Structural basis for self-cleavage prevention by tag:anti-tag pairing complementarity in type VI Cas13 CRISPR systems.
Mol Cell
; 81(5): 1100-1115.e5, 2021 03 04.
Article
em En
| MEDLINE
| ID: mdl-33472057
ABSTRACT
Bacteria and archaea apply CRISPR-Cas surveillance complexes to defend against foreign invaders. These invading genetic elements are captured and integrated into the CRISPR array as spacer elements, guiding sequence-specific DNA/RNA targeting and cleavage. Recently, in vivo studies have shown that target RNAs with extended complementarity with repeat sequences flanking the target element (taganti-tag pairing) can dramatically reduce RNA cleavage by the type VI-A Cas13a system. Here, we report the cryo-EM structure of Leptotrichia shahii LshCas13acrRNA in complex with target RNA harboring taganti-tag pairing complementarity, with the observed conformational changes providing a molecular explanation for inactivation of the composite HEPN domain cleavage activity. These structural insights, together with in vitro biochemical and in vivo cell-based assays on key mutants, define the molecular principles underlying Cas13a's capacity to target and discriminate between self and non-self RNA targets. Our studies illuminate approaches to regulate Cas13a's cleavage activity, thereby influencing Cas13a-mediated biotechnological applications.
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MEDLINE
Assunto principal:
Proteínas de Bactérias
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RNA Guia de Cinetoplastídeos
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Leptotrichia
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Endodesoxirribonucleases
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Proteínas Associadas a CRISPR
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Sistemas CRISPR-Cas
Idioma:
En
Ano de publicação:
2021
Tipo de documento:
Article