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Updated: Jun 4, 2025

Gene Digital Circuits Based on CRISPR-Cas Systems and Anti-CRISPR Proteins
Published on: October 18, 2022
Cell cycle-dependent regulation of CRISPR-Cas9 repetitive activation by anti-CRISPR and Cdt1 fusion in the CRISPRa
Kanae Kishi1, Kiyomi Nigorikawa1,2, Yuki Hasegawa2
1Graduate School of Biomedical and Health Sciences, Hiroshima University, Japan.
Abstract:
CRISPR-Cas9 is a widely used genome-editing tool. We previously developed a method with improved homology-directed repair efficiency and reduced off-target effects by utilizing a fusion protein of AcrIIA4, a Cas9 inhibitor, and Cdt1, which accumulates in the G1 phase and activates Cas9 only in the S/G2 phase. However, it is unknown whether Cas9 inhibition by AcrIIA4 + Cdt1 occurs repeatedly in the G1 phase as the cell cycle progresses. In this study, we used the CRISPRa system to monitor changes in the interaction between Cas9 and AcrIIA4 + Cdt1 at single-cell resolution and in real time. Our findings are among the few examples of successful detection of fluctuating protein-protein interactions that oscillate over time.
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