GreenGene, Inc., Seoul, Republic of Korea.
NUS Synthetic Biology for Clinical and Technological Innovation (SynCTI) and Department of Biochemistry, National University of Singapore, Singapore, Singapore.
Nat Plants. 2024 Nov;10(11):1652-1658. doi: 10.1038/s41477-024-01808-7. Epub 2024 Sep 26.
CRISPR-free, protein-only cytosine base editors (CBEs) or adenine base editors, composed of DNA-binding proteins such as zinc finger proteins or transcription activator-like effectors (TALEs) and nucleobase cytosine or adenine deaminases, respectively, enable organellar DNA editing in cultured cells, animals and plants. TALE-linked double-stranded DNA deaminase toxin A (DddA)-derived CBEs (DdCBEs) and TALE-linked adenine deaminases (TALEDs) install C-to-T and A-to-G single-nucleotide conversions, respectively, in mitochondria and chloroplasts. Interestingly, whereas TALEDs exclusively induce A-to-G conversions without C-to-T conversions in mammalian mitochondrial DNA, they often install unwanted C-to-T edits in addition to intended A-to-G edits in plastid DNA. Here we show that uracil DNA glycosylase (UDG)-fused TALEDs (UDG-TALEDs) minimize C-to-T conversions without reducing the A-to-G editing efficiency and install a mutation in the chloroplast psbA gene that encodes a single-amino-acid substitution (S264G), which confers herbicide resistance in the resulting plants.
无 CRISPR 、仅包含蛋白质的胞嘧啶碱基编辑器（CBEs）或腺嘌呤碱基编辑器，由 DNA 结合蛋白（如锌指蛋白或转录激活因子样效应物（TALEs））和核碱基胞嘧啶或腺嘌呤脱氨酶分别组成，可实现在培养细胞、动物和植物中的细胞器 DNA 编辑。TALE 连接的双链 DNA 脱氨酶毒素 A（DddA）衍生的 CBE（DdCBE）和 TALE 连接的腺嘌呤脱氨酶（TALED）分别在线粒体和叶绿体中安装 C 到 T 和 A 到 G 的单核苷酸转换。有趣的是，虽然 TALED 在哺乳动物线粒体 DNA 中仅诱导 A 到 G 的转换而不诱导 C 到 T 的转换，但它们经常在质体 DNA 中除了预期的 A 到 G 编辑之外还安装不需要的 C 到 T 编辑。在这里，我们展示了与尿嘧啶 DNA 糖基化酶（UDG）融合的 TALED（UDG-TALED）在不降低 A 到 G 编辑效率的情况下最小化 C 到 T 转换，并在编码单个氨基酸取代（S264G）的叶绿体 psbA 基因中安装突变，这在产生的植物中赋予了除草剂抗性。
