Departments of Pathology, Biochemistry & Molecular Biology, and Molecular Microbiology & Immunology and the Section of Molecular & Computational Biology, Department of Biological Sciences, Norris Comprehensive Cancer Center, University of Southern California Keck School of Medicine, Los Angeles, CA 90033, United States.
Departments of Pathology, Biochemistry & Molecular Biology, and Molecular Microbiology & Immunology and the Section of Molecular & Computational Biology, Department of Biological Sciences, Norris Comprehensive Cancer Center, University of Southern California Keck School of Medicine, Los Angeles, CA 90033, United States.
DNA Repair (Amst). 2021 Oct;106:103193. doi: 10.1016/j.dnarep.2021.103193. Epub 2021 Jul 26.
The nonhomologous DNA end joining pathway is required for repair of most double-strand breaks in the mammalian genome. Here we use a purified biochemical NHEJ system to compare the joining of free DNA with recombinant mononucleosomal and dinucleosomal substrates to investigate ligation and local DNA end resection. We find that the nucleosomal state permits ligation in a manner dependent on the presence of free DNA flanking the nucleosome core particle. Local resection at DNA ends by the Artemis:DNA-PKcs nuclease complex is completely suppressed in all mononucleosome substrates regardless of flanking DNA up to a length of 14 bp. Like mononucleosomes, dinucleosomes lacking flanking free DNA are not joined. Therefore, the nucleosomal state imposes severe constraints on NHEJ nuclease and ligase activities.
非同源末端连接途径是哺乳动物基因组中大多数双链断裂修复所必需的。在这里，我们使用纯化的生化非同源末端连接系统来比较游离 DNA 与重组单核小体和双核小体底物的连接，以研究连接和局部 DNA 末端切除。我们发现核小体状态允许连接，这种连接方式依赖于核小体核心颗粒侧翼游离 DNA 的存在。由 Artemis:DNA-PKcs 核酸酶复合物在所有单核小体底物中进行的 DNA 末端局部切除完全受到抑制，无论侧翼 DNA 的长度可达 14 个碱基对。与单核小体一样，缺乏侧翼游离 DNA 的双核小体也不连接。因此，核小体状态对 NHEJ 核酸酶和连接酶活性施加了严格的限制。
