Cas9 activates the p53 pathway and selects for p53-inactivating mutations

Oana M. Enache, Veronica Rendo, Mai Abdusamad, Daniel Lam, Desiree Davison, Sangita Pal, Naomi Currimjee, Julian Hess, Sasha Pantel, Anwesha Nag, Aaron R. Thorner, John G. Doench, Francisca Vazquez, Rameen Beroukhim, Todd R. Golub, Uri Ben-David*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

Abstract

Cas9 is commonly introduced into cell lines to enable CRISPR–Cas9-mediated genome editing. Here, we studied the genetic and transcriptional consequences of Cas9 expression itself. Gene expression profiling of 165 pairs of human cancer cell lines and their Cas9-expressing derivatives revealed upregulation of the p53 pathway upon introduction of Cas9, specifically in wild-type TP53 (TP53-WT) cell lines. This was confirmed at the messenger RNA and protein levels. Moreover, elevated levels of DNA repair were observed in Cas9-expressing cell lines. Genetic characterization of 42 cell line pairs showed that introduction of Cas9 can lead to the emergence and expansion of p53-inactivating mutations. This was confirmed by competition experiments in isogenic TP53-WT and TP53-null (TP53−/−) cell lines. Lastly, Cas9 was less active in TP53-WT than in TP53-mutant cell lines, and Cas9-induced p53 pathway activation affected cellular sensitivity to both genetic and chemical perturbations. These findings may have broad implications for the proper use of CRISPR–Cas9-mediated genome editing.

Original languageEnglish
Pages (from-to)662-668
Number of pages7
JournalNature Genetics
Volume52
Issue number7
DOIs
StatePublished - 1 Jul 2020

Funding

FundersFunder number
Gray Matters Brain Cancer Foundation
Richard Eimert Research Fund on Solid Tumors
Tel-Aviv University
National Institutes of HealthR01 CA18828, CA215489, CA219943
Howard Hughes Medical InstituteInvestigator grant
National Institute on Drug AbuseR01DA018828
Pediatric Brain Tumor Foundation
Novartis
Horowitz Foundation for Social Policy
Israel Cancer Association
Azrieli Foundation

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