Fork Restart at Replication Barriers and Effects on Genome Stability

项目摘要 基因组的复制是生命所必需的基本过程。在复制 当数百万个DNA碱基对组成基因组时，复制叉遇到了障碍， 必须克服的问题，包括DNA结构，强结合蛋白质，或受损或 交联基。解决或绕过障碍后，复制分叉必须重新启动， 以尽可能准确地避免染色体断裂;发生这种情况的一种机制是 包括通常用于同源重组过程的蛋白质， 是由DNA链侵入引起的最近，人们认识到，这一过程可能是 受核位置的调节，停滞的复制叉重新定位到核周边 进行重新启动。本申请的长期目标是了解 分叉再启动，其通过核定位的调节，以及其在防止基因组不稳定性和 有害的染色体断裂。具体目的是：（1）阐明核孔的作用 定位在叉重新启动和重组的调节，和（2）调查的影响， fork restart on repeat不稳定我们已经表明，扩展的CAG重复序列， 形成DNA结构，特别依赖于叉重新启动，以防止拷贝数的变化， 染色体脆性因此，研究构造形成阶段的再启动问题具有重要意义 重复，因为它们是基因组中突变的热点，它们的扩展可以导致 退行性疾病，如亨廷顿病、强直性肌营养不良和ALS。此外，本发明还提供了一种方法， 结构形成重复序列是脆弱的位点，可以破坏并引起重要DNA的缺失 在癌症发生的早期阶段，复制应激会加剧癌症的发生。 发生在癌细胞中。这项研究的结果将是更好地了解细胞 用于科普复制压力的机制，以及这些机制如何出错， 导致基因突变。这些信息应该提供如何支持或干预的见解 通过这些过程，来防止突变或靶向突变细胞以消除。

PROJECT SUMMARY Replication of the genome is a fundamental process necessary for life. During the replication of the millions of DNA base-pairs that make up a genome, the replication fork encounters barriers that must be overcome, including DNA structures, strongly bound proteins, or damaged or cross-linked bases. After the barrier is resolved or bypassed, the replication fork must restart as accurately as possible to avoid chromosome breakage; one mechanism by which this happens involves proteins that are normally used for a process called homologous recombination, which occurs by DNA strand invasion. Recently, it has been recognized that this process may be regulated by nuclear location, with the stalled replication fork relocating to the nuclear periphery to undergo restart. The long-term objective of this application is to understand the process of fork restart, its regulation by nuclear location, and its role in preventing genome instability and deleterious chromosome breaks. The specific aims are (1) to elucidate the role of nuclear pore localization in fork restart and regulation of recombination, and (2) to investigate the effect of fork restart on repeat instability. We have shown that expanded CAG repeat sequences, which form DNA structures, are particularly reliant on fork restart to prevent copy number changes and chromosome fragility. Thus it is of critical importance to study restart at structure-forming repeats as they are hotspots for mutation in the genome, and their expansion can cause degenerative diseases such as Huntington's disease, myotonic dystrophy, and ALS. In addition, structure-forming repeats are fragile sites that can break and provoke deletion of vital DNA sequences during early steps of cancer initiation, which is exacerbated by replication stress occurring in cancer cells. The outcome of this research will be a better understanding of cellular mechanisms used to cope with replication stress, and how these mechanisms can go wrong to cause genetic mutation. This information should provide insights into how to bolster or interfere with these processes, to either prevent mutation or target mutated cells for elimination.