Investigating contributions of BRCA2 DNA-binding domains toward maintaining genome integrity

摘要 存在着消除DNA损伤的保守机制，如果不修复，可能导致基因突变， 基因组重排以触发肿瘤细胞转化和肿瘤发生。特别是同源的 重组（HR）代表了用于修复DNA双链断裂（DSB）、链间断裂和链间断裂的高保真工具。 交联和折叠的DNA复制叉。HR的正确执行需要肿瘤抑制因子BRCA2 （乳腺癌2）。BRCA2的生殖系突变会导致乳腺癌和卵巢癌， 范可尼综合征贫血此外，体细胞驱动BRCA2突变在多种癌症类型中发现。 需要进一步的工作来阐明BRCA2在HR修复中的机制作用。重要的是，研究 本文提出的将有助于确定两个不同的DNA结合结构域对功能的贡献， BRCA2作为“重组介体”，特别是促进RAD51复合物的组装 重组酶作用于来源于DSB加工的单链DNA。我们的核心假设是， BRCA2内的两个DNA结合结构域在指导BRCA2依赖性DSB修复中起重要作用， 通过单链DNA和DNA结构的参与， 已处理的DSB。我们将在两个特定的条件下进行各种生化、遗传和细胞生物学研究。 旨在检验这一核心原则。在这些实验中，我们将描述基因突变是如何在 每个DNA结合域影响HR和复制叉保存的关键方面。结果是 奖学金项目不仅将扩大我们对BRCA2 DNA结合结构域如何有助于 基因组稳定性的维持，但他们也将阐明这些结构域中的突变是如何发生的。 最终导致恶性肿瘤。此外，我们希望我们的研究结果有助于确定新的目标和途径支点 新的癌症治疗方法的发展点。

ABSTRACT Conserved mechanisms exist to eliminate DNA damage, which, if left unrepaired, can cause gene mutations and genome rearrangements to trigger neoplastic cell transformation and oncogenesis. In particular, homologous recombination (HR) represents a high-fidelity tool for the repair of DNA double-strand breaks (DSBs), interstrand crosslinks, and collapsed DNA replication forks. Proper execution of HR requires the tumor suppressor BRCA2 (Breast Cancer 2). Germline mutations in BRCA2 lead to breast and ovarian cancer and also the cancer-prone syndrome Fanconi anemia. Moreover, somatic driver BRCA2 mutations are found in a variety of cancer types. Further work is necessary to elucidate the mechanistic role of BRCA2 in HR repair. Importantly, the research proposed herein will help define the contributions of two distinct DNA binding domains toward the function of BRCA2 as a “recombination mediator,” specifically in facilitating the assembly of complexes of the RAD51 recombinase on single-stranded DNA derived from the processing of DSBs. Our central hypothesis posits that the two DNA-binding domains within BRCA2 play an essential role in guiding BRCA2-dependent DSB repair and replication fork preservation through the engagement of single-stranded DNA and DNA structures present in processed DSBs. We will conduct a variety of biochemical, genetic, and cell biological studies under two specific aims to test this central tenet. Throughout these experiments, we will delineate how genetic mutations within each DNA binding domain affect key aspects of HR and replication fork preservation. The results from this fellowship project will not only expand our understanding of how the BRCA2 DNA-binding domains contribute to the maintenance of genome stability, but they will also shed light on how mutations within these domains ultimately cause malignancy. Furthermore, we expect our findings to help identify new targets and pathway pivot points for the development of novel cancer therapeutics.