Identification of two novel mutations in the methylmalonyl-CoA mutase gene with decreased levels of mutant mRNA in methylmalonic acidemia.
Identification of two novel mutations in the methylmalonyl-CoA mutase gene with decreased levels of mutant mRNA in methylmalonic acidemia.
复制标题
甲基丙二酰辅酶 A 变位酶基因中两个新突变的鉴定，在甲基丙二酸血症中突变 mRNA 水平降低。
DOI：
10.1093/hmg/3.6.867
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发表时间：
1994
影响因子：
3.5
通讯作者：
K. Narisawa
中科院分区：
文献类型：
--
作者：
M. Ogasawara;Y. Matsubara;H. Mikami;K. Narisawa
甲基丙二酸单酰辅酶A β（MCM）基因的遗传缺陷导致甲基丙二酸血症，这是一种常染色体隐性遗传疾病。我们调查了从两个日本患者与MCM缺陷的成纤维细胞培养。北方印迹分析未检测到MCM mRNA，表明MCM mRNA显著降低。逆转录/聚合酶链反应（RT-PCR）的MCM mRNA的荧光片段分析仪上的分析表明，MCM mRNA的水平在这些成纤维细胞是不到1%的正常对照。通过巢式RT-PCR成功地扩增了这微量的MCM mRNA，并进行了一级结构分析。序列分析发现了两个新的突变：在核苷酸位置425处的G到T取代和在核苷酸位置769和770处的2bp缺失。第一个突变（G425 T）导致氨基酸位置117处的终止密码子被谷氨酸取代。第二个突变（769 Δ CA）导致移码，在蛋白质C-末端上游产生提前终止密码子508个氨基酸。患者1为G425 T纯合子，患者2为G425 T和769 Δ CA的复合杂合子。我们的报告是第一个确定MCM突变影响MCM mRNA的稳定性。对16名日本患者的分析显示，6名患者存在G425 T，表明日本患者中该突变的发生率相对较高。这与先前的报道形成鲜明对比，该报道描述了高加索人中MCM突变的多样性。
Genetic defects in the methylmalonyl-CoA mutase (MCM) gene result in methylmalonic acidemia which is inherited as an autosomal recessive disease. We investigated fibroblast cultures obtained from two Japanese patients with MCM deficiency. MCM mRNA was not detected by Northern blot analysis, suggesting that MCM mRNA was markedly decreased. Reverse transcription/polymerase chain reaction (RT-PCR) of MCM mRNA followed by analysis on a fluorescent fragment analyzer indicated that the level of MCM mRNA in these fibroblasts was less than 1% of normal controls. This minute amount of MCM mRNA was successfully amplified by nested RT-PCR and subjected to primary structure analysis. Sequence analysis revealed two novel mutations: a G-to-T substitution at nucleotide position 425 and a 2 bp deletion at nucleotide positions 769 and 770. The first mutation (G425T) resulted in the substitution of a termination codon for glutamic acid at amino acid position 117. The second mutation (769 delta CA) resulted in a frame shift which created a premature termination codon 508 amino acid upstream of the C-terminus of the protein. Patient 1 was homozygous for G425T and patient 2 was a compound heterozygote for G425T and 769 delta CA. Our report is the first to identify MCM mutations that affect the stability of MCM mRNA. An analysis of 16 Japanese patients revealed the presence of G425T in six patients, suggesting a relatively high incidence of the mutation among Japanese patients. This is in sharp contrast to a previous report describing diverse heterogeneity of MCM mutations among Caucasians.