薄基底膜肾病家系新发COL4A4基因突变及致病机制探究  

Identification of a novel COIAA4 mutation in a family with thin- basement- membrane nephropathy and the pathogenic mechanism

作　　者：张国庆[1,2] 林楠[1] 国敏[1] 徐岩[1] 

机构地区：[1]青岛大学附属医院肾内科,青岛266000 [2]蓬莱市人民医院重症医学科

出　　处：《中华肾脏病杂志》2016年第2期94-98,共5页Chinese Journal of Nephrology

基　　金：国家自然科学基金面上项目（81170688、81470973）;山东省自然科学基金（ZR2011HM053）

摘　　要：目的对显性遗传的薄基底膜肾病（thin - basement - membrane nephropathy，TBMN）家系进行COL4A3和COL4A4基因分析，寻找突变位点并阐述其致病机制。方法提取先证者外周血基因组DNA，PCR法扩增COL4A3和COL4A4基因所有外显子，与GenBank数据库中的标准序列比对寻找突变位点，确定COL4A4基因突变后对其他6例家系成员和200例健康对照者进行该位点测序。提取先证者外周血RNA，反转录后PCR扩增含突变位点的基因片段进行TA克隆，挑选阳性单克隆菌落进行测序分析RNA。结果DNA测序显示先证者及其母亲、大姐均发现COL4A4基因1459位点G杂合突变为A，其他4例家庭成员和200例健康对照均未见该位点突变。先证者RNA分析显示COLAA4（c．1459＋G〉A）突变基因位于第21号与22号外显子之间的剪接位点，该位点突变导致剪接错误，其mRNA序列第21号外显子缺失，相应编码的氨基酸序列也全部缺失，20号外显子后编码的氨基酸序列也随之发生移码突变。结论本研究报道的COL4A4（c．1459＋G〉A）为TBMN患者的致病基因突变，其致病机制进一步丰富了TBMN发病机制，为将来临床诊断提供参考。Objective To explore a new pathogenic gene mutationin in COIAA3 and COL4A4 genes of a family with thin- basement- membrane nephropathy （TBMN）, and explain its mechanism. Methods Genomic DNA was extracted from blood samples. Mutation screening for all the exons in COL4A3 and COL4A4 of the proband was carried out by direct PCR sequencing. The sequences of the proband were compared with standard sequences in GenBank. After identifying the mutation in COL4A4, screening for the mutation site in 200 healthy controls and the rest of family members were conducted. RNA sequence of the proband was analyzed by reverse transcription PCR and TA cloning. The positive clones were sequenced for RNA screening. Results There was a G to A mutation in the 1459 site of COLAA4 （c.1459＋G 〉 A） in the proband, her motber, and the elder sister, whereas the mutation was not found in other family members and healthy people. RNA screening showed that the COIAA4 （c.1459＋G 〉 A） mutation was a heterozygous substitution in position ＋1 of exon 21, in the splicing region. This mutation leaded to eliminating of exon 21 from the COL4A4 mRNA, causing the exon 21 deletion and frameshift mutation following the exon 20 in its amino acids sequence. Conclusions It is described that COIAA4 （c.1459＋G 〉 A） is a new pathogenic mutationin TBMN, which further help understanding the pathogenesis and clinical diagnosis of TBMN.

关 键 词：血尿 胶原Ⅳ型 突变 薄基底膜肾病 

分 类 号：R692[医药卫生—泌尿科学]