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

甘氨酸脱羧酶基因（GLDC）突变引起非酮症高甘氨酸血症（NKH），这是一种以严重神经功能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，并在NKH的分子分析中研究了它们的表达。GLDC基因全长135 kb，由25个外显子组成。所有的供体和受体位点都遵守经典的GT-AG规则，除了内含子21的供体位点，其中使用变体形式GC代替GT。通过引物延伸分析，转录起始位点被分配到翻译起始三联体上游163 bp的残基。psiGLDC基因不含内含子，与功能性GLDC编码区的同源性为97.5%，表明psiGLDC是一个经过加工的假基因，起源于400 ~ 800万年前的GLDC转录本。RNA印迹分析显示GLDC在人的肝、肾、脑和胎盘中均有表达。然后，我们检查了一名NKH患者，他的淋巴母细胞中没有检测到GLDC mRNA。该患者的功能性GLDC基因的外显子1至3未被PCR扩增，而对照受试者的外显子被扩增。这些结果表明，在患者中存在大的纯合缺失（至少30 kb）。然后，我们设计了一个半定量PCR估计GLDC等位基因的数量使用psiGLDC作为内部控制，并确认了纯合性和杂合性的缺失，在病人和他的父母，分别。GLDC和psiGLDC的结构信息应有助于NKH的分子分析。

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.