多巴反应性肌张力障碍三磷酸鸟苷环化水解酶Ⅰ基因突变检测

The mutations in GTP cyclohydrolase Ⅰ gene in Chinese patients with dopa responsive dystonia

目的 检测多巴反应性肌张力障碍 (dopa responsivedystonia,DRD)三磷酸鸟苷环化水解酶Ⅰ(guanosinetriphosphatecyclohydrolaseⅠ, GCH1)基因编码区的突变。方法 对 2个DRD家系的 5例患者和 6例散发患者及其 18名亲属和 20名健康对照者的GCH1基因编码区进行聚合酶链反应 单链构象多态性(PCR SSCP)分析;对PCR SSCP异常的外显子进行PCR产物直接测序,若患者 6个外显子PCR SSCP均无异常,则对所有外显子测序;为了确证突变,引入SphⅠ限制性内切酶位点进行聚合酶链反应 限制性片段长度多态性 (PCR RFLP)分析。结果 在 1个呈常染色体显性遗传DRD家系中发现 1个新的杂合型点突变(A224G)。此突变位于 1号外显子,由酪氨酸错义突变为半胱氨酸(Tyr75Cys), 20名健康对照者等位基因无此突变。另 1家系和其他散发患者在GCH1基因编码区未发现基因突变。序列分析提示与 2号外显子邻近的 1号内含子部分和与 3号外显子邻近的 3号内含子部分存在基因多态性。结论 我们描述了一个新的错义突变Tyr75Cys,GCH1基因编码区突变能解释部分DRD患者的发病原因。

Objective To detect the mutations in coding region of the guanosine triphosphate cyclohydrolase Ⅰ (GCH1) gene in Chinese patients with dopa-responsive dystonia (DRD).Methods Two families with five affected family members and six patients with sporadic DRD were examined, as well as their eighteen relatives and twenty normal members. Mutation screening was performed using single-strand conformation polymorphism analysis followed by direct sequencing of the presumably mutated exons; in patients whose results showed a normal pattern on single-strand conformation polymorphism analysis, the entire coding region of the GCH1 gene was sequenced. To confirm the mutation, a pair of primers was designed, which produced a restrictive site for SphI.Results DNA sequencing revealed a new heterozygous A224G missense mutation (Tyr75Cys) located within exon 1 in one family with autosomal-dominant inheritance. The mutation was confirmed with restriction enzyme analysis; it was not present in 20 control alleles. Restriction enzyme analysis also detects two systematic gene mutation carriers. In patients from the other family and patients with sporadic DRD, no alterations in the translated portion of the GCH1 gene were observed. Direct sequencing also showed that there existed gene polymorphisms in intron 1 and intron 3 which neared the exon2 and exon3 respectively in Chinese.Conclusions We describe a new missense mutation (Tyr75Cys) in the GCH1 gene. Mutation in the coding region of the gene might be accounted for a part of patients with DRD.