Mechanisms of SIRT2 Function in the DNA Damage Response

项目摘要 该建议的目的是确定SIRT 2在指导DNA损伤中发挥的关键作用 通过乙酰基组的翻译后修饰，将其置于 细胞代谢、衰老、致癌和肿瘤细胞对抗癌剂的抗性的发展。 SIRT 2是一种沉默调节蛋白家族脱乙酰酶，其感知能量需求并指导细胞过程以维持 代谢平衡值得注意的是，缺乏Sirt 2的小鼠会患乳腺癌、肝癌和其他癌症，这意味着 SIRT 2是一种肿瘤抑制因子巧合的是，SIRT 2在许多人类癌症中过表达，包括 此外，我们的数据表明，SIRT 2高表达与乳腺癌的低表达相关。 接受辅助电离辐射（IR）治疗的乳腺癌患者的生存率， 许多类型的癌症治疗，包括IR，PARP抑制剂和化疗。然而，精确 SIRT 2指导DDR控制肿瘤细胞治疗抗性的机制，以及SIRT 2如何指导DDR控制肿瘤细胞治疗抗性的机制。 SIRT 2本身在DDR中受到调控，人们对此知之甚少。在上一个资助期，我们定义了一部小说， SIRT 2通过ATRIP的乙酰化状态在指导复制应激反应（RSR）中的作用， CDK 9，并且进一步表明体细胞SIRT 2突变损害SIRT 2在维持CDK 9表达中的活性。 基因组完整性在此更新申请中，我们建议通过调查新角色来进一步开展这项工作 对于SIRT 2通过MRE 11的脱乙酰化指导DNA双链断裂（DSB）的修复， 鉴定控制SIRT 2活性的上游泛素化信号传导事件，并确定SIRT 2是否 抑制是治疗对常规疗法有抗性的乳腺癌的有效治疗策略。 在这方面，我们已经表明SIRT 2脱乙酰酶活性介导癌细胞对IR的抗性， PARP抑制剂通过促进同源重组修复中的末端切除步骤。我们进一步确认 一种新的DDR蛋白网络，通过SIRT 2去乙酰化以响应IR，包括MRE 11。而且我们 数据表明SIRT 2活性而不是水平在DDR中通过蛋白质单泛素化调节。我们 假设SIRT 2响应上游泛素化信号传导以维持基因组完整性并控制 乳腺癌治疗抗性通过指导DDR通过脱乙酰化的关键底物，包括 MRE 11，可用于改善乳腺癌控制。我们建议：（1）确定 SIRT 2在DDR中指导MRE 11; 2）剖析SIRT 2由以下调节的机制： 3）建立SIRT 2作为乳腺癌耐药的新分子靶点; DNA破坏剂。完成这项工作将提供一个详细的机械理解， SIRT 2指导DSB修复以维持基因组完整性并控制肿瘤细胞治疗抗性，定义了一种新的治疗方法。 通过DDR中的蛋白质单泛素化调节SIRT 2的新机制，并建立 SIRT 2抑制作为克服乳腺癌治疗耐药性的新治疗方法的概念。

PROJECT SUMMARY The objective of this proposal is to define the critical role that SIRT2 plays in directing the DNA damage response (DDR) through post-translational modification of the acetylome, which places it at the confluence of cellular metabolism, aging, carcinogenesis, and development of tumor cell resistance to anticancer agents. SIRT2 is a sirtuin family deacetylase that sense energy requirements and direct cellular processes to maintain metabolic homeostasis. Significantly, mice deficient in Sirt2 develop breast, liver, and other cancers, implying that SIRT2 is a tumor suppressor. Paradoxically, SIRT2 is overexpressed in many human cancers, including breast malignancies, and furthermore, our data suggest that high SIRT2 expression is associated with poor survival in breast cancer patients treated with adjuvant ionizing radiation (IR) and contributes to resistance to many types of cancer treatments, including IR, PARP inhibitor, and chemotherapy. However, the precise mechanisms by which SIRT2 directs the DDR to govern tumor cell treatment resistance, and moreover, how SIRT2 itself is regulated in the DDR are poorly understood. In the previous funding period, we defined a novel role for SIRT2 in directing the replication stress response (RSR) through the acetylation status of ATRIP and CDK9, and furthermore, showed that somatic SIRT2 mutations impair the activity of SIRT2 in maintaining genome integrity. In this renewal application, we propose to carry this work further by investigating a new role for SIRT2 in directing the repair of DNA double-strand breaks (DSB) through deacetylation of MRE11, identifying the upstream ubiquitination signaling events governing SIRT2 activity, and determining if SIRT2 inhibition is an effective therapeutic strategy for treatment of breast cancers resistant to conventional therapies. In this regard, we have shown that SIRT2 deacetylase activity mediates resistance of cancer cells to IR and PARP inhibitor by facilitating the end resection step in homologous recombination repair. We further identified a novel network of DDR proteins deacetylated by SIRT2 in response to IR, including MRE11. Moreover, our data suggest that SIRT2 activity but not levels is regulated in the DDR via protein monoubiquitination. We hypothesize that SIRT2 responds to upstream ubiquitination signaling to maintain genome integrity and govern breast cancer treatment resistance by directing the DDR through deacetylation of key substrates, including MRE11, which may be exploited to improve breast cancer control. We propose to: 1) Determine the role of SIRT2 in directing MRE11 in the DDR; 2) Dissect the mechanism by which SIRT2 is regulated by monoubiquitination in the DDR; 3) Establish SIRT2 as a new molecular target for breast cancer resistant to DNA damaging agents. Completion of this work will provide a detailed mechanistic understanding of how SIRT2 directs DSB repair to maintain genome integrity and govern tumor cell treatment resistance, define a novel mechanism for regulation of SIRT2 via protein monoubiquitination in the DDR, and establish proof of concept for SIRT2 inhibition as novel therapeutic approach for overcoming breast cancer treatment resistance.