RNA targeting unleashes indiscriminate nuclease activity of CRISPR–Cas12a2

Bravo JPK, Hallmark T, Naegle B, Beisel CL, Jackson RN, Taylor DW. 2023. RNA targeting unleashes indiscriminate nuclease activity of CRISPR–Cas12a2. Nature. 613(7944), 582–587.

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Author
Bravo, JackISTA ; Hallmark, Thomson; Naegle, Bronson; Beisel, Chase L.; Jackson, Ryan N.; Taylor, David W.
Abstract
Cas12a2 is a CRISPR-associated nuclease that performs RNA-guided, sequence-nonspecific degradation of single-stranded RNA, single-stranded DNA and double-stranded DNA following recognition of a complementary RNA target, culminating in abortive infection<jats:sup>1</jats:sup>. Here we report structures of Cas12a2 in binary, ternary and quaternary complexes to reveal a complete activation pathway. Our structures reveal that Cas12a2 is autoinhibited until binding a cognate RNA target, which exposes the RuvC active site within a large, positively charged cleft. Double-stranded DNA substrates are captured through duplex distortion and local melting, stabilized by pairs of ‘aromatic clamp’ residues that are crucial for double-stranded DNA degradation and in vivo immune system function. Our work provides a structural basis for this mechanism of abortive infection to achieve population-level immunity, which can be leveraged to create rational mutants that degrade a spectrum of collateral substrates.
Publishing Year
Date Published
2023-01-04
Journal Title
Nature
Volume
613
Issue
7944
Page
582-587
ISSN
eISSN
IST-REx-ID

Cite this

Bravo JPK, Hallmark T, Naegle B, Beisel CL, Jackson RN, Taylor DW. RNA targeting unleashes indiscriminate nuclease activity of CRISPR–Cas12a2. Nature. 2023;613(7944):582-587. doi:10.1038/s41586-022-05560-w
Bravo, J. P. K., Hallmark, T., Naegle, B., Beisel, C. L., Jackson, R. N., & Taylor, D. W. (2023). RNA targeting unleashes indiscriminate nuclease activity of CRISPR–Cas12a2. Nature. Springer Nature. https://doi.org/10.1038/s41586-022-05560-w
Bravo, Jack Peter Kelly, Thomson Hallmark, Bronson Naegle, Chase L. Beisel, Ryan N. Jackson, and David W. Taylor. “RNA Targeting Unleashes Indiscriminate Nuclease Activity of CRISPR–Cas12a2.” Nature. Springer Nature, 2023. https://doi.org/10.1038/s41586-022-05560-w.
J. P. K. Bravo, T. Hallmark, B. Naegle, C. L. Beisel, R. N. Jackson, and D. W. Taylor, “RNA targeting unleashes indiscriminate nuclease activity of CRISPR–Cas12a2,” Nature, vol. 613, no. 7944. Springer Nature, pp. 582–587, 2023.
Bravo JPK, Hallmark T, Naegle B, Beisel CL, Jackson RN, Taylor DW. 2023. RNA targeting unleashes indiscriminate nuclease activity of CRISPR–Cas12a2. Nature. 613(7944), 582–587.
Bravo, Jack Peter Kelly, et al. “RNA Targeting Unleashes Indiscriminate Nuclease Activity of CRISPR–Cas12a2.” Nature, vol. 613, no. 7944, Springer Nature, 2023, pp. 582–87, doi:10.1038/s41586-022-05560-w.
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