Novel therapeutic means against Staphylococcus aureus infections are urgently needed due to the emergence of drug-resistant S. aureus. We report the development of a CRISPR RNA-guided cytidine deaminase (pnCasSA–BEC), enabling highly efficient gene inactivation and point mutations in S. aureus. We engineered a fusion of a Cas9 nickase (Cas9D10A) and a cytidine deaminase (APOBEC1) that can be guided to a target genomic locus for gene inactivation via generating a premature stop codon. The pnCasSA–BEC system nicks the non-edited strand of the genomic DNA, directly catalyzes the conversion of cytidine (C) to uridine (U), and relies on DNA replication to achieve C → T (G → A) conversion without using donor repair templates. The development of the base-editing system will dramatically accelerate drug-target exploration in S. aureus and provides critical insights into the development of base-editing tools in other microbes.

中文翻译：

---

使用工程CRISPR RNA引导的胞苷脱氨酶 对金黄色葡萄球菌进行高效碱基编辑†

由于耐药性金黄色葡萄球菌的出现，迫切需要针对金黄色葡萄球菌感染的新型治疗手段。我们报道了一种由CRISPR RNA引导的胞苷脱氨酶（pnCasSA–BEC）的开发，该酶可在金黄色葡萄球菌中实现高效的基因失活和点突变。我们设计了Cas9切口酶（Cas9D10A）和胞苷脱氨酶（APOBEC1）的融合体，可以将其引导至目标基因组位点，以通过产生过早的终止密码子。pnCasSA–BEC系统切割基因组DNA的未编辑链，直接催化胞苷（C）转化为尿苷（U），并依靠DNA复制来实现C→T（G→A）转化，而无需使用供体维修模板。基础编辑系统的开发将大大加快金黄色葡萄球菌药物靶标的探索，并为其他微生物基础编辑工具的开发提供重要见解。