DNA double-strand break chromatin alterations and genome integrity

联系PD/PI：Greenberg，Roger A. 摘要 进化保守的DNA修复机制在整个细胞周期中协同工作，忠实地维持 基因组完整性。然而，对于受损的染色质如何引导一系列 决定后续修复机制的事件。这种知识上的差距与一个糟糕的 已知的同源定向修复机制变得活跃，以维持端粒长度 大约15%的人类癌症。另一种端粒延长(ALT)发生在非规范的 间充质来源癌症中同源基因引导的DNA修复机制。重组机制 负责端粒维持的基因在ALT依赖型癌症中具有独特的脆弱性。 我们已经开发了实验系统，能够定量、实时地可视化 同源基因定向修复受损的端粒。我们已发表的研究表明，ALT是由dna启动的。 损伤依赖的同源定向修复合成，单向进行到 染色体。我们将这种机制命名为Break诱导的端粒合成，并表明它对 利用丙氨酸氨基转移酶的细胞端粒延长。在这里，我们研究了DNA损伤如何反应的问题 组装在端粒染色质上，指导长链同源定向修复端粒合成。 我们假设，Break诱导的端粒合成是通过组装几个不同的元件而发生的 DNA修复机制在端粒染色质上实现同源定向修复合成。我们未出版的 结果明确支持这一前提，显示了广泛的增殖细胞核抗原泛素化和5‘-3’的招募 受损端粒上的核酸酶SNM1A和5‘-3’解旋酶FANCJ促进DNA修复合成和 端粒完整性。我们将研究它们与端粒和端粒损伤相关性的基础。 使用多方面方法在端粒重组依赖修复合成中的功能 US在细胞和体外高纯度系统中检查端粒损伤反应的每一步 在确定的底物上组装断裂诱导的复制体。这些基础研究旨在 了解Break诱导端粒合成的关键要素以及它如何使端粒延长 用于依赖ALT的细胞的持续增殖。

Contact PD/PI: Greenberg, Roger A. Summary Evolutionary conserved DNA repair mechanisms cooperate throughout the cell cycle to faithfully maintain genome integrity. However, a minimal understanding exists for how damaged chromatin directs the series of events that dictate ensuing repair mechanism. This gap in knowledge is particularly relevant to a poorly understood homology directed repair mechanism that becomes active to maintain telomere length in approximately 15% of human cancers. Alternative lengthening of telomeres (ALT) occurs by noncanonical homology directed DNA repair mechanism in cancers of mesenchymal origin. The recombination mechanisms responsible for telomere maintenance represent a unique vulnerability in ALT reliant cancers. We have developed experimental systems that enable quantitative, real-time visualization of each step in the homology directed repair of damaged telomeres. Our published studies reveal that ALT is initiated by DNA damage dependent homology directed repair synthesis that proceeds unidirectionally to the end of the chromosome. We named this mechanism Break Induced Telomere Synthesis and showed that it is critical for telomere lengthening in cells that utilize ALT. Here, we examine the question of how DNA damage responses assemble on telomeric chromatin to direct long-tract homology directed repair telomere synthesis. We hypothesize that Break Induced Telomere Synthesis occurs by assembling elements of several different DNA repair mechanisms on telomeric chromatin to achieve homology directed repair synthesis. Our unpublished results definitively support this premise, showing extensive PCNA- ubiquitination and recruitment of the 5’-3’ nuclease SNM1A and 5’-3’ helicase FANCJ at damaged telomeres to promote DNA repair synthesis and telomere integrity. We will investigate the basis for their damage dependent association with telomeres and functions in recombination dependent repair synthesis at telomeres using a multifaceted approach that allows us to examine each step of the telomere damage response in cells and in vitro in highly purified systems that assemble the break induced replisome on defined substrates. These fundamental studies are designed to understand the critical elements of Break Induced Telomere Synthesis and how it allows telomere lengthening for sustained proliferation in cells that rely on ALT.