Discovery of potent and versatile CRISPR-Cas9 inhibitors engineered for chemically controllable genome editing.
Discovery of potent and versatile CRISPR-Cas9 inhibitors engineered for chemically controllable genome editing.
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DOI：
10.1093/nar/gkac099
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发表时间：
2022-03-21
影响因子：
14.9
通讯作者：
Tian Y
中科院分区：
文献类型：
--
作者：
Song G;Zhang F;Tian C;Gao X;Zhu X;Fan D;Tian Y
抗CRISPR(ACR)蛋白由噬菌体、质粒等多种移动遗传元件编码，可对抗原核生物利用的CRISPR-Cas适应性免疫系统，为CRISPR-Cas的应用提供了强有力的工具。在这里，我们从链球菌MGES中发现了9个不同的II-A抗CRISPR(AcrIIA24-32)家族，并发现大多数ACRs在细菌和人类细胞中都能有效地抑制链球菌的II-A Cas9同源物(SpyCas9、St1Cas9或St3Cas9)。其中，AcrIIA26、AcrIIA27、AcrIIA30和AcrIIA31能阻断Cas9与DNA的结合，而AcrIIA24能阻断Cas9对DNA的切割。值得注意的是，AcrIIA25.1和AcrIIA32.1能够抑制SpyCas9的DNA结合和DNA切割活性，表现出独特的抗CRISPR特性。重要的是，我们开发了几种基于AcrIIA25.1和AcrIIA32.1的化学诱导的抗CRISPR变体，通过包含ACR蛋白和4-羟基他莫昔芬响应的内含素的杂交，这使得能够对CRISPR-Cas9介导的人类细胞基因组编辑进行翻译后控制。综上所述，我们的工作扩大了II-A型抗CRISPR家族和ACR蛋白工具箱的多样性，用于化学诱导控制基于Cas9的应用。
Anti-CRISPR (Acr) proteins are encoded by many mobile genetic elements (MGEs) such as phages and plasmids to combat CRISPR–Cas adaptive immune systems employed by prokaryotes, which provide powerful tools for CRISPR–Cas-based applications. Here, we discovered nine distinct type II-A anti-CRISPR (AcrIIA24–32) families from Streptococcus MGEs and found that most Acrs can potently inhibit type II-A Cas9 orthologs from Streptococcus (SpyCas9, St1Cas9 or St3Cas9) in bacterial and human cells. Among these Acrs, AcrIIA26, AcrIIA27, AcrIIA30 and AcrIIA31 are able to block Cas9 binding to DNA, while AcrIIA24 abrogates DNA cleavage by Cas9. Notably, AcrIIA25.1 and AcrIIA32.1 can inhibit both DNA binding and DNA cleavage activities of SpyCas9, exhibiting unique anti-CRISPR characteristics. Importantly, we developed several chemically inducible anti-CRISPR variants based on AcrIIA25.1 and AcrIIA32.1 by comprising hybrids of Acr protein and the 4-hydroxytamoxifen-responsive intein, which enabled post-translational control of CRISPR–Cas9-mediated genome editing in human cells. Taken together, our work expands the diversity of type II-A anti-CRISPR families and the toolbox of Acr proteins for the chemically inducible control of Cas9-based applications.
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影响因子：
14.9
作者：
Birkholz, Nils;Fagerlund, Robert D.;Fineran, Peter C.
通讯作者：
Fineran, Peter C.
影响因子：
56.9
作者：
Jinek, Martin;Chylinski, Krzysztof;Charpentier, Emmanuelle
通讯作者：
Charpentier, Emmanuelle
影响因子：
16
作者：
Jiang, Fuguo;Liu, Jun-Jie;Doudna, Jennifer A.
通讯作者：
Doudna, Jennifer A.
通过反式编码的小 RNA 和宿主因子 RNase III 进行 CRISPR RNA 成熟。
影响因子：
64.8
作者：
Deltcheva, Elitza;Chylinski, Krzysztof;Sharma, Cynthia M.;Gonzales, Karine;Chao, Yanjie;Pirzada, Zaid A.;Eckert, Maria R.;Vogel, Joerg;Charpentier, Emmanuelle
通讯作者：
Charpentier, Emmanuelle
影响因子：
14.8
作者：
Davis, Kevin M.;Pattanayak, Vikram;Thompson, David B.;Zuris, John A.;Liu, David R.
通讯作者：
Liu, David R.