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Increasing the efficiency and targeting range of cytidine base editors through fusion of a single-stranded DNA-binding protein domain.
- Source :
-
Nature cell biology [Nat Cell Biol] 2020 Jun; Vol. 22 (6), pp. 740-750. Date of Electronic Publication: 2020 May 11. - Publication Year :
- 2020
-
Abstract
- Cytidine base editors are powerful genetic tools that catalyse cytidine to thymidine conversion at specific genomic loci, and further improvement of the editing range and efficiency is critical for their broader applications. Through insertion of a non-sequence-specific single-stranded DNA-binding domain from Rad51 protein between Cas9 nickase and the deaminases, serial hyper cytidine base editors were generated with substantially increased activity and an expanded editing window towards the protospacer adjacent motif in both cell lines and mouse embryos. Additionally, hyeA3A-BE4max selectively catalysed cytidine conversion in TC motifs with a broader editing range and much higher activity (up to 257-fold) compared with eA3A-BE4max. Moreover, hyeA3A-BE4max specifically generated a C-to-T conversion without inducing bystander mutations in the haemoglobin gamma gene promoter to mimic a naturally occurring genetic variant for amelioration of β-haemoglobinopathy, suggesting the therapeutic potential of the improved base editors.
- Subjects :
- Animals
Cell Differentiation
Cytidine chemistry
DNA-Binding Proteins genetics
Embryo, Mammalian cytology
Embryo, Mammalian metabolism
Female
HEK293 Cells
Humans
Mice
Mice, Inbred C57BL
Mice, Inbred ICR
Protein Domains
Rad51 Recombinase genetics
CRISPR-Cas Systems
Cytidine genetics
DNA-Binding Proteins metabolism
Gene Editing
Mutation
Rad51 Recombinase metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 1476-4679
- Volume :
- 22
- Issue :
- 6
- Database :
- MEDLINE
- Journal :
- Nature cell biology
- Publication Type :
- Academic Journal
- Accession number :
- 32393889
- Full Text :
- https://doi.org/10.1038/s41556-020-0518-8