Department of Molecular Biosciences, University of Texas at Austin, Austin, TX, 78712-1597, USA.
Interdiscplinary Life Sciences Graduate Programs, University of Texas at Austin, Austin, TX, 78712-1597, USA.
Nat Commun. 2022 May 20;13(1):2829. doi: 10.1038/s41467-022-30402-8.
CRISPR-Cas systems are adaptive immune systems that protect prokaryotes from foreign nucleic acids, such as bacteriophages. Two of the most prevalent CRISPR-Cas systems include type I and type III. Interestingly, the type I-D interference proteins contain characteristic features of both type I and type III systems. Here, we present the structures of type I-D Cascade bound to both a double-stranded (ds)DNA and a single-stranded (ss)RNA target at 2.9 and 3.1 Å, respectively. We show that type I-D Cascade is capable of specifically binding ssRNA and reveal how PAM recognition of dsDNA targets initiates long-range structural rearrangements that likely primes Cas10d for Cas3' binding and subsequent non-target strand DNA cleavage. These structures allow us to model how binding of the anti-CRISPR protein AcrID1 likely blocks target dsDNA binding via competitive inhibition of the DNA substrate engagement with the Cas10d active site. This work elucidates the unique mechanisms used by type I-D Cascade for discrimination of single-stranded and double stranded targets. Thus, our data supports a model for the hybrid nature of this complex with features of type III and type I systems.
CRISPR-Cas 系统是一种适应性免疫系统，可保护原核生物免受外来核酸（如噬菌体）的侵害。两种最常见的 CRISPR-Cas 系统包括 I 型和 III 型。有趣的是，I 型-D 干扰蛋白包含 I 型和 III 型系统的特征。在这里，我们展示了分别与双链 DNA（dsDNA）和单链 RNA（ssRNA）靶标结合的 I 型-D Cascade 的结构，分辨率分别为 2.9 和 3.1Å。我们表明，I 型-D Cascade 能够特异性结合 ssRNA，并揭示了 dsDNA 靶标 PAM 识别如何引发长程结构重排，这可能使 Cas10d 为 Cas3'结合和随后的非靶链 DNA 切割做好准备。这些结构使我们能够模拟抗 CRISPR 蛋白 AcrID1 如何通过竞争性抑制 DNA 底物与 Cas10d 活性位点的结合来阻止靶 dsDNA 结合。这项工作阐明了 I 型-D Cascade 用于区分单链和双链靶标的独特机制。因此，我们的数据支持了该复合物具有 III 型和 I 型系统特征的混合性质的模型。
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