Chromatin's Role in Repair of Radiation-induced Damage

染色质在修复辐射引起的损伤中的作用

• 批准号: 6623276
• 负责人: Jessica K Tyler
• 金额: $ 27.17万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-05-01 至 2006-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6623276
• 关键词: DNA damage; DNA repair; DNA replication; chromatin; chromosome aberrations; developmental genetics; endonuclease; enzyme induction /repression; eukaryote; fungal genetics; gene expression; gene targeting; genetic manipulation; genetic mapping; genetic models; histones; protein biosynthesis; protein structure function; radiation genetics; radiation sensitivity; yeasts

The goal of this research is to define the fundamental role that chromatin plays during the repair of radiation-induced double-strand breaks (DSBs). Cell survival and maintenance of genome integrity are critically dependent on the repair of DSBs by the non-homologous end joining (NHEJ) pathway in mammalian cells. If repaired incorrectly, DSBs result in abberations such as chromosomal rearrangements and can lead to formation of cancers. In order to fully understand the repair of radiation- induced DSBs, it is important to consider the natural context - chromatin. The packaging of the genome into chromatin is likely to influence DNA repair processes by analogy to the situation with gene expression. Accordingly, we have discovered a novel chromatin assembly factor, termed ASF1, essential for the repair of radiation-induced, endogenous and developmentally-programmed DNA damage in vivo. Surprisingly, ASF1 is even required for DNA repair processes, such as NHEJ, that do not invoke the assembly of chromatin onto newly-synthesized DNA. We will test the hypothesis that chromatin structure is altered by the ASF1 chromatin assembly factor during NHEJ and that changes to the chromatin structure are essential and intrinsic to the repair of DNA damage. In order to gain insight into the repair of radiation-induced DNA damage, the approach will be to induce a highly specific endonuclease within yeast to generate a synchronous defined DSB that can only be repaired by NHEJ. Using this model system, we will map for the first time the changes to the chromatin structure that precede, accompany and follow the repair of a DSB by NHEJ. Finally, we will determine the generality of the influence of chromatin structure on NHEJ, by examining the role of ASF1 and chromatin structure in mammalian cells. The findings of the proposed experiments using model genetic systems to induce a defined DSB will provide the foundation for understanding the fundamental, yet previously overlooked, role of chromatin structure during the repair of radiation-induced DSBs. As such, these studies are directly applicable to human diseases that result from radiation-induced loss of genome integrity, including many forms of cancer.

这项研究的目的是确定染色质在修复辐射引起的双链断裂 (DSB) 过程中所发挥的基本作用。 细胞存活和基因组完整性的维持严重依赖于哺乳动物细胞中非同源末端连接 (NHEJ) 途径对 DSB 的修复。 如果修复不正确，DSB 会导致染色体重排等畸变，并可能导致癌症的形成。 为了充分了解辐射诱导的 DSB 的修复，重要的是要考虑自然环境 - 染色质。 与基因表达的情况类似，基因组包装到染色质中可能会影响 DNA 修复过程。 因此，我们发现了一种新的染色质组装因子，称为 ASF1，对于体内辐射诱导的内源性和发育程序性 DNA 损伤的修复至关重要。 令人惊讶的是，ASF1 甚至是 DNA 修复过程所必需的，例如 NHEJ，这些过程不会调用染色质组装到新合成的 DNA 上。 我们将检验这样的假设：染色质结构在 NHEJ 过程中被 ASF1 染色质组装因子改变，并且染色质结构的改变对于 DNA 损伤的修复是必不可少的和固有的。 为了深入了解辐射引起的 DNA 损伤的修复，该方法将在酵母内诱导高度特异性的核酸内切酶，以生成只能由 NHEJ 修复的同步定义的 DSB。 使用这个模型系统，我们将首次绘制 NHEJ 修复 DSB 之前、伴随和之后的染色质结构变化。 最后，我们将通过检查 ASF1 和染色质结构在哺乳动物细胞中的作用来确定染色质结构对 NHEJ 影响的普遍性。 使用模型遗传系统诱导确定的 DSB 的拟议实验的结果将为理解染色质结构在辐射诱导的 DSB 修复过程中的基本作用（但之前被忽视）奠定基础。 因此，这些研究可直接适用于由辐射引起的基因组完整性丧失而导致的人类疾病，包括许多形式的癌症。