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

Oana M Enache1, Veronica Rendo2, Mai Abdusamad1, Daniel Lam1, Desiree Davison1, Sangita Pal2, Naomi Currimjee2, Julian Hess1, Sasha Pantel3, Anwesha Nag4, Aaron R Thorner4, John G Doench3, Francisca Vazquez1, Rameen Beroukhim1,2,5, Todd R Golub1,5,6,7, Uri Ben-David8,9

  1. Cancer Program, Broad Institute of Harvard and MIT, Cambridge, MA, USA.
  2. Department of Cancer Biology, Dana-Farber Cancer Institute, Boston, MA, USA.
  3. Genetic Perturbation Platform, Broad Institute of Harvard and MIT, Cambridge, MA, USA.
  4. Center for Cancer Genome Discovery, Dana-Farber Cancer Institute, Boston, MA, USA.
  5. Harvard Medical School, Boston, MA, USA.
  6. Howard Hughes Medical Institute, Chevy Chase, MD, USA.
  7. Department of Pediatric Oncology, Dana-Farber Cancer Institute, Boston, MA, USA.
  8. Cancer Program, Broad Institute of Harvard and MIT, Cambridge, MA, USA. ubendavid@tauex.tau.ac.il.
  9. Department of Human Molecular Genetics & Biochemistry, Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel. ubendavid@tauex.tau.ac.il.

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.

Presented By Veronica Rendo

Genetics & Genome Biology, GenomicsNicholas Liao2020-07 (Jul)