Cellular responses to DNA replication stress

项目概要 基因组不稳定性会导致癌症、衰老和其他人类疾病的进展。我们的诚信 基因组在复制时尤其面临风险，因为复制叉经常遇到阻碍 进展，包括 DNA 损伤、难以复制的序列、转录中间体或蛋白质-DNA 复合物。这些遭遇常常导致 DNA 断裂、严重的染色体重排和 非整倍性，这是癌症发生的关键事件。正确修复停滞或崩溃的复制叉 通过基于同源重组 (HR) 的机制在防止复制方面发挥着重要作用 应激引起的基因组不稳定性和 HR 机制组件中的许多突变已被 与癌症倾向有关。在缺乏完整的人力资源机制的情况下，容易出错的机制 例如非同源末端连接（NHEJ）往往会被过度利用，导致广泛的基因组 不稳定。因此，对 DNA 损伤招募 HR 和 NHEJ 因子的调控基础是 基因组生物学和癌症研究的核心兴趣，不仅是为了解释 肿瘤发生，而且还为癌症治疗提供了有希望的途径，正如最近所证明的那样 PARP 抑制剂现已被批准用于治疗具有 BRCA1 突变的卵巢癌患者 欧洲和美国。该提案将研究重组 DNA 调控的新机制 修复和修复途径的选择。中心假设是进化上保守的脚手架 蛋白质 TopBP1 在 HR 介导的修复和 DNA 的控制中发挥着核心作用，但在很大程度上尚未被探索。 修复途径选择。在酵母、人类细胞系和 基因组编辑小鼠，我们将定义 TopBP1 及其酵母直系同源物 Dpb11 的分子机制 控制DNA修复。我们的研究将为关键HR的作用提供无与伦比的分子理解 和 NHEJ 因素相互协调，将揭示信号网络如何整合 DNA 的控制 复制、检查点信号传导和 DNA 修复。拟议的实验将揭示新的修复机制 通路选择和重组修复对于抑制基因组不稳定和癌症至关重要。 产生的结果将对 HR 修复功能障碍引起的肿瘤发生研究产生影响 并应该为治疗提供新的理由。

PROJECT SUMMARY Genomic instabilities drive the progression of cancer, aging, and other human diseases. The integrity of our genome is especially at risk while it is being replicated, as replication forks often encounter obstacles to their progression, including DNA lesions, hard-to-replicate sequences, transcription intermediates, or protein-DNA complexes. These encounters often result in DNA breaks, gross chromosomal rearrangements and aneuploidy, which are key events in cancer initiation. The proper repair of stalled or collapsed replication forks through homologous recombination (HR)-based mechanisms plays a major role in preventing replication stress-induced genomic instabilities and many mutations in components of the HR machinery have been associated with cancer predisposition. In the absence of an intact HR-machinery, error-prone mechanisms such as non-homologous end joining (NHEJ) tend to become hyper-utilized, leading to extensive genomic instability. The regulatory basis of the recruitment of HR and NHEJ factors to DNA lesions is therefore of central interest to genome biology and cancer research, not only for explaining the mechanisms of tumorigenesis, but also for providing promising avenues for cancer therapy, as recently demonstrated for PARP inhibitors that are now approved for treatment of ovarian cancer patients with BRCA1 mutations in Europe and the US. This proposal will investigate a new mechanism for regulation of recombinational DNA repair and repair pathway choice. The central hypothesis is that the evolutionarily conserved scaffolding protein TopBP1 plays a central, yet largely unexplored, role in the control of HR-mediated repair and DNA repair pathway choice. Utilizing biochemical, proteomic and genetic approaches in yeast, human cell lines and genome-edited mice, we will define the molecular mechanism by which TopBP1 and its yeast ortholog Dpb11 control DNA repair. Our studies will provide unparalleled molecular understanding of how the action of key HR and NHEJ factors are coordinated and will reveal how signaling networks integrate the control of DNA replication, checkpoint signaling and DNA repair. Proposed experiments will reveal novel mechanisms of repair pathway choice and recombinational repair that are crucial to suppress genomic instability and cancer. Generated outcomes will have implications in the study of tumorigenesis caused by dysfunctions in HR repair and should provide new rationale for therapy.