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Basic study on the mechanisms of carcinogenesis and aging with special reference to the function of RecQ family helicases

致癌和衰老机制的基础研究，特别是RecQ家族解旋酶的功能

基本信息

批准号：
11307055
负责人：
ENOMOTO Takemi
金额：
$ 24.75万
依托单位：
Tohoku University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (A)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2001
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11307055/
关键词：
RecQ family Helicase Werner syndrome WHIP WRN carcinogenesis aging yeast Sgs1 RecQL1 ワーナー症候群 Whip ブルーム症候群

项目摘要

In this study, we tried to clarify the function of RecQ family helicases to understand the genomic instability resulting in predisposition to cancer and premature aging due to the defect in RecQ family helicases, and obtained following results by using budding yeasts and chicken DT40 cells, which are easily applicable to genetic analysis, as well as cultured human cells.1. It has been indicated by generating and analyzing BLM-/- and BLM-/-/RAD54-/- DT40 cells that the majority of increased SCEs in BLM disrupted cells are formed via homologous recombination and BLM functions to decrease the formation of DNA double strand breaks during DNA replication.2. Werner syndrome gene product, WRN, interacted with Ubc9 and SUMO-1 and was actually conjugated with SUMO-1.3. A novel Werner helicase interacting protein, WHIP, which we found, was co-localized with WRN in the nuclei, and the interaction of these proteins required ATP, indicating a dynamic interaction of these proteins.4. The genetic analyzes using yeast mutants that overexpression of WHIP in mutants of DNA polymerase δ, RFC, or PCNA resulted in the lethality of the cells indicated the existence of functional relationship between WHIP and DNA polymerase δ, RFC, and PCNA. In addition, physical interaction between WHIP and DNA polymerase δ was indicated by the yeast-two hybrid system.5. Analyzes using yeasts also indicated the genetical interaction of the yeast BLM/WRN homologue, Sgs1 with Mre11, Sld3, and Sld5 which are involved in the initiation of DNA replication.

在本研究中，我们试图阐明RecQ家族解旋酶的功能，以了解由于RecQ家族解旋酶缺陷导致的基因组不稳定性导致的癌症易感性和过早衰老，并通过使用易于适用于遗传分析的芽殖酵母和鸡DT 40细胞以及培养的人类细胞获得了以下结果。通过对BLM-/-和BLM-/-/RAD 54-/-DT 40细胞的产生和分析表明，BLM破坏细胞中增加的SCE大多数是通过同源重组形成的，BLM的功能是减少DNA复制过程中DNA双链断裂的形成. Werner综合征基因产物WRN与Ubc 9和SUMO-1相互作用，实际上与SUMO-1.3结合。我们发现一种新的Werner解旋酶相互作用蛋白WHIP与WRN共定位于细胞核中，并且这些蛋白的相互作用需要ATP，表明这些蛋白之间存在动态相互作用.用酵母突变体进行的DNA聚合酶δ、RFC和PCNA突变体中WHIP过表达导致细胞死亡的遗传分析表明，WHIP与DNA聚合酶δ、RFC和PCNA之间存在功能关系。此外，酵母双杂交系统表明WHIP与DNA聚合酶δ之间存在物理相互作用.使用酵母的分析还表明酵母BLM/WRN同源物Sgs 1与参与DNA复制起始的Mre 11、Sld 3和Sld 5的遗传相互作用。