Diagnosis of xeroderma pigmentosum patients and carriers by PCR-RFLP analysis

最近克隆出了导致色素性干皮病（XP） A组的基因，并将其命名为XPA基因。既往研究表明，大多数日本XPA患者存在XPA基因内含子3剪接位点的纯合突变，该突变被限制性内切酶（RE） AlwNI （AlwNI突变）识别。数据中发现的其他突变有RE HphI识别的6外显子228密码子无义突变（HphI突变）和RE MseI识别的3外显子116密码子突变（MseI突变）。采用聚合酶链反应限制性片段长度多态性（PCR-PFLP）分析XPA基因的点突变。(1) 17例患者中12例为AlwNI突变纯合子，4例为AlwNI突变与HphI突变复合杂合子，1例为AlwNI突变与MseI突变复合杂合子。临床特征分析表明，4例HphI突变患者临床表现较轻。PCR-RFLP分析了3例无症状XPA患者的兄弟姐妹的XPA基因，发现其中2人携带突变的XPA等位基因，1人不携带。(2) A组两兄妹的临床表现有显著差异，妹妹的病情较轻。姐姐从4岁开始严格防晒，而妹妹从2岁开始。PCR-RFLP分析显示，两例患者具有相同的XPA基因突变。这些数据表明，在XP患者中，防晒开始得越早，皮肤癌发展得越晚。(3)我们诊断了一个胎儿的XPA基因突变，其兄弟姐妹是XPA患者。羊水DNA PCR-RFLP分析显示纯合突变。(4) P53蛋白在DNA损伤修复、细胞周期和细胞凋亡中起着至关重要的作用。western blot检测UVB对不同基因突变的XPA细胞中p53蛋白的诱导作用。UVB对HphI/AlwNI杂合突变细胞的p53蛋白诱导作用弱于AlwNI纯合突变细胞或AlwNI/MseI杂合突变细胞。少

The gene responsible for xeroderma pigmentosum (XP) group A has recently been cloned and designated XPA gene. Previous studies have shown that most Japanese XPA patients have homozygous mutations for the splicing site of intron 3 of the XPA gene, which was recognized by restriction endonuculease (RE) AlwNI (AlwNI mutation). Other mutations found to data have been the nonsense mutation at codon 228 in exon 6, recognized by RE HphI (HphI mutation), and at codon 116 in exon 3, recognized by RE MseI (MseI mutation) . Using polymerase chain reaction restriction fragment length polymorphism (PCR-PFLP) analysis, we examined the point mutations of the XPA gene.(1) We found that 12 patients of 17 patients were homozygous for the AlwNI mutation, four were compound heterozygotes for the AlwNI mutation and the HphI mutation, and one was a compound heterozygote for the AlwNI mutation and the MseI mutation. Investigation of their clinical features suggested that the four patients with HphI mutation … More had milder clinical manifestations. PCR-RFLP analysis of the XPA gene in the three asymptomatic siblings of the XPA patients revealed that two were carriers of the mutated XPA allele, and one was not a carrier.(2) We found two siblings with XP group A who showed a significant difference in clinical manifestations and younger sister was much milder. The elder sister started strict sun protection at 4 years of age, whereas the younger began at 2 years of age. PCR-RFLP analysis revealed that both patients had the identical mutation in XPA gane. These data suggent, that in patients with XP the earlier sun protection begins had the later skin cancer develops.(3) We diagnosed the XPA gene mutation of a fetus whose sibling is a XPA patient. PCR-RFLP analysis of DNA from amniotic fluid showed the homozygous mutation.(4) P53 protein is critical for DNA damage repair, cell cycle and apoptosis. The p53 protein induction by UVB in XPA cells with various gene mutations was examined using western blot analysis. Cells with HphI/AlwNI heterozygous mutations showed weaker induction of p53 protein by UVB than cells with AlwNI homozygous mutations or AlwNI/MseI heterozygous mutations. Less