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The CRISPR-associated DNA-cleaving enzyme Cpf1 also processes precursor CRISPR RNA

Fonfara, Ines, Richter, Hagen, Bratovič, Majda, Le Rhun, Anaïs, Charpentier, Emmanuelle·2016·Nature
DOI 10.1038/nature17945
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Abstract

CRISPR-Cas systems that provide defence against mobile genetic elements in bacteria and archaea have evolved a variety of mechanisms to target and cleave RNA or DNA. The well-studied types I, II and III utilize a set of distinct CRISPR-associated (Cas) proteins for production of mature CRISPR RNAs (crRNAs) and interference with invading nucleic acids. In types I and III, Cas6 or Cas5d cleaves precursor crRNA (pre-crRNA) and the mature crRNAs then guide a complex of Cas proteins (Cascade-Cas3, type I; Csm or Cmr, type III) to target and cleave invading DNA or RNA. In type II systems, RNase III cleaves pre-crRNA base-paired with trans-activating crRNA (tracrRNA) in the presence of Cas9 (refs 13, 14). The mature tracrRNA-crRNA duplex then guides Cas9 to cleave target DNA. Here, we demonstrate a novel mechanism in CRISPR-Cas immunity. We show that type V-A Cpf1 from Francisella novicida is a dual-nuclease that is specific to crRNA biogenesis and target DNA interference. Cpf1 cleaves pre-crRNA upstream of a hairpin structure formed within the CRISPR repeats and thereby generates intermediate crRNAs that are processed further, leading to mature crRNAs. After recognition of a 5'-YTN-3' protospacer adjacent motif on the non-target DNA strand and subsequent probing for an eight-nucleotide seed sequence, Cpf1, guided by the single mature repeat-spacer crRNA, introduces double-stranded breaks in the target DNA to generate a 5' overhang. The RNase and DNase activities of Cpf1 require seq…

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Metadata

Authors
Fonfara, I., Richter, H., Bratovič, M., Le Rhun, A., & Charpentier, E.
Venue
Nature
Publisher
Springer Science and Business Media LLC
Published
2016-04
Volume
532
Issue
7600
Pages
517-521
ISSN
  • 0028-0836
  • 1476-4687
DOI
10.1038/nature17945
Cited by
988
References
5
Language
en
Licence
www.springer.com/tdm (tdm)

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