Abstract

CRISPR-based genetic screening has revolutionized cancer drug target discovery, yet reliable, multiplex gene editing to reveal synergies between gene targets remains a major challenge. Here, we present a simple and robust CRISPR-Cas12a-based approach for combinatorial genetic screening in cancer cells. By engineering the CRISPR-AsCas12a system with key modifications to the Cas protein and its CRISPR RNA (crRNA), we can achieve high efficiency combinatorial genetic screening. We demonstrate the performance of our optimized AsCas12a (opAsCas12a) through double knockout screening against epigenetic regulators. This screen reveals synthetic sick interactions between Brd9&Jmjd6, Kat6a&Jmjd6, and Brpf1&Jmjd6 in leukemia cells.

Reliable, multiplexed gene editing to uncover synergies between targets remains challenging. Here, the authors engineer AsCas12a and the crRNA to improve double knockout for synthetic sick/lethal interaction genetic screening.

Details

Title
High-performance CRISPR-Cas12a genome editing for combinatorial genetic screening
Author
Gier, Rodrigo A 1   VIAFID ORCID Logo  ; Budinich, Krista A 2 ; Evitt, Niklaus H 3   VIAFID ORCID Logo  ; Cao Zhendong 2 ; Freilich, Elizabeth S 2 ; Chen Qingzhou 2 ; Qi, Jun 4 ; Yemin, Lan 5 ; Kohli, Rahul M 6   VIAFID ORCID Logo  ; Shi Junwei 2   VIAFID ORCID Logo 

1  University of Pennsylvania, Department of Cancer Biology, Perelman School of Medicine, Philadelphia, USA (GRID:grid.25879.31) (ISNI:0000 0004 1936 8972); University of Pennsylvania, Epigenetics Institute, Department of Cell and Developmental Biology, Perelman School of Medicine, Philadelphia, USA (GRID:grid.25879.31) (ISNI:0000 0004 1936 8972); University of Pennsylvania, Abramson Family Cancer Research Institute, Perelman School of Medicine, Philadelphia, USA (GRID:grid.25879.31) (ISNI:0000 0004 1936 8972); University of Pennsylvania, Department of Bioengineering, Philadelphia, USA (GRID:grid.25879.31) (ISNI:0000 0004 1936 8972) 
2  University of Pennsylvania, Department of Cancer Biology, Perelman School of Medicine, Philadelphia, USA (GRID:grid.25879.31) (ISNI:0000 0004 1936 8972); University of Pennsylvania, Epigenetics Institute, Department of Cell and Developmental Biology, Perelman School of Medicine, Philadelphia, USA (GRID:grid.25879.31) (ISNI:0000 0004 1936 8972); University of Pennsylvania, Abramson Family Cancer Research Institute, Perelman School of Medicine, Philadelphia, USA (GRID:grid.25879.31) (ISNI:0000 0004 1936 8972) 
3  University of Pennsylvania, Department of Cancer Biology, Perelman School of Medicine, Philadelphia, USA (GRID:grid.25879.31) (ISNI:0000 0004 1936 8972); University of Pennsylvania, Epigenetics Institute, Department of Cell and Developmental Biology, Perelman School of Medicine, Philadelphia, USA (GRID:grid.25879.31) (ISNI:0000 0004 1936 8972); University of Pennsylvania, Abramson Family Cancer Research Institute, Perelman School of Medicine, Philadelphia, USA (GRID:grid.25879.31) (ISNI:0000 0004 1936 8972); University of Pennsylvania, Department of Medicine, Perelman School of Medicine, Philadelphia, USA (GRID:grid.25879.31) (ISNI:0000 0004 1936 8972) 
4  Harvard Medical School, Department of Cancer Biology, Dana-Farber Cancer Institute, Department of Medicine, Boston, USA (GRID:grid.38142.3c) (ISNI:000000041936754X) 
5  University of Pennsylvania, Epigenetics Institute, Department of Cell and Developmental Biology, Perelman School of Medicine, Philadelphia, USA (GRID:grid.25879.31) (ISNI:0000 0004 1936 8972) 
6  University of Pennsylvania, Epigenetics Institute, Department of Cell and Developmental Biology, Perelman School of Medicine, Philadelphia, USA (GRID:grid.25879.31) (ISNI:0000 0004 1936 8972); University of Pennsylvania, Department of Medicine, Perelman School of Medicine, Philadelphia, USA (GRID:grid.25879.31) (ISNI:0000 0004 1936 8972) 
Publication year
2020
Publication date
2020
Publisher
Nature Publishing Group
e-ISSN
20411723
Source type
Scholarly Journal
Language of publication
English
DOI
https://doi.org/10.1038/s41467-020-17209-1
ProQuest document ID
2423335421
Copyright
© The Author(s) 2020. 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.