Bacteria and archaea employ CRISPR (clustered, regularly, interspaced, short palindromic repeats)-Cas (CRISPR-associated) systems as a type of adaptive immunity to target and degrade foreign nucleic acids. While a myriad of CRISPR-Cas systems have been identified to date, type I-C is one of the most commonly found subtypes in nature. Interestingly, the type I-C system employs a minimal Cascade effector complex, which encodes only three unique subunits in its operon. Here, we present a 3.1 Å resolution cryo-EM structure of the Desulfovibrio vulgaris type I-C Cascade, revealing the molecular mechanisms that underlie RNA-directed complex assembly. We demonstrate how this minimal Cascade utilizes previously overlooked, non-canonical small subunits to stabilize R-loop formation. Furthermore, we describe putative PAM and Cas3 binding sites. These findings provide the structural basis for harnessing the type I-C Cascade as a genome-engineering tool.

细菌和古细菌采用CRISPR（聚类的，规则的，间隔的，短回文重复序列）-Cas（CRISPR相关的）系统作为一种靶向和降解外来核酸的适应性免疫。迄今为止，已经确定了无数的CRISPR-Cas系统，但IC型是自然界中最常见的亚型之一。有趣的是，类型IC系统采用了最小的Cascade效应器复合物，该复合物仅在其操纵子中编码三个唯一的亚基。在这里，我们介绍了寻常脱硫弧菌的3.1Å分辨率冷冻EM结构IC级联反应，揭示了RNA定向复杂装配基础的分子机制。我们演示了这种最小的级联如何利用先前被忽视的非规范小亚基来稳定R环的形成。此外，我们描述了假定的PAM和Cas3结合位点。这些发现为利用IC Cascade类型作为基因组工程工具提供了结构基础。