CHARACTERIZATION OF A SPLICING MUTATION IN GROUP-A XERODERMA-PIGMENTOSUM
CHARACTERIZATION OF A SPLICING MUTATION IN GROUP-A XERODERMA-PIGMENTOSUM
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DOI：
10.1073/pnas.87.24.9908
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发表时间：
1990-12-01
影响因子：
11.1
通讯作者：
OKADA, Y
中科院分区：
文献类型：
--
作者：
SATOKATA, I;TANAKA, K;OKADA, Y
通过比较来自 A 组 XP 患者的 XP A 组互补基因 (XPAC) 的多个克隆的核苷酸序列与正常基因的核苷酸序列，研究了 A 组着色性干皮病 (XP) 的分子基础。克隆显示G.fwdarw。内含子 3 的 3'' 剪接受体位点发生 C 取代，将必需的 AG 受体二核苷酸更改为 AC。对聚合酶链式反应扩增的 cDNA 进行核苷酸测序表明，这种单碱基取代消除了规范的 3'' 剪接位点，从而产生了两种异常剪接的 mRNA 形式。较大的形式与正常 mRNA 相同，只是在外显子 4 的 5'' 末端有一个二核苷酸缺失。这种缺失会导致外显子 4 中翻译提前终止的移码。较小的形式删除了整个外显子 3 和外显子 4 的 5'' 末端的二核苷酸。转染研究的结果提供了额外的证据，证明这种单碱基取代是致病突变。这种单碱基取代为限制性核酸酶 AlwNI 创建了一个新的切割位点。 AlwNI限制性片段长度多态性分析显示，该突变在 A XP 组日本患者中出现频率较高：21 名不相关的日本患者中，该突变有 16 名纯合子，4 名杂合子。然而，A XP 组的 11 名白人和 2 名黑人没有这种突变等位基因。结论是，多态性 AlwNI 限制性片段可用于诊断日本受试者（包括携带者的产前病例）A 组 XP。
The molecular basis of group A xeroderma pigmentosum (XP) was investigated by comparison of the nucleotide sequences of multiple clones of the XP group A complementing gene (XPAC) from a patient with group A XP with that of a normal gene. The clones showed a G .fwdarw. C substitution at the 3'' splice acceptor site of intron 3, which altered the obligatory AG acceptor dinucleotide to AC. Nucleotide sequencing of cDNAs amplified by the polymerase chain reaction revealed that this single base substitution abolishes the canonical 3'' splice site, thus creating two abnormally spliced mRNA forms. The larger form is identical with normal mRNA except for a dinucleotide deletion at the 5'' end of exon 4. This deletion results in a frameshift with premature translation termination in exon 4. The smaller form has a deletion of the entire exon 3 and the dinucleotide at the 5'' end of exon 4. The result of a transfection study provided additional evidence that this single base subsitution is the disease-causing mutation. This single base substitution creates a new cleavage site for the restriction nuclease AlwNI. Analysis of AlwNI restriction fragment length polymorphism showed a high frequency of this mutation in Japanese patients with group A XP: 16 of 21 unrelated Japanese patients were homozygous and 4 were heterozygous for this mutation. However, 11 Caucasians and 2 Blacks with group A XPdid not have this mutant allele. The polymorphic AlwNI restriction fragments are concluded to be useful for diagnosis of group A XP in Japanese subjects, including prenatal cases of carriers.