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Published on: September 8, 2012
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A scoutRNA Is Required for Some Type V CRISPR-Cas Systems.
Lucas B Harrington1, Enbo Ma1, Janice S Chen1
1Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA.
Molecular Cell
|July 10, 2020
Summary
CRISPR-Cas12c/d systems require a novel RNA cofactor, scoutRNA, for DNA cutting and pre-crRNA maturation. This discovery expands our understanding of bacterial adaptive immunity and genome editing tools.
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- CRISPR-Cas12c/d proteins are distinct from widely used Cas12a and Cas9 nucleases.
- Genome editing and DNA detection activities of Cas12c/d have not been previously demonstrated.
Purpose of the Study:
- To investigate the DNA cleavage and genome editing capabilities of CRISPR-Cas12c/d systems.
- To identify and characterize the RNA components essential for Cas12c/d function.
Main Methods:
- Biochemical assays to assess DNA cleavage activity.
- Cell-based experiments for genome editing and pre-crRNA maturation analysis.
- RNA structure and sequence analysis of the novel cofactor.
Main Results:
- Cas12d-catalyzed DNA cutting requires both CRISPR RNA (crRNA) and a short-complementarity untranslated RNA (scoutRNA).
- scoutRNA possesses a unique secondary structure and a conserved essential sequence, distinct from known tracrRNAs.
- scoutRNA acts as an essential cofactor for Cas12c-catalyzed pre-crRNA maturation and crRNA-directed DNA recognition.
- Cas12c/d systems do not require host factors like ribonuclease III.
Conclusions:
- scoutRNA represents a third class of CRISPR-associated transcripts.
- This finding elucidates the mechanism of Cas12c/d systems in bacterial adaptive immunity.
- The unique requirements of Cas12c/d systems offer new avenues for genome editing and DNA detection technologies.
Keywords:
CRISPR-casCandidate Phyla Radiation (CPR) bacteriaCas12c (C2c3)Cas12d (CasY)RuvC nuclease domaincrRNAscoutRNAtracrRNAMore Related Videos
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