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CRISPR-Cas9 Circular Permutants as Programmable Scaffolds for Genome Modification.
Oakes, Benjamin L; Fellmann, Christof; Rishi, Harneet; Taylor, Kian L; Ren, Shawn M; Nadler, Dana C; Yokoo, Rayka; Arkin, Adam P; Doudna, Jennifer A; Savage, David F.
Affiliation
  • Oakes BL; Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA; Innovative Genomics Institute, University of California, Berkeley, Berkeley, CA 94720, USA.
  • Fellmann C; Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA; Gladstone Institutes, San Francisco, CA 94158, USA.
  • Rishi H; Graduate Group in Biophysics, University of California, Berkeley, Berkeley, CA 94720, USA.
  • Taylor KL; Innovative Genomics Institute, University of California, Berkeley, Berkeley, CA 94720, USA.
  • Ren SM; Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA.
  • Nadler DC; Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA.
  • Yokoo R; Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA.
  • Arkin AP; Department of Bioengineering, University of California, Berkeley, Berkeley, CA 94720, USA; Environmental Genomics and Systems Biology Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA.
  • Doudna JA; Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA; Innovative Genomics Institute, University of California, Berkeley, Berkeley, CA 94720, USA; Gladstone Institutes, San Francisco, CA 94158, USA; Graduate Group in Biophysics, University of Californi
  • Savage DF; Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA. Electronic address: savage@berkeley.edu.
Cell ; 176(1-2): 254-267.e16, 2019 01 10.
Article in En | MEDLINE | ID: mdl-30633905
ABSTRACT
The ability to engineer natural proteins is pivotal to a future, pragmatic biology. CRISPR proteins have revolutionized genome modification, yet the CRISPR-Cas9 scaffold is not ideal for fusions or activation by cellular triggers. Here, we show that a topological rearrangement of Cas9 using circular permutation provides an advanced platform for RNA-guided genome modification and protection. Through systematic interrogation, we find that protein termini can be positioned adjacent to bound DNA, offering a straightforward mechanism for strategically fusing functional domains. Additionally, circular permutation enabled protease-sensing Cas9s (ProCas9s), a unique class of single-molecule effectors possessing programmable inputs and outputs. ProCas9s can sense a wide range of proteases, and we demonstrate that ProCas9 can orchestrate a cellular response to pathogen-associated protease activity. Together, these results provide a toolkit of safer and more efficient genome-modifying enzymes and molecular recorders for the advancement of precision genome engineering in research, agriculture, and biomedicine.
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Full text: 1 Collection: 01-internacional Database: MEDLINE Main subject: Clustered Regularly Interspaced Short Palindromic Repeats / CRISPR-Cas Systems / Gene Editing Language: En Journal: Cell Year: 2019 Document type: Article Affiliation country: Estados Unidos
Fulltext
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Full text: 1 Collection: 01-internacional Database: MEDLINE Main subject: Clustered Regularly Interspaced Short Palindromic Repeats / CRISPR-Cas Systems / Gene Editing Language: En Journal: Cell Year: 2019 Document type: Article Affiliation country: Estados Unidos