Novel regulations of DNA damage repair

项目总结 DNA损伤反应和DNA修复缺陷是基因组不稳定和 肿瘤发生学。更好地了解DNA修复所涉及的途径不仅可以增加我们的 对癌症病因的了解，也为癌症治疗提供了新的靶点。一种关键的蛋白质参与了 DNA修复和肿瘤发生是P53结合蛋白1，即53BP1。 我的实验室多年来一直在研究53BP1。我们的团队是最早证明 53BP1在DNA损伤反应中的作用我们建立了第一只53BP1基因敲除(KO)小鼠；我们还 揭示了53BP1是DNA修复所必需的，并在体内起到了肿瘤抑制作用。此外，我们 阐明了53BP1在DNA损伤后的调控。我们和其他人证明了依赖H_2AX的 DNA损伤信号通路控制着53BP1在DNA断裂部位的募集和积聚。 然而，53BP1也可以以不依赖于H2 AX的方式定位到DNA损伤部位，尽管 潜在的机制仍有待确定。此外，我们还证明了53BP1对于特定的 修复过程称为类交换重组，表明53BP1参与了一种特殊的DNA修复 与典型的非同源末端连接途径明显不同的途径。我们最近的研究 其他人的研究表明，53BP1抑制BRCA1缺陷的同源重组修复 细胞，这对基于聚(ADP-核糖)聚合酶抑制剂的癌症的反应至关重要 治疗。总之，这些数据突显了53BP1在对抗同源重组中的重要性 修复以应对DNA损伤。在这个提案中，我们计划将重点放在53BP1上，并在分子水平上阐明 53BP1在DNA损伤后如何受到调控，并有助于DNA修复和基因组维护。 为了进一步了解53BP1在DNA损伤部位的定位和功能调节，我们最近 进行串联亲和纯化和质谱分析，以鉴定能够 与53BP1区域特异性结合，这是其定位到DNA所必需和充分的 损毁地点。令人惊讶的是，我们发现了几种新的53BP1结合蛋白，包括NUDT16， NUDT16L1和DEK.在本提案中，我们将1)进一步确定NUDT16L1和NUDT16在 53BP1的调控和在DNA损伤反应中的作用；2)阐明DEK和DEK的功能意义。 其他新发现的53BP1相关蛋白在损伤诱导的53BP1定位、DNA修复和 保持基因组的完整性。这些研究将帮助我们了解起作用的关键成分 它位于53BP1的上游，与53BP1一起参与DNA修复和基因组维护。

PROJECT SUMMARY Defects in DNA damage response and DNA repair are the driving forces of genomic instability and tumorigenesis. Gaining a better understanding of the pathways involved in DNA repair not only increases our understanding of cancer etiology, but also provides new targets for cancer therapies. A key protein involved in DNA repair and tumorigenesis is p53-binding protein 1, i.e. 53BP1. My laboratory has been working on 53BP1 for many years. Our group was one of the first to demonstrate the role of 53BP1 in DNA damage response. We established the first 53bp1 knockout (KO) mice; we also revealed that 53BP1 is required for DNA repair and acts as a tumor suppressor in vivo. In addition, we elucidated the regulation of 53BP1 after DNA damage. We and others demonstrated that the H2AX-dependent DNA damage signaling pathway controls the recruitment and accumulation of 53BP1 at sites of DNA breaks. However, 53BP1 can also localize to DNA damage sites in an H2AX-independent manner, although the underlying mechanisms remain to be determined. Moreover, we showed that 53BP1 is critical for a particular repair process called class-switch recombination, indicating that 53BP1 is involved in a special DNA repair pathway that is distinctly different from the canonical nonhomologous end-joining pathway. Our recent studies and those of others suggest that 53BP1 suppresses homologous recombination repair in BRCA1-deficient cells, which is critically important for response to poly (ADP-ribose) polymerase inhibitor-based cancer therapies. Together, these data highlight the importance of 53BP1 in counteracting homologous recombination repair in response to DNA damage. In this proposal, we plan to focus on 53BP1 and elucidate at the molecular level how 53BP1 is regulated after DNA damage and contributes to DNA repair and genome maintenance. To further understand the regulation of 53BP1 localization and function at DNA damage sites, we recently performed tandem affinity purification coupled with mass spectrometry analysis to identify proteins that would specifically associate with a region of 53BP1, which is necessary and sufficient for its localization to DNA damage sites. Surprisingly, we uncovered several novel 53BP1-binding proteins, including NUDT16, NUDT16L1, and DEK. In this proposal, we will 1) further determine the roles of NUDT16L1 and NUDT16 in 53BP1 regulation and in the DNA damage response, and 2) elucidate the functional significance of DEK and other newly discovered 53BP1-associated proteins in damage-induced 53BP1 localization, DNA repair, and the maintenance of genomic integrity. These studies will help us understand the key components that act upstream of 53BP1 and function together with 53BP1 in DNA repair and genome maintenance.