Building Endogenous Gene Connections through RNA Self-Assembly Controlled CRISPR/Cas9 Function
Building Endogenous Gene Connections through RNA Self-Assembly Controlled CRISPR/Cas9 Function
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DOI：
10.1021/jacs.1c09041
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发表时间：
2021-12-01
影响因子：
15
通讯作者：
Xu, Liang
中科院分区：
文献类型：
--
作者：
Lin, Jiao;Wang, Wei-Jia;Xu, Liang
构建能够人工建立内源性基因连接的合成电路对于引入新的细胞行为表型至关重要。鉴于内源基因的多样性，它缺乏一个通用的和易于设计的工具箱来操纵遗传网络。在这里，我们提出了一种类型的自组装诱导的RNA电路，可以直接建立内源基因之间的调节连接。受CRISPR/Cas9复合物中指导RNA的天然组装过程的启发，该设计采用独立的触发RNA链来诱导三元指导RNA组装体的形成，用于CRISPR/Cas9的功能控制。根据这一基本原理，在大肠杆菌中，内源基因的表达调控可以由完全独立的内源小RNA和mRNA来控制。大肠杆菌通过可激活的CRISPR/Cas9功能。此外，E.通过引入新的基因连接，成功地对大肠杆菌进行了编程。此外，该设计的功能也在哺乳动物系统中得到了验证。这种基于自组装的RNA电路具有很大的灵活性和设计简单性，并提供了一种独特的方法来建立内源性基因连接，这为操纵细胞遗传网络铺平了道路。
Construction of synthetic circuits that can artificially establish endogenous gene connections is essential to introduce new phenotypes for cellular behaviors. Given the diversity of endogenous genes, it lacks a general and easy-to-design toolbox to manipulate the genetic network. Here we present a type of self-assembly-induced RNA circuit that can directly build regulatory connections between endogenous genes. Inspired from the natural assembling process of guide RNA in the CRISPR/Cas9 complex, this design employs an independent trigger RNA strand to induce the formation of a ternary guide RNA assembly for functional control of CRISPR/Cas9. With this general principle, expressional regulations of endogenous genes can be controlled by totally independent endogenous small RNAs and mRNAs in E. coli via activatable CRISPR/Cas9 function. Moreover, the cellular phenotype of E. coli is successfully programmed with introduction of new gene connections. In addition, the functionality of this design is also verified in the mammalian system. This self-assembly-based RNA circuit exhibits a great flexibility and simplicity of design and provides a unique approach to build endogenous gene connections, which paves a broad way toward manipulation of cellular genetic networks.