Transcription-Associated Genome Instability

 描述(由申请人提供) 转录和复制是基本的生理过程，但矛盾的是，它们都对基因组的稳定性构成了威胁。高转录的DNA基因座与重组和突变水平的增加有关，复制叉在每个细胞周期都会遇到DNA损伤和其他类型的复制应激。参与复制和转录的蛋白质复合体之间的物理冲突也威胁到基因组的稳定性。最近的研究表明，在S时期，转录的有害效应可能通过R-环、含有RNADNA杂交体的三链核酸结构和单链DNA区来介导。R-环存在于哺乳动物细胞的基因组中，调节基因表达的各个方面，但在某些情况下，它们的形成与DNA损伤有关。R环如何影响复制和转录，它们是如何被解决的，以及它们最终是如何导致DNA损伤的，这些都是悬而未决的问题。这项研究计划的长期目标是了解细胞如何区分和处理调节和有害的R环，以及这是如何在人类疾病中受到干扰的。人们假设R-环是动态结构，但当转录、RNA加工或DNA复制受到干扰时，它们会积累到有毒的水平。在这些条件下，累积的R-环对复制和转录构成威胁，并导致DNA损伤，最终导致基因组不稳定。这项应用的目标是确定R-环是如何被细胞识别和处理的，以及这些途径中的缺陷如何影响DNA复制、转录和基因组稳定性。在第一个目标中，细胞核酸酶参与了对R环的处理 将探索DNA修复。在第二个目标中，将利用最近开发的一个新系统来研究细胞中复制和转录机制之间的碰撞，以及R环对这些碰撞的影响。最后，在第三个目标中，将探讨R-环处理对转录、复制和基因组稳定性的影响。这些研究将利用先进的基因组方法来绘制基因组中R-环、转录活性和双链断裂的空间分布图。

 DESCRIPTION (provided by applicant) Transcription and replication are fundamental physiological processes, yet paradoxically they both pose a threat to the stability of the genome. Highly transcribed DNA loci are associated with increased levels of recombination and mutagenesis, and replication forks encounter DNA lesions and other types of replication stress every cell cycle. Physical conflicts between the protein complexes involved in replication and transcription also threaten genome stability. Recent studies suggest that the deleterious effects of transcription could be mediated during S phase by R-loops, three-stranded nucleic acid structures containing an RNA-DNA hybrid and region of single-stranded DNA. R-loops occur throughout the genome of mammalian cells and regulate various aspects of gene expression, but in some circumstances their formation is correlated with DNA damage. How R-loops affect replication and transcription, how they are resolved, and how they ultimately lead to DNA damage are unresolved questions. The long-term goal of this research program is to understand how cells distinguish and process regulatory and deleterious R-loops, and how this is perturbed in human disease. It is hypothesized that R-loops are dynamic structures, but that they accumulate to toxic levels when transcription, RNA processing or DNA replication is perturbed. Under these conditions, accumulated R-loops pose threats to replication and transcription and cause DNA damage, ultimately resulting in genome instability. The objective of this application is to define how R-loops are recognized and processed by the cell, and how defects in these pathways affect DNA replication, transcription and genome stability. In the first aim, the processing of R-loops by cellular nucleases involved in DNA repair will be explored. In the second aim, a novel system recently developed will be utilized to study collisions between the replication and transcription machineries in cells, and th impact of an R-loop on these collisions. Finally, in the third aim, the effects of R- loop processing on transcription, replication and genome stability will be probed. These studies will take advantage of cutting-edge genomic approaches to map the spatial distribution of R-loops, transcriptional activity and double-strand breaks in the genome.