Role of Telomerase is DSB Repair

双链断裂(DSB)是DNA损伤最有害的形式之一。他们出现了 在暴露于电离辐射或模拟放射性化学物质后，也会作为一种 正常细胞代谢的副产品。双链断裂的忠实修复是保存基因组的关键 正直，抑制恶变。对DSB修复机制的准确理解 不仅对了解肿瘤的发生很重要，而且还影响肿瘤对辐射的反应。 治疗和大多数化疗药物。在DSB修复过程中，端粒酶-一种正常催化的酶 端粒DNA在染色体末端的延伸--能够混杂地添加端粒 在染色体内DSB处重复，潜在地干扰准确的修复。以前的研究 研究表明，将端粒重复序列添加到染色体内区域(称为端粒 序列插入或TSI)会导致染色体断裂、重组和重排。 因此，TSI通常被抑制，以防止基因组不稳定。然而，它的作用是 端粒酶在DSB修复中的作用一直被忽视，目前还不清楚端粒酶是如何介导的 在人类和哺乳动物细胞中，DSB的TSI被抑制。我们最近的发现有力地支持了这一点 TSI是由于端粒酶在DSB上错误地添加端粒重复序列而引起的。此外，我们 已经确定MLH1是端粒酶表达细胞中的TSI抑制因子。使用特定于域的 MLH1的突变，我们还发现MLH1募集到DSB是抑制TSI所必需的。这个 该建议的中心假设是，作为对DSB诱导的反应，MLH1定位于 染色体内断裂部位以防止端粒酶在DSB上增加端粒重复， 从而确保准确的修复和保护基因组的稳定性。本R03的目标是 建议对TSI进行第二次分析，并建立一个明确的分子系统来 研究TSI背后的调控机制。在目标1中，我们将开发一个可诱导的DSB 修复系统，研究TSI。在目标2中，我们将定义各种DSB修复蛋白在 TSI抑制。拟议中的研究结果将获得新的见解和准确的 了解DSB修复和基因组不稳定性，并为开发新的 肿瘤管理的治疗策略。

Double-strand breaks (DSBs) represent one of the most deleterious forms of DNA damage. They arise following exposure to ionizing radiation or radio-mimetic chemicals, and are also generated as a byproduct of normal cellular metabolism. Faithful repair of DSBs is critical for preserving genome integrity and suppressing malignant transformation. Accurate understanding of DSB repair mechanisms is not only important for understanding tumorigenesis, but also impacts tumor response to radiation therapy and most chemotherapy agents. During DSB repair, telomerase — an enzyme normally catalyzing the extension of telomeric DNA at chromosome ends — is capable of promiscuously adding telomeric repeats at intra-chromosomal DSBs, potentially interfering with accurate repair. Previous studies have shown that addition of telomeric repeats into intra-chromosomal regions (known as telomere sequence insertion, or TSI) causes chromosome breakage, recombination, and rearrangements. Therefore, TSI is normally suppressed to prevent genome instability. However, the role of telomerase in DSB repair has long been neglected, and it remains unknown how telomerase-mediated TSI at DSBs is suppressed in human and mammalian cells. Our recent findings strongly support that TSI is caused by erroneous addition of telomeric repeats at DSBs by telomerase. In addition, we have identified MLH1 as a TSI suppressor in telomerase-expressing cells. Using domain-specific mutations of MLH1, we also find that MLH1 recruitment to DSBs is required for suppressing TSI. The central hypothesis of this proposal is that in response to DSB induction, MLH1 localizes at intra-chromosomal break sites to prevent telomerase from adding telomeric repeats at DSBs, therefore ensuring accurate repair and protecting genome stability. The objective of this R03 proposal is to perform a second analysis of TSI and establish a defined molecular system to investigate the regulatory mechanism underlying TSI. In Aim 1, we will develop an inducible DSB repair system to study TSI. In Aim 2, we will define the roles of various DSB repair proteins in TSI suppression. Findings from the proposed research will gain novel insights and accurate understanding of DSB repair and genome instabilities, and offer guidance in developing new therapeutic strategies for tumor management.