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蛋白，可合成端粒G-RICH链DNA DE NOVO。它是一种专门的逆转录酶，其中包含催化成分，TERT和模板RNA，可以更好地了解端粒酶的调节机制，在这项研究中，我们首先建立了TERT^<-1->基因敲除小鼠。第一代TERT^<-1->基因敲除小鼠没有表现出任何表型，表明端粒酶催化蛋白的缺乏本身不会影响小鼠的发育。但是，当将TERT^<-1->小鼠配对以保持端粒酶不存在的线时，第五代小鼠显示出明显的萎缩生殖系细胞。这些结果表明生殖细胞对端粒长度的减小特别敏感。我们和其他人已经证明，TERT基因表达水平在调节细胞中端粒酶活性中具有主要作用。尽管据报道，包括MYC，SP1，ER在内的几个转录因子都涉及……更多的TERT表达，但调节TERT的机制的细节，尤其是在正常的体细胞中，仍然在很大程度上未知。我们已经研究了正常的人外周淋巴细胞作为模型系统。我们发现，转录因子USF1和2在体外积极调节TERT启动子。已知USF1和2，Mini-USF1和Mini-USF2的截短形式是通过替代剪接从同一基因产生的。有趣的是，我们证明了Mini-USF1和2在体外对启动子进行负调节。人外周淋巴细胞主要是静止，不具有端粒酶活性。将外部刺激应用于静止淋巴细胞时，它们开始增殖并同时开始显示端粒酶活性。我们发现阴性调节剂Mini-USF存在于静息细胞中，但在活化的淋巴细胞中不存在。相比之下，在静止和生长的淋巴细胞中都存在正调节剂，全长USF。这些结果表明，在控制TERT表达方面，Mini-USF可能对全USF产生主要的负面影响。较少的