CRISPR-induced on-target large deletions can be controlled by modulating PolQ and RPA

Yuan, Baolei; Bi, Chongwei; Li, M.

CRISPR-induced on-target large deletions can be controlled by modulating PolQ and RPA

Type

Presentation

Authors

Yuan, Baolei

Bi, Chongwei

Li, M.

KAUST Department

King Abdullah University of Science and Technology
Bioscience Program
Biological and Environmental Science and Engineering (BESE) Division

Date

2022-12-01

Abstract

CRISPR-Cas9, an efficient genome editing tool, has been widelyused in research and holds great promise in the clinic. However, thelarge unintended rearrangements of the genome after CRISPR-Cas9editing occur frequently and their potential risk cannot be ignored. In this study, we detected large deletions (LDs) induced by Cas9 inhuman embryonic stem cells (hESCs) and found micro-homologyend joining (MMEJ) repair pathway plays a predominant role inLD. We genetically targeted PARP1, RPA, PolQ and Lig3, whichplays critical roles in MMEJ, during CRISPR-Cas9 editing. Wefound that knocking down PARP1 and Lig3 does not alter LDfrequency, while knocking down or inhibiting PolQ dramaticallyreduces Cas9-induced LD frequency. Knocking down RPA in-creases LD frequency, and consistently, overexpression of RPAsreduces the frequency of LD. In conclusion, RPAs and PolQ playopposite roles in Cas9-induced LD and may be promising targets forreducing large rearrangement frequency during genome editing.