Fukuhara Yasuyuki, Miura Ai, Yamazaki Narutoshi, So Tetsumin, Kosuga Motomichi, Yanagi Kumiko, Kaname Tadashi, Yamagata Takanori, Sakuraba Hitoshi, Okuyama Torayuki
Division of Medical Genetics, National Center for Child Health and Development, 2-10-1 Okura, Setagaya-ku, Tokyo 157-8535, Japan.
Department of Clinical Laboratory Medicine, National Center for Child Health and Development, 2-10-1 Okura, Setagaya-ku, Tokyo, 157-8535, Japan.
Mol Genet Metab Rep. 2020 Dec 10;25:100692. doi: 10.1016/j.ymgmr.2020.100692. eCollection 2020 Dec.
We previously showed that the genotype-phenotype correlation in MPS II is well-conserved in Japan (Kosuga et al., 2016). Almost all of our patients with attenuated MPS II have missense variants, which is expected to result in residual activity of iduronate-2-sulfatase. In contrast, our patients with severe MPS II have so-called null-type disease-associated variants, such as nonsense variants, frame-shifts, gene insertions, gene deletions and rearrangement with pseudogene (), none of which are expected to result in residual activity. However, we recently encountered a patient with attenuated MPS II who had a presumable null-type disease-associated variant and 76-base deletion located in exon 1 that extended into intron 1. To investigate this discordance, we extracted RNA from the leukocytes of the patient and performed reverse transcription polymerase chain reaction. One of the bands of the cDNA analysis was found to include a nucleotide sequence whose transcript was expected to generate an almost full-length IDS mature peptide lacking only part of its signal peptide as well as only one amino acid at the end of the N-terminus. This suggests that an alternative splicing donor site is generated in exon 1 upstream of the deleted region. Based on these observations, we concluded that the phenotype-genotype discordance in this patient with MPS II was due to the decreased amount of IDS protein induced by the low level of the alternatively spliced mRNA, lacking part of the region coding for the signal peptide but including the region coding almost the full mature IDS protein. The first 25 amino acids at the N-terminus of IDS protein are a signal peptide. The alternative splice transcript has only 13 (1 M-13 L) of those 25 amino acids; 14G-25G are missing, suggesting that the exclusively hydrophobic 1 M-13 L of the signal peptide of IDS might have a crucial role in the signal peptide.
我们之前的研究表明，黏多糖贮积症II型（MPS II）的基因型-表型相关性在日本人群中得到了很好的保留（小菅等人，2016年）。几乎所有我们研究的症状较轻的MPS II患者都有错义变异，预计这会导致艾杜糖醛酸-2-硫酸酯酶有残余活性。相比之下，我们研究的症状严重的MPS II患者有所谓的无效型疾病相关变异，如无义变异、移码突变、基因插入、基因缺失以及与假基因的重排（），预计这些变异都不会产生残余活性。然而，我们最近遇到了一名症状较轻的MPS II患者，其有一个可能的无效型疾病相关变异，且外显子1中有一个76个碱基的缺失，该缺失延伸到了内含子1中。为了研究这种不一致性，我们从该患者的白细胞中提取了RNA，并进行了逆转录聚合酶链反应。在cDNA分析的条带中发现其中一个包含一个核苷酸序列，其转录本预计会产生一个几乎全长的IDS成熟肽，该肽仅缺少部分信号肽以及N端末尾的一个氨基酸。这表明在缺失区域上游的外显子1中产生了一个可变剪接供体位点。基于这些观察结果，我们得出结论，该MPS II患者的表型-基因型不一致是由于可变剪接mRNA水平较低导致IDS蛋白量减少所致，该mRNA缺少部分编码信号肽的区域，但包含几乎完整的成熟IDS蛋白编码区域。IDS蛋白N端的前25个氨基酸是信号肽。可变剪接转录本仅包含这25个氨基酸中的13个（1M - 13L）；14G - 25G缺失，这表明IDS信号肽中仅有的疏水性1M - 13L可能在信号肽中起关键作用。
