Study on pathomechanism of Dyschromatosis Symmetrica Hereditaria caused by gene mutation of RNA editing enzyme, DSRAD

对称遗传性色素沉着症（DSH）是一种常染色体显性遗传的色素疾病。2003年，我们首次报道了双链rna特异性腺苷脱氨酶（DSRAD/ADAR1）是DSH的负责基因。ADAR1蛋白在双链RNA底物中催化腺苷脱氨为肌苷，导致剪接位点的改变或密码子的改变，从而导致蛋白质的功能改变。本研究旨在阐明ADAR1基因突变患者DSH发生的病理机制。(1)据报道，人类ADAR1蛋白包括两种主要形式，一种是干扰素诱导的全长150-kDa蛋白（p150），另一种是组成型表达的n端截短110-kDa蛋白（p110）。这两种形式的产生是由于不同的促进剂的存在。在本研究中，我们发现了两个突变引起t细胞的移码变化，更多的是p150的合成，但对p110蛋白的合成有影响。因此我们认为全长p150一定与DSH有关。(2)为了鉴定p150ADAR1的RNA底物，我们尝试使用以下两种策略筛选几种候选底物：(a)干扰素诱导的p150ADAR1基因在培养的黑色素细胞中的寡核苷酸阵列分析。由于p150ADAR1是由干扰素诱导的，而转染siADAR1有报道能显著抑制ADAR1的表达，因此我们利用人基因阵列分析比较了用干扰素培养的正常人黑素细胞与p150ADAR1短抑制mRNA （sirna）的细胞总RNA表达。(b) A-to-I编辑位点rna的计算鉴定。原则上，可以使用表达序列标签（est）和rna的大规模数据库来检测编辑。当一个序列与基因组对齐时，编辑位点就会出现：当DNA读取a时，测序将编辑位点中的肌苷识别为鸟苷。如上所述的两次实验结果，我们列出了许多候选RNA，但没有确定其中特定的RNA底物。少

Dyschromatosis symmmetrica hereditaria (DSH) is a pigmentary disorder with an autosomal dominant inheritance. In 2003, we for the first time reported that the double-stranded RNA-specific adenosine deaminase (DSRAD/ADAR1) is the responsible gene for DSH. ADAR1 protein catalyzes the deamination of adenosine to inosine in double-stranded RNA substrates, which results in the creation of alternative splicing sites or alternations of the codon and thus leads to functional changes in the protein. Purpose of this study was to clarify the pathomechanism how DSH developed in the patient with the gene mutation of ADAR1.(1) Human ADAR1 protein has been reported to include two major forms, one is the interferon-inducible full-length 150-kDa protein (p150) and the other is the constitutively expressed N-terminally truncated 110-kDa protein (p110). The production of the two forms is due to the presence of different promoters. In this study, we have found two mutations causing frameshift changes in t … More he synthesis of p150, but an effect on the synthesis of p110 protein. Thus we can suggest that the full-length p150 must be involved with DSH.(2) In order to identify the RNA substrate for p150ADAR1, we tried to screen out several candidates as the substrate(s) using two strategies as follows: (a) Oligonucleotide array analyses of interferon-inducible p150ADAR1 genes in cultured melanocytes. As the p150ADAR1 is induced by interferon, and transfection with siADAR1 was reported to dramatically inhibite the expression of ADAR1, we compared the total cellular RNA expression in normal human melanocytes cultured with interferon and that with the short inhibitory mRNA (siRNAs) of p150ADAR1 by analyses using the human gene arrays. (b) Computational identification of RNAs with A-to-I editing sites. In principle, editing can be detected using the large-scale database of expressed sequence tags (ESTs) and RNAs. Editing sites show up when a sequence is aligned with the genome: while the DNA reads A, sequencing identifies the inosine in the edited site as guanosine.As the results of two experiments as mentioned above, we listed many candidate RNAs, but have identified no specific RNA substrate(s) among them. Less