Molecular studies on mammalian telomerase

哺乳动物端粒酶的分子研究

基本信息

批准号：
10308031
负责人：
ISHIKAWA Fuyuki
金额：
$ 20.03万
依托单位：
Tokyo Institute of Technology
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (A).
财政年份：
1998
资助国家：
日本
起止时间：
1998 至 2000
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10308031/
关键词：
Telomere Telomerase TERT RAP1 RIF1 TAP1 テロメレース ES細胞 mvc MZF2

项目摘要

Telomerase is a ribonucleoprotein complex that synthesizes telomeric G-rich strand DNA de novo. It is a specialized reverse transcriptase, containing the catalytic component, TERT and the template RNA, TR.To better understand regulation mechanisms of telomerase, in this study, we first established TERT^<-1-> knockout mice. The first generation of TERT^<-1-> knockout mice did not show any phenotype, indicating that the absence of telomerase catalytic protein by itself does not affect mouse development. However, when TERT^<-1-> mice were mated to keep the line absent for telomerase, the fifth generation mice showed significantly hypotrophic germ line cells. These results indicate that germ cells are particularly sensitive to telomere length reduction. We and others have already demonstrated that TERT gene expression levels have a primary role in regulating telomerase activity in cells. Although it has been reported that several transcription factors, including Myc, Sp1, ER, are involved … More in TERT expression, details of mechanisms regulating TERT, especially in normal somatic cells, remain largely unknown. We have studied normal human peripheral lymphocytes as a model system. We found that the transcription factors USF1 and 2 positively regulate the TERT promoter in vitro. Truncated forms of USF1 and 2, mini-USF1 and mini-USF2, are known to be produced from the same genes by alternative splicing. Interestingly, we demonstrated that mini-USF1 and 2 negatively regulate the promoter in vitro. Human peripheral lymphocytes are mostly resting and do not possess telomerase activity. Upon external stimuli applied to resting lymphocytes, they start proliferation and simultaneously begin to show telomerase activity. We found that the negative regulator, mini-USFs are present in resting cells but not in activated lymphocytes. In contrast, the positive regulator, full length USFs are present both in resting and growing lymphocytes. These results suggest that mini-USFs might have dominant negative effects on full USFs in controlling TERT expression. Less

端粒酶是一种核糖核蛋白复合物，从头合成端粒富含G链DNA，是一种特殊的逆转录酶，含有催化成分TERT和模板RNA TR。为了更好地了解端粒酶的调控机制，在本研究中，我们首先研究端粒酶的调控机制。建立TERT^<-1->敲除小鼠第一代TERT^<-1->敲除小鼠没有表现出任何表型，表明端粒酶的缺失。然而，当TERT^-1-小鼠交配以保持该系不存在端粒酶时，第五代小鼠表现出显着的生殖系细胞营养不良。我们和其他人已经证明 TERT 基因表达水平在调节细胞中的端粒酶活性中起主要作用，尽管有报道称，包括 Myc、Sp1、ER 在内的几种转录因子对端粒长度缩短很敏感。尽管参与 TERT 表达，但调节 TERT 的机制（尤其是在正常体细胞中）的细节仍然很大程度上未知。我们研究了正常人外周淋巴细胞作为模型系统，我们发现转录因子 USF1 和 2 正向调节 TERT 启动子。在体外，已知 USF1 和 2（mini-USF1 和 mini-USF2）实际上是由相同基因通过选择性剪接产生的，我们证明 mini-USF1 和 2 对启动子具有负调控作用。人类外周淋巴细胞大多处于静息状态，不具有端粒酶活性。当对静息淋巴细胞施加外部刺激时，它们开始增殖，同时开始表现出端粒酶活性，但静息细胞中存在负调节因子。相反，正调节因子全长 USF 存在于静息淋巴细胞和生长淋巴细胞中。这些结果表明，迷你 USF 在控制 TERT 方面可能对全长 USF 具有显着的负面影响。表达量少