Abstract

The CRISPR-Cas9 system provides the ability to edit, repress, activate, or mark any gene (or DNA element) by pairing of a programmable single guide RNA (sgRNA) with a complementary sequence on the DNA target. Here we present a new method for small-molecule control of CRISPR-Cas9 function through insertion of RNA aptamers into the sgRNA. We show that CRISPR-Cas9-based gene repression (CRISPRi) can be either activated or deactivated in a dose-dependent fashion over a >10-fold dynamic range in response to two different small-molecule ligands. Since our system acts directly on each target-specific sgRNA, it enables new applications that require differential and opposing temporal control of multiple genes.

Control of CRISPR-Cas9 activity allows for fine-tuning of editing and gene expression. Here the authors use gRNAs modified with RNA aptamers to enable small molecule control in bacterial systems.

Details

Title
Controlling CRISPR-Cas9 with ligand-activated and ligand-deactivated sgRNAs
Author
Kale, Kundert 1   VIAFID ORCID Logo  ; Lucas, James E 2 ; Watters, Kyle E 3   VIAFID ORCID Logo  ; Fellmann Christof 3   VIAFID ORCID Logo  ; Ng, Andrew H 2   VIAFID ORCID Logo  ; Heineike, Benjamin M 4 ; Fitzsimmons, Christina M 5 ; Oakes, Benjamin L 3 ; Qu Jiuxin 6 ; Prasad Neha 6 ; Rosenberg, Oren S 7   VIAFID ORCID Logo  ; Savage, David F 3 ; El-Samad, Hana 8 ; Doudna, Jennifer A 9   VIAFID ORCID Logo  ; Kortemme Tanja 10   VIAFID ORCID Logo 

1  University of California San Francisco, Graduate Group in Biophysics, San Francisco, USA (GRID:grid.266102.1) (ISNI:0000 0001 2297 6811) 
2  University of California San Francisco, UC Berkeley – UCSF Graduate Program in Bioengineering, San Francisco, USA (GRID:grid.266102.1) (ISNI:0000 0001 2297 6811) 
3  University of California Berkeley, Department of Molecular and Cell Biology, Berkeley, USA (GRID:grid.47840.3f) (ISNI:0000 0001 2181 7878) 
4  University of California San Francisco, Bioinformatics Graduate Program, San Francisco, USA (GRID:grid.266102.1) (ISNI:0000 0001 2297 6811) 
5  University of California San Francisco, Chemistry and Chemical Biology Graduate Program, San Francisco, USA (GRID:grid.266102.1) (ISNI:0000 0001 2297 6811); National Institutes of Health, Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, Bethesda, USA (GRID:grid.94365.3d) (ISNI:0000 0001 2297 5165) 
6  University of California San Francisco, Department of Medicine, San Francisco, USA (GRID:grid.266102.1) (ISNI:0000 0001 2297 6811) 
7  University of California San Francisco, Department of Medicine, San Francisco, USA (GRID:grid.266102.1) (ISNI:0000 0001 2297 6811); Chan Zuckerberg Biohub, San Francisco, USA (GRID:grid.266102.1) 
8  Chan Zuckerberg Biohub, San Francisco, USA (GRID:grid.47840.3f); University of California San Francisco, Department of Biochemistry and Biophysics, San Francisco, USA (GRID:grid.266102.1) (ISNI:0000 0001 2297 6811) 
9  University of California Berkeley, Department of Molecular and Cell Biology, Berkeley, USA (GRID:grid.47840.3f) (ISNI:0000 0001 2181 7878); University of California Berkeley, Howard Hughes Medical Institute, Berkeley, USA (GRID:grid.47840.3f) (ISNI:0000 0001 2181 7878) 
10  Chan Zuckerberg Biohub, San Francisco, USA (GRID:grid.47840.3f); University of California San Francisco, Department of Bioengineering and Therapeutic Sciences, San Francisco, USA (GRID:grid.266102.1) (ISNI:0000 0001 2297 6811) 
Publication year
2019
Publication date
2019
Publisher
Nature Publishing Group
e-ISSN
20411723
Source type
Scholarly Journal
Language of publication
English
DOI
https://doi.org/10.1038/s41467-019-09985-2
ProQuest document ID
2222645115
Copyright
© The Author(s) 2019. This work is published under http://creativecommons.org/licenses/by/4.0/ (the “License”). Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License.