KU, TELOMERE MAINTENANCE AND CELLULAR SENESCENCE

本计划的目的是研究Ku异源二聚体（Ku 70/80） 在哺乳动物端粒维持和细胞衰老中的作用。 Ku参与端粒维护的支持是由最近的 证明酵母的Ku同系物是端粒所必需的 上维护此外，已经表明酵母Ku与 端粒DNA在体内，表明Ku在端粒长度的直接作用 调控引人注目的是，尽管相对较低的序列保守性 酵母和人Ku亚基在氨基酸水平上的差异， 人Ku亚基在酵母中的表达拯救了Ku无效突变。在这 根据该提案，申请人提出了最近获得的新的初步证据， 在人类细胞中Ku与端粒DNA的关联。端粒 维持对于衰老是至关重要的，因为端粒酶的激活， 负责端粒DNA合成的酶，已被证明 直接控制原代人类细胞的复制衰老。申请人 对已知元素之间的相互作用进行了详细的研究 的端粒和Ku，以确定这些蛋白质在 端粒维持和复制衰老。申请人的具体目标 主要研究：1）Ku/端粒DNA在哺乳动物端粒上的相互作用 使用体内和体外实验方法; 2）是否已知哺乳动物 端粒结合蛋白介导Ku和端粒之间的相互作用; 3）胚胎干细胞中端粒的维持与胚胎干细胞的衰老 Ku 70、Ku 80、DNA-PKcs和 端粒酶RNA部分。该提案汇集了两个集团， 补充专业知识：伊丽莎白布莱克本博士的小组在大学 加州，旧金山弗朗西斯科，在端粒和端粒酶方面有专长， 洛斯阿拉莫斯国家实验室的大卫陈博士的小组在Ku方面有专长 异源二聚体和DNA-PKcs。这项工作的目标是形成一种理解， 这些关键成分，端粒和Ku异源二聚体， 在哺乳动物细胞中。这些研究对于进一步了解 端粒的维护，这将提供更好的理解细胞 衰老

The goal of this proposal is to study the Ku heterodimer's (Ku70/80) role in telomere maintenance and cellular senescence in mammals. Support for Ku's involvement in telomere maintenance is provided by the recent demonstration that the Ku homologs of yeast are required for telomere maintenance. Moreover, it has been shown that yeast Ku is in close proximity to telomeric DNA in vivo, suggesting a direct role for Ku in telomere length regulation. Strikingly, despite the relatively low sequence conservation between yeast and human Ku subunits at the amino acid level, the exogenous expression of human Ku subunits in yeast rescues a Ku null mutation. In this proposal, the applicant presents recently obtained novel preliminary evidence for an association of Ku with telomeric DNA in human cells. Telomere maintenance is critical for senescence, since activation of telomerase, the enzyme responsible for the synthesis of telomeric DNA, has been shown to directly control replicative senescence in primary human cell. The applicant proposes a detailed investigation into the interactions between known elements of the telomere and Ku to determine the role played by these proteins in telomere maintenance and replicative senescence. The applicant's specific aims are to investigate: 1) Ku/telomeric DNA interactions at the mammalian telomere using in vivo and in vitro experimental approaches; 2) whether known mammalian telomere binding proteins mediate an interaction between Ku and the telomere; and 3) telomere maintenance in embryonic stem cells and cellular senescence in mouse embryonic fibroblasts with various mutations in Ku70, Ku 80, DNA-PKcs and the RNA moiety of telomerase. This proposal brings together two groups with complementary expertise: Dr. Elizabeth Blackburn's group at the University of California, San Francisco with expertise in the telomere and telomerase, and Dr. David Chen's group at Los Alamos National Lab with expertise in the Ku heterodimer and DNA-PKcs. The goal of this work is to form an understanding of the interaction of these key components, the telomere and the Ku heterodimer, in mammalian cells. These studies are critical for a further understanding of telomere maintenance, which will provide a better understanding of cellular senescence.