Telomere Structure and Function in Arabidopsis

描述(申请人提供)：端粒是更高级的核蛋白 在物理上覆盖染色体末端并有助于保存的结构 基因组完整性。在具有无限增殖能力的细胞中，包括95 在人类癌症中，端粒是由端粒酶维持的。相反， 正常体细胞和端粒的大多数细胞都不存在端粒酶 无情地缩短，直到染色体末端变得没有上限和难以区分 从双链断裂。端到端融合被诱导，最终引领 到细胞周期停滞。在这里，我们建议利用阿拉伯的偶像来阐明 端粒帽的结构和功能，以更好地了解该设备如何 为基因组提供稳定性，并促进细胞的持续增殖。 我们的初步结果表明，拟南芥具有异常高的 对端粒功能障碍的耐受性。这一发现，再加上法西斯 拟南芥的遗传学、其完全测序的基因组和 转基因工具表明，这个系统可以提供独特的机会。 用于研究端粒生物学中的必要基因。的前两个目标 该方案利用了我们开发的端粒酶缺失的拟南芥模型 探索有帽和无帽染色体之间的分子转换。在AIM I，我们将检测细胞中端粒重组的频率 端粒变短，但功能正常。我们还将确定以下方面的贡献 体细胞和减数分裂重组对端粒维持的影响 端粒酶。在目标2中，我们将通过以下方式探索端粒融合的机制 在染色体连接处对DNA进行测序，并通过评估 这一过程中的非同源末端连接途径。一种遗传策略也 建议研究含有染色体融合的细胞的繁殖。这个 最后两个目标集中在形成染色体帽的蛋白质上。在《目标3》中，我们将 通过研究拟南芥中两个可能的POT1同源基因的功能 阐明它们的DNA结合特性和分子伙伴，并通过 确定过度表达和基因破坏的后果。在目标4中， 生物化学和遗传学方法被描述为发现 便于染色体封顶。因为几乎所有涉及到的哺乳动物基因 在端粒动态平衡中有一个拟南芥的对应物，这些实验 应该揭示所有多细胞生物体共有的机制。

DESCRIPTION (provided by applicant): Telomeres are higher order nucleoprotein structures that physically cap the chromosome terminus and help to perserve genome integrity. In cells with unlimited proliferative capacity, including 95 percent of human cancers, telomeres are maintained by telomerase. By contrast, telomerase is absent from most cells of the normal soma and telomeres inexorably 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 Arab idopsis to elucidate the structure and function of the telomere cap to better understand how this device provides stability to the genome, and facilitates continued cell proliferation. Our preliminary results indicate that Arabidopsis has an exceptionally high tolerance for telomere dysfunction. This finding, coupled with the fascile genetics of Arabidopsis, its completely sequenced genome, and arsenal of transgenic tools demonstrate that this system can offer unique opportunities for investigating essential genes in telomere biology. The first two goals of the proposal exploit the telomerase-deficient Arabidopsis model we developed to probe the molecular transition between capped and uncapped chromosomes. In Aim I, we will examine the frequency of telomere recombination in cells with shortened, but functional telomeres. We will also determine the contribution of somatic and meiotic recombination to telomere maintenance in the absence of telomerase. In Aim 2, we will explore the mechanism of telomere fusions by sequencing DNA at chromosome junctions, and by assessing the role of the non-homologous end-joining pathway in this process. A genetic strategy is also proposed to examine the propagation of cells harboring chromosome fusions. The last two aims focus on proteins that form the chromosome cap. In Aim 3, we will examine the function of two putative Pot1 orthologs in Arabidopsis by elucidating their DNA binding properties and molecular partners, and by determining the consequences of over-expression and gene disruption. In Aim 4, biochemical and genetic approaches are described to discover genes that facilitate chromosome capping. Because virtually every mammalian gene involved in telomere homeostasis has an Arabidopsis counterpart, these experiments should uncover mechanisms common to all multicellular organisms.