由ALK1基因内含子突变引起的遗传性出血性毛细血管扩张症

Hereditary hemorrhagic telangiectasia caused by mutation in intron 4 of ALK1 gene:analysis of a HTT family

目的探讨遗传性出血性毛细血管扩张症发病的分子机制。方法 (1)用聚合酶链式反应-单链构像多态性分析(PCR-SSCP)寻找异常突变的外显子及其相邻剪接位点。(2)通过 DNA 测序确定突变的类型。(3)使用逆转录-聚合酶链反应(RT-PCR)方法确定剪接方式。结果用 PCR-SSCP 方法发现扩增 ALK1基因第4外显子及相邻的部分内含子片段有突变位点。对有突变该片段,用 DNA 测序检测,发现第4外显子的给位剪接点相邻碱基 A>T(IVS4+3 a>t)的突变,并用反向测序确认。用 RT-PCR 方法检测 ALK1基因表达 mRNA 情况,电泳和测序发现第4外显子的丢失。结论 ALK1基因的ⅣS4+3 a>t 突变,导致 ALK1基因表达异常,形成无功能截短蛋白,引起 HHT2。

Objective To investigate the molecular pathogenesis of hereditary hemorrhagic telangiectasia（HHT）Methods Peripheral blood samples were collected from a HTT family,including the proband,female,aged 48,and her mother ,elder brother,elder sister,younger brother,and son HHT gene mutations were identified by PCR-SSCP and DNA sequencing and confirmed by reverse sequencing.Ectopic transcripts of RT-PCR were used to confiom the characteristics of the mutation in non-canonical splicing site（ⅣS4＋3a〉t）Results A mutational segment of PCR producl of exon 4,exon-intron bonndaries and the 3＇5＇untranslated sequence of ALK1 gene was identified by PCR-SSCP,The mutational segment was analyzed by DNA sequencing,An ⅣS4＋3a〉t mutation was found,causing splicing abnormality of intron 4 and exon 3 skipping,Conclusion A splicing pattern of the Ⅳ S4＋3a〉t mutation has been reported among Chinese HHT2 patients for the first time.