Chromatin-mediated mechanisms of genome integrity

摘要 我们的研究集中在阐明的机制，其中局部染色质环境中的双脱氧核糖核酸- 模板化过程包括DNA复制、转录和DNA修复。虽然取得了相当大的进展， 尽管我们对指导体外DNA复制的机制的理解已经取得了很大进展，但我们对 DNA复制的起始位点（起点）是如何在染色体中被选择和调节的。的 我的研究小组开创的基因组方法为研究这些机制提供了新的见解， 通过这种方法，可以确定染色体中局部染色质的状态和结构（核小体和转录因子占有率）， 在包括S.酿酒酵母和果蝇。我们 最近开发了一种新的方法来“足迹"真核基因组-同时揭示基因组- 核小体和较小的DNA结合因子（如起始和转录）对DNA的广泛占据 因素）。与利用一个或两个定义的DNA模板的生化重建实验不同，我们能够 全面观察细胞周期调节的染色质变化的级联反应，发生在每个起源周围 酵母基因组中的复制。我们未来的研究将集中在识别和表征染色质 在解旋酶加载后启动DNA复制所需的介导事件。我们还将研究如何 染色质结构在整个基因组中重新建立，以在传代后保持表观遗传完整性。 DNA复制叉DNA复制也是双链断裂（DSB）的一个潜在来源， 修复，可能导致基因组不稳定。我们处于独特的地位，能够识别和理解 在诱导位点特异性DSB及其随后的同源修复后， 重组或非同源末端连接。最后，与Hartemink实验室（杜克，CS）合作， 我们正在使用同步的酵母菌群，通过细胞周期来开发强大的统计AP， 这将使我们能够模拟细胞周期依赖的基因表达变化，从染色质占用的方法 数据

Abstract Our research is focused on elucidating the mechanisms by which the local chromatin environment influences DNA- templated processes including DNA replication, transcription and DNA repair. While considerable progress has been made in our understanding of the mechanisms that direct DNA replication in vitro, we know very little about how start sites of DNA replication (origins) are selected and regulated in the context of the chromosome. The genomic approaches that my research group have pioneered have provided new insights into the mechanisms by which the local chromatin state and structure (nucleosome and transcription factor occupancy) influences key steps in regulating the DNA replication program in multiple species including S. cerevisiae and Drosophila. We have recently developed a novel approach to `footprint' a eukaryotic genome – simultaneously revealing genome- wide occupancy of DNA for both nucleosomes and smaller DNA binding factors (e.g. initiation and transcription factors). Unlike biochemical reconstitution experiments utilizing one or two defined DNA templates, we are able to comprehensively view the cell cycle regulated cascade of chromatin changes that occur surrounding each origin of replication in the yeast genome. Our future research will focus on identifying and characterizing the chromatin mediated events required for initiation of DNA replication following helicase loading. We will also investigate how chromatin structure is re-established throughout the genome to preserve epigenetic integrity following passage of the DNA replication fork. DNA replication is also a potent source of double-stranded breaks (DSB) which, if not repaired, may lead to genomic instability. We are uniquely positioned to identify and understand the dynamics of chromatin structure following the induction of site-specific DSBs and their subsequent repair by homologous recombination or non-homologous end joining. Finally, in collaboration with the Hartemink laboratory (Duke, CS) we are using synchronous populations of yeast proceeding through the cell cycle to develop robust statistical ap- proaches that will enable us to model cell cycle-dependent changes in gene expression from chromatin occupancy data.