利用 CRISPR 技术绘制 DNA 双链断裂的细胞反应图谱。
Mapping cellular responses to DNA double-strand breaks using CRISPR technologies.
机构信息
Department of Biophysics and Biophysical Chemistry, Johns Hopkins University, Baltimore, MD, USA.
Department of Biophysics and Biophysical Chemistry, Johns Hopkins University, Baltimore, MD, USA; Department of Biomedical Engineering, Johns Hopkins University, Baltimore, MD, USA; Department of Biophysics, Johns Hopkins University, Baltimore, MD, USA; Howard Hughes Medical Institute, Baltimore, MD, USA.
出版信息
Trends Genet. 2023 Jul;39(7):560-574. doi: 10.1016/j.tig.2023.02.015. Epub 2023 Mar 25.
DNA double-strand breaks (DSBs) are one of the most genotoxic DNA lesions, driving a range of pathological defects from cancers to immunodeficiencies. To combat genomic instability caused by DSBs, evolution has outfitted cells with an intricate protein network dedicated to the rapid and accurate repair of these lesions. Pioneering studies have identified and characterized many crucial repair factors in this network, while the advent of genome manipulation tools like clustered regularly interspersed short palindromic repeats (CRISPR)-CRISPR-associated protein 9 (Cas9) has reinvigorated interest in DSB repair mechanisms. This review surveys the latest methodological advances and biological insights gained by utilizing Cas9 as a precise 'damage inducer' for the study of DSB repair. We highlight rapidly inducible Cas9 systems that enable synchronized and efficient break induction. When combined with sequencing and genome-specific imaging approaches, inducible Cas9 systems greatly expand our capability to spatiotemporally characterize cellular responses to DSB at specific genomic coordinates, providing mechanistic insights that were previously unobtainable.
DNA 双链断裂 (DSB) 是最具遗传毒性的 DNA 损伤之一，可导致从癌症到免疫缺陷等多种病理缺陷。为了应对 DSB 引起的基因组不稳定性，进化赋予了细胞一个复杂的蛋白质网络，专门用于快速准确地修复这些损伤。开创性的研究已经鉴定和描述了该网络中的许多关键修复因子，而诸如成簇规律间隔短回文重复 (CRISPR)-CRISPR 相关蛋白 9 (Cas9) 等基因组操作工具的出现，重新激发了人们对 DSB 修复机制的兴趣。这篇综述调查了利用 Cas9 作为 DSB 修复研究的精确“损伤诱导物”所获得的最新方法学进展和生物学见解。我们强调了可快速诱导的 Cas9 系统，该系统可实现同步和高效的断裂诱导。当与测序和基因组特异性成像方法结合使用时，诱导型 Cas9 系统极大地扩展了我们在特定基因组坐标处对 DSB 诱导的细胞反应进行时空特征描述的能力，提供了以前无法获得的机制见解。
Zotero 插件