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Molecular Genetics of Human Albinism

人类白化病的分子遗传学

基本信息

批准号：
63480471
负责人：
KIKUCHI Hideaki
金额：
$ 3.97万
依托单位：
Tohoku University
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1988
资助国家：
日本
起止时间：
1988 至 1990
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-63480471/
关键词：
ALBINISM TYROSINASE TRANSGENIC MOUSE チロシナーゼ

项目摘要

1. A human tyrosinase gene containing a 5'-flanking region and the first exon was isolated from a normal human liver library using mouse tyrosinase cDNA as a probe for screening. In the 5'-upstream region of transcription start sites, there is a putative TATA box and CAAT box and some palindromic sequences.2. Enzymatic DNA amplification and direct DNA sequencing were used to detect a mutation in the tyrosinase gene of a Japanese patient with tyrosinase-negative albinism. Single-base change could be detected by direct sequencing. This base change (G to A) is thought to result in an amino acid change (Arg to Gln) in tyrosinase of the patient. The mutant allele can be also identified by Southem blot hybridization using MspI because the change destroys the MspI site.3. We introduced a human-mouse fused minigene, hg-tyrs-J, in which the human genomic 5' non-coding flanking sequence was fused to a mouse tyrosinase cDNA, into fertilized eggs of albino mice. We obtained two pigmented mice and established some transgenic lines. Histological analysis revealed that melanization was restricted to melanocytes in hair bulbs and eyes in transgenic mice. These results suggested that this minigene encodes active throsinase protein and that its 5'-flanking region contains the regulatory sequences for the expression and that there is a common or very similar mechanism between mice and humans in tyrosinase gene regulation.

1.以小鼠酪氨酸酶cDNA为探针，从正常人肝文库中克隆了一个含有5 ′侧翼区和第一外显子的人酪氨酸酶基因。在转录起始位点的5 '上游区域，存在一个推测的TATA盒和CAAT盒以及一些回文序列.酶的DNA扩增和直接DNA测序被用来检测酪氨酸酶基因的突变与酪氨酸酶阴性白化病的日本患者。通过直接测序可以检测到单个碱基的变化。认为这种碱基变化（G至A）导致患者酪氨酸酶中的氨基酸变化（Arg至Gln）。突变等位基因也可以通过使用MspI的Southem印迹杂交来鉴定，因为该变化破坏了MspI位点。我们将人基因组5'端非编码侧翼序列与小鼠酪氨酸酶cDNA融合的人-小鼠融合小基因hg-tyrs-J导入白化病小鼠受精卵中。我们获得了两只有色小鼠，并建立了一些转基因品系。组织学分析显示转基因小鼠的黑化仅限于毛球和眼睛中的黑素细胞。这些结果表明，该小基因编码活性酪氨酸酶蛋白，其5 '侧翼区含有表达调控序列，小鼠和人类之间在酪氨酸酶基因调控中存在共同或非常相似的机制。