Mechanism for Telomeric G-overhang Generation in Human Cells

描述（由申请人提供）：端粒通过防止染色体发生不适当的融合、降解和重组来保护基因组稳定性。端粒保护的失败导致基因组不稳定，导致癌症和过早衰老等疾病。端粒通过形成称为t环的独特DNA-蛋白质结构来实现其保护功能。T环的破坏导致端粒末端暴露，触发DNA损伤反应并诱导细胞衰老或凋亡。因此，诱导端粒功能障碍是一种很有前途的癌症治疗方法。T环的一个组成部分，单链富含G的突出端（G-突出端），在调节端粒稳态中具有多种重要作用：（1）它对于T环结构的形成是必需的，（2）它对于端粒酶与端粒的结合是必需的，（3）它对于在某些癌细胞中使用端粒酶非依赖性途径延长端粒是必需的，（4）G-突出端的产生过程有助于端粒缩短。因此，破译G-突出端生成的机制对于理解端粒在端粒酶表达和非表达癌细胞中的维持是至关重要的。然而，在人类细胞中产生G-突出端的分子机制知之甚少。缺乏这种知识是设计有效的癌细胞治疗策略的一个重要障碍。 我们的初步研究表明，端粒G-突出端是由细胞周期中多个严格调控的步骤产生的。我们还揭示了细胞周期蛋白依赖性激酶1（CDK 1）在控制G-突出端生成和维持癌细胞中功能性端粒中的未鉴定作用。核心假设是G-突出端是在CDK 1的控制下通过细胞周期中的多个严格调控步骤产生的。具体目标是：（1）定义在细胞周期期间负责在前导和滞后子端粒处产生G-突出端的分子事件。(2)确定CDK 1调节G-突出端生成的分子途径。我们将结合联合收割机生物化学和分子生物学的方法来确定CDK 1调节端粒维持的途径。 这些研究结果将进一步加深我们对维持端粒完整性和基因组稳定性的机制的理解，有助于我们理解癌细胞中端粒的维持机制，并有助于开发新的癌症治疗方案。 公共卫生相关性：端粒完整性的维持对于癌细胞的无限生长至关重要。该提案旨在对癌细胞中的端粒维持、细胞生长控制和肿瘤发生产生新的见解。它可能会导致新的抗癌疗法的发展。

DESCRIPTION (provided by applicant): Telomeres protect genome stability by preventing chromosomes from inappropriate fusions, degradation, and recombination. Failure of telomere protection results in genome instability that leads to diseases such as cancer and premature aging. Telomeres achieve their protection function through the formation of distinct DNA-protein structures termed t-loops. Disruption of t-loop leads to exposure of telomere ends, triggering DNA damage response and inducing cellular senescence or apoptosis. Therefore, inducing telomere dysfunction is a promising approach for cancer therapy. One component of t-loop, the single-stranded G-rich overhang (G- overhang), has multiple vital roles in regulating telomere homeostasis: (1) it is essential for the formation of the t-loop structure, (2) it is required for telomerase binding to telomeres, (3) it is needed for lengthening telomeres in certain cancer cells using telomerase-independent pathway, and (4) the process for generating G-overhang contributes to telomere shortening. Thus, deciphering the mechanism for G-overhang generation is crucial for understanding telomere maintenance in both telomerase-expressing and non-expressing cancer cells. However, the molecular mechanism for generating G-overhangs is poorly understood in human cells. Lack of such knowledge is a significant barrier for devising effective therapeutic strategies for cancer cells. Our preliminary studies indicate that telomeric G-overhangs are generated by multiple tightly regulated steps during the cell cycle. We have also revealed the unidentified role of cyclin-dependent kinase 1 (CDK1) in controlling G-overhang generation and in maintaining functional telomeres in cancer cells. The central hypothesis is that G-overhangs are generated by multiple tightly regulated steps during the cell cycle under the control of CDK1. The specific aims are: (1) Define the molecular events responsible for G-overhang generation at leading and lagging daughter telomeres during the cell cycle. (2) Determine the molecular pathway by which CDK1 regulates G-overhang generation. We will combine biochemical and molecular approaches to determine the pathway by which CDK1 regulates telomere maintenance. The findings from these studies will advance our understanding in the mechanism for maintaining telomere integrity and genome stability, aid us in understanding telomere maintenance mechanism in cancer cells and in developing new regimens for cancer therapy. PUBLIC HEALTH RELEVANCE: Maintenance of telomere integrity is essential for the unlimited growth of cancer cells. This proposal aims to yield new insights into telomere maintenance in cancer cells, cell growth control and oncogenesis. It may lead to the development of new anti-cancer therapy.