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RNA targeting unleashes indiscriminate nuclease activity of CRISPR-Cas12a2.

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 infection¹. 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.

SUBMITTER: Bravo JPK

PROVIDER: S-EPMC9849127 | biostudies-literature | 2023 Jan

REPOSITORIES: biostudies-literature

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RNA targeting unleashes indiscriminate nuclease activity of CRISPR-Cas12a2.

Bravo Jack P K JPK Hallmark Thomson T Naegle Bronson B Beisel Chase L CL Jackson Ryan N RN Taylor David W DW

Nature 20230104 7944

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<sup>1</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 ...[more]

PMID: 36599980

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