Maroteaux-Lamy综合征的ARSB基因分析及新突变的致病性鉴定  

ARSB Gene Analysis and Novel Mutation Pathogenicity Identification of Maroteaux-Lamy Syndrome

作　　者：郭东炜 谢杰[1] 潘伟绵 唐佳 艾阳[1] 李荣[1] 杜敏联[4] 郭奕斌[1] GUO Dong-wei;XIE Jie;PAN Wei-mian;TANG Jia;AI Yang;LI Rong;DU Min-lian;GUO Yi-bin(MPS Team,Department of Medical Genetics,Zhongshan School of Medicine,Sun Yat-sen University,Guangzhou 510080;Clinical Medicine,Grade 2014,Medical College,Xiamen University,Xiamen 361102;Department of Immune Hematology,The Second Affiliated Hospital,Fujian University of Medical Science,Quanzhou 362000;Department of Pediatrics,The First Affiliated Hospital,SUN Yat-sen University,Guangzhou 510620,China)

机构地区：[1]中山大学中山医学院遗传室MPS科研组,广东广州510080 [2]厦门大学医学院,福建厦门361102 [3]福建医科大学附属二院免疫血液科,福建泉州362000 [4]中山大学附属第一医院儿科,广东广州510080

出　　处：《中山大学学报（医学版）》2018年第6期844-853,共10页Journal of Sun Yat-Sen University:Medical Sciences

基　　金：国家自然科学基金(30772069);闽粤合作科研基金(71010025和71020010)。

摘　　要：【目的】对7家拟诊为Maroteaux_Lamy综合征(MPSⅥ)的患儿及其父母进行ARSB基因的突变检测和新突变的致病性鉴定,以揭示其分子发病机制,为将来的产前/植入前基因诊断等创造前提条件。【方法】在临床初诊及GAG尿检和MPS酶检的基础上,抽取患儿及其父母EDTA抗凝血,进行ARSB基因的PCR扩增和Sanger测序。对所发现的新突变,在经HGMD、1000G和ExAC等数据库核实排查后,首先用SWISS-MODEL软件分析、比对突变蛋白和正常蛋白的空间构象,然后用Clustal X软件分析跨物种氨基酸的保守性,用PROVEAN、SIFT、PolyPhen-2软件预测新突变的致病性,最后用ACMG标准对新突变的致病性进行综合分析鉴定。【结果】1)7个家系先证者的基因检测结果分别为:No1:c.574T> C,p.C192R纯合错义突变;No2:c.160G> A/p.D54N(来自其母)和c.1197C> G/p.F399L(来自其父)的复合杂合子;No3:仅检出c.1072G> A/p.V358M和IVS5 as(-27)A>C变异,但酶检和临床表型符合Ⅵ型;No4:为c.281C> T,p.S94L(新突变,来自其母)和IVS5 as(-27)A> C(来自其父)的复合杂合子;No5:为c.1197 C> G,p.F399L纯合错义突变;No6:为c.1197 C> G,p.F399L(来自其母)和c.1379 C> T,p.S460F(新突变,来自其父)的复合杂合子;No7:为c.499 G> A,p.G167R(来自其父)和c.1325C>T,p.T442M(来自其母)的复合杂合子。2)新突变鉴定结果:对正常ARSB酶蛋白和p.S94L突变酶蛋白的空间构象的预测比对结果显示,两者有明显区别;跨物种保守性分析结果显示,p.94突变点所在氨基酸(S)在物种进化过程中具有高度保守性;PROVEAN、SIFT和PolyPhen-2软件对p.S94L预测结果分别为:Deleterious、Damaging和Probably damaging。用上述方法对p.S460F的预测结果及ACMG的综合分析结果也显示该突变可能是致病性的。【结论】1)家系4的p.S94L和家系6的p.S460F新突变可能都是新的致病性突变,有可能都是引起患儿发病的内在原因之一。2)家系1,2,5,6,7可以确诊为MPS Ⅵ型,其基因型和�【Objective】To reveal the molecular genetic mechanism of 7 cases that suspected Maroteaux_Lamy syndrome(MPSⅥ),we analyze the ARSB gene of proband and their parents and identify the pathogenicity of novel mutation,which lay foundation for prenatal diagnosis or preimplantation genetic diagnosis(PGD)in the future.【Methods】On the basis of clinical preliminary diagnosis,GAG urine test and the detection of MPS enzyme activity,and the EDTA anti-coagulated blood was collected from proband and their parents then perform PCR amplification and Sanger sequencing of the ARSB gene.After the novel mutations were detected and verified by databases such as HGMD,1000G and ExAC.At first,spatial conformation of mutant protein and normal protein were analyzed and compared by SWISSMODEL.Then conservation analysis of cross-species amino acids by Clustal X.What’s more,pathogenicity prediction methods including PROVEAN,SIFT and PolyPhen-2 were applied.Finally,the pathogenicity of the novel mutation was comprehensively analyzed and identified by ACMG standard.【Results】1)The genetic test results of the 7 family proband were as follows.Family 1,c.574T>C/p.C192R homozygous missense mutation.Family 2,c.160G>A/p.D54N(from mother)and c.1197C>G/p.F399L(from father)compound heterozygote.Family 3,except for c.1072G>A/p.V358M and IVS5 as(-27)A>C,no other mutations were found in the rest exons but the enzymatic and clinical phenotypes were in accordance with MPS type VI.Family 4,c.281C>T/p.S94L(novel mutation,from mother)and IVS5 as(-27)A>C(from father)compound heterozygote.Family 5,c.1197 C>G/.F399L homozygous missense mutation.Family 6,c.1197C>G/p.F399L(from mother)and c.1379C>T/p.S460F(novel mutation,from father)compound heterozygote.Family 7,c.499 G>A/p.G167R(from father)and c.1325C>T,p.T442M(from mother)compound heterozygote.2)Identification results of novel mutation.The spatial conformation predictions for normal ARSB enzyme protein and p.S94L mutant enzyme protein show that there is a clear difference between them,cross-species

关 键 词：Maroteaux_Lamy综合征 ARSB基因 新突变 致病性鉴定 

分 类 号：R394.3[医药卫生—医学遗传学]