Telomere structure and function in Arabidopsis

拟南芥端粒结构和功能

• 批准号: 7096299
• 负责人: Dorothy Shippen
• 金额: $ 28.78万
• 依托单位: TEXAS A&M UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-05-01 至 2010-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7096299
• 关键词: Arabidopsis; DNA; cell cycle; cell growth regulation; chemical stability; chemical structure function; chromosome disorders; chromosomes; genetic recombination; genetically modified plants; intermolecular interaction; laboratory rabbit; meiosis; molecular cloning; molecular site; nucleic acid structure; nucleoproteins; plant genetics; plant growth /development; plant physiology; plant proteins; polymerase chain reaction; southern blotting; telomerase; telomere; yeast two hybrid system

DESCRIPTION (provided by applicant): Telomeres are higher order nucleoprotein structures that physically cap the chromosome terminus and help to preserve genome integrity. In cells with unlimited proliferative capacity, including 95% of human cancers, telomeres are maintained by telomerase. By contrast, telomerase is absent from most of the normal soma and telomeres shorten until chromosome ends become uncapped and indistinguishable from double strand breaks. End-to-end fusions are induced, ultimately leading to cell cycle arrest. Here we propose to exploit Arabidopsis to investigate essential components of the telomere cap in a genetically tractable higher eukaryote. Arabidopsis has an exceptionally high tolerance for telomere dysfunction. This finding, coupled with its facile genetics, completely sequenced genome, and arsenal of transgenic tools offer unique opportunities for investigating essential genes in telomere biology. In this renewal application, we will focus on the recently discovered Protection of telomeres 1 (Pot1) protein, which binds the extreme terminus of the chromosome and plays a central role in chromosome end protection and telomere length regulation. Arabidopsis encodes three strikingly different Pot proteins, AtPot1, AtPot2 and AtPot3, which appear to be functionally non-redundant. We hypothesize that the separation of function of Arabidopsis Pot proteins derives from their distinct interactions with telomeric DNA and with other protein components of the telomere. The proposal is comprised of five Specific Aims. The first two focus on defining the interactions of Pot proteins with telomeric DNA in vitro and in vivo, and elucidating interaction partners in the telomere complex in vivo. For the last three Aims, we will exploit a combination of genetic and biochemical approaches to examine the mechanism of positive regulation of telomerase by AtPot1 (Aim 3), the contribution of AtPot2 to chromosome end protection (Aim 4), and the apparent developmental regulation of AtPot3 as it impacts this protein's function at telomeres (Aim 5). As part of Aim 5, we will also test the hypothesis that telomeres are uncapped in a specific stage of the plant life cycle. Given the strong conservation in telomere architecture and composition in plants and humans, these studies should uncover mechanisms common to all higher eukaryotes.

描述（由申请人提供）：端粒是高阶核蛋白结构，可以物理限制染色体末端并有助于保留基因组完整性。在不受限制的增殖能力（包括95％的人类癌症）的细胞中，端粒可以维持端粒酶。相比之下，大多数正常体体不存在端粒酶，而端粒缩短，直到染色体末端变得不封闭并且与双链断裂无法区分。诱发端到端融合，最终导致细胞周期停滞。在这里，我们建议利用拟南芥，以研究遗传上较高的真核生物中端粒帽的基本成分。拟南芥对端粒功能障碍具有极高的耐受性。这一发现，再加上其简便的遗传学，完全测序的基因组和转基因工具的阿森纳为研究端粒生物学中的基本基因提供了独特的机会。在此续订应用中，我们将重点关注最近发现的端粒1（POT1）蛋白的保护，该蛋白结合了染色体的极端末端，并在染色体终端保护和端粒长度调节中起着核心作用。拟南芥编码三种截然不同的锅蛋白ATPOT1，ATPOT2和ATPOT3，它们在功能上似乎不是冗余。我们假设拟南芥蛋白的功能分离来自它们与端粒DNA的不同相互作用以及端粒的其他蛋白质成分。该提案由五个具体目标组成。前两个重点是在体外和体内定义锅蛋白与端粒DNA的相互作用，并阐明体内端粒复合物中的相互作用伙伴。在最后三个目标中，我们将利用遗传和生物化学方法的组合，以检查topot1对端粒酶阳性调控的机制（AIM 3），ATPOT2对ATPOT2对染色体终端保护的贡献（AIM 4），以及ATPOT3的明显发育调节ATPOT3，因为它会影响该蛋白质在Telomeres的功能（AIM 5）。作为AIM 5的一部分，我们还将检验以下假设：在植物生命周期的特定阶段，端粒是未覆盖的。鉴于植物和人类中端粒结构和组成的强大保护，这些研究应揭示所有较高真核生物常见的机制。