Recurrent deletion in glycine decarboxylase gene and nonketotic hyperglycinemia Medical genetics, Research

甘氨酸脱羧酶基因反复缺失与非酮症高甘氨酸血症医学遗传学、研究

• 批准号: 10672134
• 负责人: KURE Shigeo
• 金额: $ 1.98万
• 依托单位: TOHOKU UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1998
• 资助国家: 日本
• 起止时间: 1998 至 1999
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10672134/
• 关键词: glycine decarboxylase; Alu repeats; non-ketotic hyperglycinemia; deletion; homologous recombination; glycine decarboxylase; Nonketotic hyperglycinemia; Homologous recombination; ghycime decarboxylase; nonkefotic hyperglycimemia; homologous recom

Mutations in glycine decarboxylase gene (GLDC) cause nonketotic hyperglycinemia (NKH), an inborn error of metabolism characterized by severe neurological disturbance. We determined the structures of GLDC and its pseudogene (psiGLDC) and studied their expression in molecular analysis of NKH. The GLDC gene spanned a least 135 kb and consisted of 25 exons. All donor and acceptor sites adhered to the canonical GT-AG rule except for the donor site of intron 21, where a variant form GC was used instead of GT. The transcription initiation site was assigned to a residue 163 bp upstream from the translation initiation triplet by primer extension analysis. The psiGLDC gene had no intron and shared 97.5% homology with the coding region of functional GLDC, suggesting that psiGLDC ia a processed pseudogene that arose from the GLDC transcript about 4 to 8 million years ago. RNA blotting analysis revealed that GLDC is expressed in human liver, kidney, brain and placenta. We then examined a patient with NKH with no detectable GLDC mRNA in his lymphoblasts. Exons 1 to 3 of the functional GLDC gene from this patient were not amplified by PCR, whereas those from control subjects were amplified. These results suggest a large homozygous deletion (at least 30 kb) in the patient. We then devise a semi-quantitative PCR to estimate the number of GLDC alleles using psiGLDC as an internal control, and confirmed the homozygosity and heterozygosity of the deletion in the patient and his parents, respectively. Structural information of GLDC and psiGLDC should facilitate the molecular analysis of NKH.

甘氨酸脱羧酶基因 (GLDC) 突变会导致非酮症高甘氨酸血症 (NKH)，这是一种以严重神经障碍为特征的先天性代谢缺陷。我们确定了 GLDC 及其假基因 (psiGLDC) 的结构，并研究了它们在 NKH 分子分析中的表达。 GLDC 基因跨度至少 135 kb，由 25 个外显子组成。所有供体和受体位点均遵守规范的 GT-AG 规则，但内含子 21 的供体位点除外，其中使用变体形式 GC 代替 GT。通过引物延伸分析，转录起始位点被指定为翻译起始三联体上游 163 bp 的残基。 psiGLDC基因没有内含子，与功能性GLDC的编码区有97.5%的同源性，这表明psiGLDC是一个经过加工的假基因，产生于大约4至8百万年前的GLDC转录本。 RNA印迹分析显示GLDC在人类肝脏、肾脏、大脑和胎盘中表达。然后我们检查了一名 NKH 患者，其淋巴母细胞中未检测到 GLDC mRNA。 PCR 未扩增该患者功能性 GLDC 基因的外显子 1 至 3，而对照受试者的外显子则被扩增。这些结果表明患者体内存在大量纯合缺失（至少 30 kb）。然后，我们设计了半定量PCR，以psiGLDC作为内部对照来估计GLDC等位基因的数量，并分别确认了患者及其父母中缺失的纯合性和杂合性。 GLDC 和 psiGLDC 的结构信息应有助于 NKH 的分子分析。

项目成果

Kure S,et al.: "Prenatal diagnosis of nonketotic hyperglycinemia・・"Prenat. Diagn.. 19. 717-720 (1999)

Kure S 等人：“非酮症高甘氨酸血症的产前诊断……”Diagn.. 19. 717-720 (1999)

Kure S,et al.: "Tetrahydrobiopterin-responsive phenylalanine hydroxylase・・"J. Pediatr. 135. 375-378 (1999)

Kure S 等人：“四氢生物蝶呤响应性苯丙氨酸羟化酶……”J. Pediatr. 135. 375-378 (1999)

Kure S,at.al.: "A subtype of pyridoxime dependent epilepsy with・・・" J Inher Metabol Dis. 21. 431-432 (1998)

Kure S,at.al.：“吡哆肟依赖性癫痫亚型......”J Inher Metabol Dis. 21. 431-432 (1998)

Kure S.et al.: "Tetrahydrobiopterine-responsive phenylalanine hydrosylase deficiency"Journal of Pediatrics. 135. 357-378 (1999)

Kure S.等人：“四氢生物蝶呤反应性苯丙氨酸水解酶缺乏症”儿科杂志。

Fujii K, Matsubara Y, Akanuma J, Takahashi K, Kure S, Suzuki Y, Imaizumi M, Iinuma K, Sakatsme O, Rinaldo P, Kuniaki Narisawa.: "Mutation detection by TaqMan-allele specific amplification: its application to the molecular diagnosis of glycogen storage dis

Fujii K、Matsubara Y、Akanuma J、Takahashi K、Kure S、Suzuki Y、Imaizumi M、Iinuma K、Sakatsme O、Rinaldo P、Kuniaki Narisawa。：“TaqMan 等位基因特异性扩增的突变检测：其在分子诊断中的应用