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威胁。高度转录的DNA基因座与重组和诱变水平的增加相关，并且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物之间的物理冲突也威胁着基因组的稳定性。最近的研究表明，转录的有害影响可以介导的R-环，三链核酸结构包含一个RNA-DNA杂交和区域的单链DNA在S期。R环存在于哺乳动物细胞的整个基因组中，并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，它们如何被分解，以及它们最终如何导致DNA损伤是尚未解决的问题。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性和有害的R环，以及这在人类疾病中是如何受到干扰的。据推测，R环是动态结构，但当转录、RNA加工或DNA复制受到干扰时，它们会积累到毒性水平。在这些条件下，积累的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。本申请的目的是确定R环如何被细胞识别和加工，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，通过参与以下的细胞核酸酶来加工R环： 将探索DNA修复。在第二个目标中，最近开发的一种新系统将被用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环加工对转录、复制和基因组稳定性的影响。这些研究将利用尖端的基因组方法来绘制基因组中R环的空间分布、转录活性和双链断裂。

 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.