致病机制。方法检测先证者及其家系成员的血浆凝血酶原时间(prothrombin time,PT)、活化部分凝血活酶时间(activated partial thromboplastin time,APTT)、纤维蛋白原(fibrinogen,FIB)含量、血浆凝血因子Ⅷ活性(coagulation factor Ⅷ activity,FⅧ∶C)、凝血因子Ⅸ活性(coagulation factor Ⅸ activity,FⅨ∶C )、FⅪ活性(FⅪ activity,FⅪ∶C)、凝血因子Ⅻ活性(coagulation factor Ⅻ activity,FⅫ∶C)和狼疮抗凝物(lupus antieoagulation,LA);ELISA法检测FⅪ抗原(FⅪ antigen,FⅪ∶Ag)等指标,明确临床表型诊断。采用DNA直接测序法分析先证者F11基因15个外显子、侧翼序列以及5′端、3′端非翻译区序列,发现突变位点后用反向测序予以证实,并检测家系成员相应的突变位点区域。采用ClustalX-2.1-win软件分析氨基酸突变位点的保守性;用PolyPhen-2 、PROVEAN、SIFT和Mutation Taster 4个在线生物信息学软件分析突变对蛋白质功能的影响;用Swiss-PdbViewer软件对突变位点进行蛋白模型和氨基酸相互作用分析。结果先证者APTT为69.6 s,明显延长,其FⅪ∶C和FⅪ∶Ag均明显下降,分别为6%和10.7%;其母亲、姐姐、大妹、弟弟、女儿和儿子APTT均稍延长,FⅪ∶C和FⅪ∶Ag也均有不同程度下降(为正常对照的50%左右)。基因分析发现先证者F11基因第7外显子的c.738G>A(p.Trp228stop)杂合无义突变及第13外显子的c.1556G>C(p.Trp501Ser)杂合错义突变;其母亲、姐姐和女儿存在p.Trp228stop突变杂合子,大妹、弟弟和儿子存在p.Trp501Ser突变杂合子;其丈夫和小妹均为野生型。保守性分析结果表明,Trp501在同源物种间高度保守。4个生物信息学软件对该突变的预测结果一致:PolyPhen-2评分结果为1.000分,PROVEAN评分结果为-11.565分,均预示此突变是可能致病性的突变;SIFObjective To identify potential mutations of F11 gene in a pedigree affected with hereditary coagulation factor Ⅺ(FⅪ) deficiency and explore its molecular pathogenesis. Methods Prothrombin time (PT), activated partial thromboplastin time (APTT), fibrinogen (FIB), coagulation factor Ⅷ activity (FⅧ∶C), coagulation factor Ⅸ activity (FⅨ∶C), coagulation factorⅪ activity (FⅪ∶C), coagulation factor Ⅻ activity (FⅫ∶C) and lupus antieoagulation (LA) of the proband and eight family members were determined. FⅪ antigen (FⅪ∶Ag) was determined by enzyme-linked immunosorbent assay (ELISA). For the proband, potential mutations in the exons, flanking introns and 5′-, 3′-untranslated regions of the F11 gene were screened by direct DNA sequencing. The results were confirmed by reverse sequencing. Suspected mutations were detected in other family members. ClustalX-2.1-win and four online bioinformatics tools (PolyPhen-2, PROVEAN, SIFT, and Mutation Taster) were used to study the conservation and possible impact of the mutations. The structure of the mutational sites was processed with Swiss-PdbViewer. Results The propositus had prolonged APTT (69.6 s), whose FⅪ∶C and FⅪ∶Ag were reduced to 6.0% and 10.7%, respectively. Her mother, elder sister, one younger sister, little brother, daughter and son showed slightly prolonged APTT and median FXI∶C and FⅪ∶Ag level. Gene sequencing revealed that the propositus carried a heterozygous nonsense mutation c. 738G>A (p.Trp228stop) in exon 7 and a heterozygous mutation c. 1556G>C (p.Trp501Ser) in exon 13. Her mother, elder sister and daughter were heterozygous for the p. Trp228stop mutation, while one younger sister and little brother and son were heterozygous for p. Trp501Ser. Her husband and the youngest sister were of the wild type. Phylegenetics analysis suggested that Trp501 was highly conserved among all homologous species. The p. Trp501Ser was predicted to be "probably damaging","deleterious","affect protein function" and "disease causing

关 键 词：凝血因子Ⅺ缺陷症 遗传性 生物信息学 模型分析 

分 类 号：R554.7[医药卫生—血液循环系统疾病]