Mechanisms of SIRT2 Function in the DNA Damage Response

项目总结 这项提案的目的是确定SIRT2在指导DNA损伤中所起的关键作用 通过乙酰体的翻译后修饰来响应(DDR)，这使其处于 细胞代谢、衰老、癌变和肿瘤细胞对抗癌药耐药性的发展。 SIRT2是sirtuin家族的一种脱乙酰酶，它感知能量需求并指导细胞过程来维持 代谢动态平衡。值得注意的是，缺乏SIRT2的小鼠会患上乳腺癌、肝癌和其他癌症，这意味着 SIRT2是一种肿瘤抑制因子。矛盾的是，SIRT2在许多人类癌症中过度表达，包括 此外，我们的数据表明，SIRT2的高表达与不良相关 接受辅助电离辐射(IR)治疗的乳腺癌患者的存活率并有助于抵抗 多种癌症治疗，包括IR、PARP抑制剂和化疗。然而，准确的 SIRT2引导DDR调控肿瘤细胞治疗耐药性的机制，以及如何 SIRT2本身在DDR中受到调控，但人们对此知之甚少。在上一个资助期，我们定义了一部小说 SIRT2通过Atrip和Atrip的乙酰化状态在指导复制应激反应(RSR)中的作用 CDK9，并进一步表明，体细胞SIRT2突变削弱了SIRT2在维持 基因组完整性。在这次续签申请中，我们建议通过调查新角色来进一步开展这项工作 对于SIRT2通过MRE11的脱乙酰基来指导DNA双链断裂(DSB)的修复， 识别控制SIRT2活性的上游泛素化信号事件，并确定SIRT2是否 抑制是治疗对传统疗法耐药的乳腺癌的有效治疗策略。 在这方面，我们已经证明了SIRT2脱乙酰酶活性介导了癌细胞对IR和 通过促进同源重组修复中的末端切除步骤来抑制PARP。我们进一步确定了 一个新的由SIRT2脱乙酰化的DDR蛋白网络响应IR，包括Mre11。而且，我们的 数据表明，在DDR中，SIRT2的活性而不是水平是通过蛋白质单泛素化来调节的。我们 假设SIRT2响应上游泛素化信号以维持基因组完整性和治理 通过关键底物的脱乙酰化引导DDR产生乳腺癌治疗耐药性，包括 Mre11，可能被用来改善乳腺癌的控制。我们建议：1)确定 SIRT2在DDR中指导MRE11；2)剖析SIRT2受 DDR中的单素化；3)将SIRT2确立为乳腺癌耐药的新分子靶点 DNA破坏剂。这项工作的完成将提供对如何 SIRT2指导DSB修复以维持基因组完整性并控制肿瘤细胞治疗耐药性，定义一种 通过DDR中的蛋白质单泛素化来调节SIRT2的新机制，并建立了 抑制SIRT2作为克服乳腺癌耐药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.