Journal Article

Structural basis for assembly of non-canonical small subunits into type I-C Cascade

Schwartz, Evan A., Wrapp, Daniel, O’Brien, Roisin E., Santos, Inês C., Brodbelt, Jennifer S., Bravo, Jack P. K., Taylor, David W.

2020

Overview

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. Type I-C Cascade (the CRISPR-associated complex for antiviral defense) is a minimal system, comprising only three unique Cas proteins. Cryo-EM structure of the Desulfovibrio vulgaris type I-C Cascade reveals the molecular mechanisms that underlie RNA-directed complex assembly.

• Publisher: Nature Publishing Group UK,Nature Publishing Group,Nature Portfolio
• Subject: 101/28 / 631/337/4041 / 631/45/173 / 631/45/500 / 631/45/535/1258/1259 / Adaptive immunity / Adaptive systems / Antiviral drugs / Archaea / Assembly / Bacterial Proteins - chemistry / Bacterial Proteins - metabolism / Binding Sites / Clustered Regularly Interspaced Short Palindromic Repeats - genetics / CRISPR / CRISPR-Associated Proteins - chemistry / CRISPR-Associated Proteins - metabolism / CRISPR-Cas Systems / Cryoelectron Microscopy / Desulfovibrio vulgaris / Desulfovibrio vulgaris - chemistry / Desulfovibrio vulgaris - genetics / DNA - chemistry / DNA - metabolism / Genome editing / Genomes / Humanities and Social Sciences / Models, Molecular / Molecular modelling / multidisciplinary / Multiprotein Complexes - chemistry / Multiprotein Complexes - metabolism / Nucleic acids / Nucleotide Motifs / Protein Conformation / Protein Subunits - chemistry / Protein Subunits - metabolism / Ribonucleic acid / RNA / RNA, Bacterial - chemistry / RNA, Bacterial - metabolism / Science / Science (multidisciplinary)