Studies on the roles of RecQ family proteins in DNA repair and recombination in relation to carcinogenesis and aging

RecQ家族蛋白在与癌变和衰老相关的DNA修复和重组中的作用研究

基本信息

批准号：
09470498
负责人：
ENOMOTO Takemi
金额：
$ 6.27万
依托单位：
Tohoku University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1997
资助国家：
日本
起止时间：
1997 至 1998
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-09470498/
关键词：
RecQ protein DNA helicase Q1 Bloom's syndrome Carcinogenesis Aging DNA topoisomerase III Werner's syndrome Rad18 ワーナー症候群

项目摘要

There exist at least three RecQ homologues in higher eukaryotic cells, DNA helicase Q1, the Bloom's syndrome gene and Werner's syndrome gene product. To clarify the functions of the eukaryotic Rec Q family DNA helicases in DNA repair and recombination and to get an insight into the molecular bases of Bloom's syndrome and Werner's syndrome, we analyzed expression of mRNAs of RecQ family proteins and DNA topoisomerase III which is speculated to function with RecQ helicases, and searched proteins which interact with RecQ family proteins. In addition, we analyzed functional domains of RecQ helicases using gene disruptants of SGS1, which is the yeast RecQ homologue.1. We cloned cDNAs encoding mouse DNA helicase Q1, Bloom's syndrome helicase, DNA topoisomerase IIIalpha and a novel DNA topoisomerase III, IIIbeta (TOP3 beta). We found that messages of these four proteins were highly expressed in the testis. The levels of these mRNAs in the testis increased after birth when the cells in the pachytene phase began to appear and increase.2. We screened for proteins using the yeast two-hybrid system with mouse Wrn as bait and identified three proteins. One is a novel protein, Wip1 (Werner interacting protein 1) and the other two are UBC9 and SUMO-1 (small ubiquitin-related modifier-1), indicating that the function of Werner helicase is regulated by the SUMO- 1 conjugation system.3. We introduced mutations, which were found in the genes of Bloom's syndrome and Werner's syndrome patients, into SGS1 gene and analyzed phenotypes of the yeasts transfected with mutated genes. We found that two different mechanisms are operated in RecQ functions, one requiring DNA helicase activity and one not.

在高等真核细胞，DNA解旋酶Q1，Bloom综合征基因和Werner综合征基因产物中，至少存在三个RECQ同源物。为了阐明真核REC Q家族DNA解旋酶在DNA修复和重组中的功能，并深入了解Bloom综合征和Werner综合征的分子碱基，我们分析了RECQ家族蛋白和DNA拓扑酶III的mRNA表达与RECQ解旋酶一起起作用，并搜索与RECQ家族蛋白相互作用的蛋白质。此外，我们使用SGS1的基因破坏因子（即Yeast RecQ同源物）分析了RECQ解旋酶的功能域。1。我们克隆了编码小鼠DNA解旋酶Q1，Bloom的综合征解旋酶，DNA拓扑异构酶IIIALPHA和新型DNA DNA拓扑异构酶III，IIIBETA（TOP3 Beta）的CDNA。我们发现，这四种蛋白质的信息在睾丸中高度表达。当Pachytene阶段的细胞开始出现并增加时，睾丸中这些mRNA的水平有所增加。2。我们使用酵母两杂交系统（用小鼠WRN作为诱饵）筛选蛋白质，并鉴定出三种蛋白质。一种是一种新型的蛋白质，WIP1（Werner相互作用蛋白1），另一种是Ubc9和Sumo-1（小型泛素相关的修饰符1），表明Werner解旋酶的功能受SUMO-1共轭系统调节。 3。我们引入了在Bloom综合征和Werner综合征患者的基因中发现的突变，并将其引入SGS1基因，并分析了用突变基因转染的酵母的表型。我们发现在RECQ功能中运行了两种不同的机制，一种需要DNA解旋酶活性，一种不需要。