Engineering of the Translesion DNA Synthesis Pathway Enables Controllable C-to-G and C-to-A Base Editing in Corynebacterium glutamicum.
Engineering of the Translesion DNA Synthesis Pathway Enables Controllable C-to-G and C-to-A Base Editing in Corynebacterium glutamicum.
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DOI：
10.1021/acssynbio.2c00265
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发表时间：
2022-09
影响因子：
4.7
通讯作者：
Yu Wang;Dongdong Zhao;Letian Sun;Jie Wang;Liwen Fan;Guimin Cheng;Zhihui Zhang;Xiaomeng Ni
中科院分区：
文献类型：
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作者：
Yu Wang;Dongdong Zhao;Letian Sun;Jie Wang;Liwen Fan;Guimin Cheng;Zhihui Zhang;Xiaomeng Ni
扩展基本转换类型有望在很大程度上拓宽基本编辑的应用范围，而这需要破译控制编辑结果的机制。在这里，我们发现DNA聚合酶V介导的跨损伤DNA合成(TLS)途径控制着大肠杆菌中糖基编辑者(GBE)对C-to-A的编辑。然而，令人惊讶的是，在谷氨酸棒杆菌中，GBE的主要产物是C-G转换，随后的基因失活确定了决定TLS酶的决定因素。将大肠杆菌TLS途径引入TLS缺失的谷氨酸杆菌突变体中，完全改变了GBE从C到G到C到A的结果。结合规范的C-to-T编辑，在谷氨酸菌中建立了一个开创性的C-to-N碱基编辑工具箱。碱基转换能力的扩大产生了更大的遗传多样性，并促进了碱基编辑在基因失活和蛋白质进化中的应用。这项研究证明了设计TLS系统来开发先进的基因组编辑工具的可能性。
Expanding the base conversion type is expected to largely broaden the application of base editing, whereas it requires decipherment of the machinery controlling the editing outcome. Here, we discovered that the DNA polymerase V-mediated translesion DNA synthesis (TLS) pathway controlled the C-to-A editing by a glycosylase base editor (GBE) in Escherichia coli. However, C-to-G conversion was surprisingly found to be the main product of the GBE in Corynebacterium glutamicum and subsequent gene inactivation identified the decisive TLS enzymes. Introduction of the E. coli TLS pathway into a TLS-deficient C. glutamicum mutant completely changed the GBE outcome from C-to-G to C-to-A. Combining the canonical C-to-T editor, a pioneering C-to-N base editing toolbox was established in C. glutamicum. The expanded base conversion capability produces greater genetic diversity and promotes the application of base editing in gene inactivation and protein evolution. This study demonstrates the possibility of engineering TLS systems to develop advanced genome editing tools.