Mechanism of the S-Phase DNA Damage Checkpoint

描述（由申请人提供）：人类癌症是通过一系列将正常细胞转化为恶性肿瘤的遗传变化而产生的。许多这些变化是由基因组重排和复制过程中的其他错误引起的。为了防止这种复制错误，细胞进化出了DNA损伤检查点，这是一套复杂的DNA质量控制机制。其中的核心是S期DNA损伤检查点，这是一种减缓DNA损伤复制的机制。人类和小鼠的遗传学证据表明，S期DNA损伤检查点对于预防癌症至关重要;具有破坏该检查点的突变的人类患者容易发生各种早发性恶性肿瘤。了解检查点的机制对于了解这些癌症的病因至关重要，并将从根本上影响随后对该检查点的研究方法。检查点有两个分支：一个是调节复制起点的激活，另一个是调节复制叉的进展。检查点的分叉调节分支的机制还不清楚。此外，这两个分支在维持基因组稳定性方面的相对重要性尚不清楚。所提出的实验被设计为i）直接确定起点激发和分叉进展的调节在多大程度上有助于响应于DNA损伤的复制减慢，ii）测试分叉分支用于诱导复制偶联重组的假设，以及iii）测量两个分支对维持基因组稳定性的相对贡献。这些实验将利用裂殖酵母粟酒裂殖酵母作为模型系统。裂变酵母和人类之间检查点的保守性使得裂变酵母成为研究这些重要DNA损伤监测途径的极好模型。裂变酵母强大的遗传和生物化学工具使其能够快速识别关键途径成员并严格测试有关其功能的假设。了解裂变酵母S期DNA损伤检查点将为理解人类检查点如何维持基因组稳定性提供重要框架。这一认识将为人类癌症的治疗和预防带来新的治疗靶点和诊断工具。

DESCRIPTION (provided by applicant): Human cancers arise through a series of genetic changes that transform normal cells into malignant tumors. Many of these changes are caused by genomic rearrangements and other errors during replication. To prevent such replication errors, cells have evolved DNA damage checkpoints, a sophisticated set of DNA quality control mechanisms. Central among them is the S-phase DNA damage checkpoint, a mechanism that slows replication in response to DNA damage. Genetic evidence in humans and mice suggest that the S-phase DNA damage checkpoint is crucial for preventing cancer; human patients with mutations that disrupt this checkpoint are prone to a variety of early-onset malignancies. Understanding the checkpoint's mechanism is essential for understanding the etiology of these cancers, and will fundamentally affect the way subsequent studies of this checkpoint are approached. The checkpoint has two branches: one that regulates the activation of replication origins and one that regulates the progression of replication forks. The mechanism of the fork-regulation branch of the checkpoint is not understood. Furthermore, the relative importance of the two branches in maintaining genomic stability is unknown. The proposed experiments are designed to i) to directly determine the extent to which regulation of origin firing and fork progression contribute to the slowing of replication in response to DNA damage, ii) to test the hypothesis that the fork branch acts to induce replication-coupled recombination and iii) to measure the relative contributions of the two branches to the maintenance of genomic stability. These experiments will take advantage of the fission yeast Schizosaccharomyces pombe as a model system. The conservation of checkpoints between fission yeast and humans makes fission yeast an excellent model for investigating these vital DNA damage surveillance pathways. The powerful genetic and biochemical tools available for fission yeast make it possible to rapidly identify key pathway members and rigorously test hypotheses about their functions. Understanding the fission yeast S-phase DNA damage checkpoint will provide an important framework for understanding how the human checkpoint maintains genomic stability. This understanding will lead to new therapeutic targets and diagnostic tools for the treatment and prevention of human cancer.