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The CRISPR-associated DNA-cleaving enzyme Cpf1 also processes precursor CRISPR RNA
Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Helmholtz Centre for Infection Research, Department of Regulation in Infection Biology, Braunschweig 38124, Germany; 3Max Planck Institute for Infection Biology, Department of Regulation in Infection Biology, Berlin 10117, Germany.
Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Helmholtz Centre for Infection Research, Department of Regulation in Infection Biology, Braunschweig 38124, Germany; 3Max Planck Institute for Infection Biology, Department of Regulation in Infection Biology, Berlin 10117, Germany.
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2016 (engelsk)Inngår i: Nature, ISSN 0028-0836, E-ISSN 1476-4687, Vol. 532, nr 7600, s. 517-520Artikkel i tidsskrift (Fagfellevurdert) Published
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Abstract [en]

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(1). 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)(2-5) 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(6-12). 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 DNA15. 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(16). The RNase and DNase activities of Cpf1 require sequence- and structure-specific binding to the hairpin of crRNA repeats. Cpf1 uses distinct active domains for both nuclease reactions and cleaves nucleic acids in the presence of magnesium or calcium. This study uncovers a new family of enzymes with specific dual endoribonuclease and endonuclease activities, and demonstrates that type V- A constitutes the most minimalistic of the CRISPR- Cas systems so far described.

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2016. Vol. 532, nr 7600, s. 517-520
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URN: urn:nbn:se:umu:diva-120803DOI: 10.1038/nature17945ISI: 000374815900051OAI: oai:DiVA.org:umu-120803DiVA, id: diva2:930906
Tilgjengelig fra: 2016-05-25 Laget: 2016-05-23 Sist oppdatert: 2018-06-07bibliografisk kontrollert

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