DNA Repair Pathway Choices in CRISPR-Cas9-Mediated Genome Editing

• Published in: Trends in genetics Vol. 37; no. 7; pp. 639 - 656
• Main Authors: Xue, Chaoyou, Greene, Eric C.
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 01.07.2021
• Subjects: CRISPR-Cas Systems - genetics; DNA Breaks, Double-Stranded; DNA End-Joining Repair - genetics; DNA Repair - genetics; Gene Editing - trends; Genome, Human - genetics; Homologous Recombination - genetics; Humans; Mutagenesis, Insertional - genetics; Signal Transduction - genetics
• ISSN: 0168-9525
• DOI: 10.1016/j.tig.2021.02.008

Many clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein 9 (Cas9)-based genome editing technologies take advantage of Cas nucleases to induce DNA double-strand breaks (DSBs) at desired locations within a genome. Further processing of the DSBs by the cellular DSB repair machinery is then necessary to introduce desired mutations, sequence insertions, or gene deletions. Thus, the accuracy and efficiency of genome editing are influenced by the cellular DSB repair pathways. DSBs are themselves highly genotoxic lesions and as such cells have evolved multiple mechanisms for their repair. These repair pathways include homologous recombination (HR), classical nonhomologous end joining (cNHEJ), microhomology-mediated end joining (MMEJ) and single-strand annealing (SSA). In this review, we briefly highlight CRISPR-Cas9 and then describe the mechanisms of DSB repair. Finally, we summarize recent findings of factors that can influence the choice of DNA repair pathway in response to Cas9-induced DSBs. Clustered regularly interspaced short palindromic repeats (CRISPR)- CRISPR-associated protein 9 (Cas9)-mediated genome editing offers a powerful approach as a potential therapy for monogenic human genetic diseases.Precise template-free base deletions can be achieved through microhomology-mediated end joining (MMEJ) repair and depend on local target site sequence.The DNA repair pathway choice in CRISPR-Cas9 induced-double-strand breaks (DSBs) is regulated by several key factors including the cell cycle, target site sequence and chromatin structure, and the identity of the donor DNA template.Homology-directed repair (HDR)-related DNA repair pathways in response to CRISPR-Cas9 induced-DSBs in mammalian cells are complicated and relatively inefficient. Different DNA repair pathways might be used to repair each end at a DSB resulting in the potential for asymmetric repair.