Engineering IscB to develop highly efficient miniature editing tools in mammalian cells and embryos.
Mol Cell
; 84(16): 3128-3140.e4, 2024 Aug 22.
Article
in En
| MEDLINE
| ID: mdl-39096898
ABSTRACT
The IscB proteins, as the ancestors of Cas9 endonuclease, hold great promise due to their small size and potential for diverse genome editing. However, their activity in mammalian cells is unsatisfactory. By introducing three residual substitutions in IscB, we observed an average 7.5-fold increase in activity. Through fusing a sequence-non-specific DNA-binding protein domain, the eIscB-D variant achieved higher editing efficiency, with a maximum of 91.3%. Moreover, engineered ωRNA was generated with a 20% reduction in length and slightly increased efficiency. The engineered eIscB-D/eωRNA system showed an average 20.2-fold increase in activity compared with the original IscB. Furthermore, we successfully adapted eIscB-D for highly efficient cytosine and adenine base editing. Notably, eIscB-D is highly active in mouse cell lines and embryos, enabling the efficient generation of disease models through mRNA/ωRNA injection. Our study suggests that these miniature genome-editing tools have great potential for diverse applications.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
CRISPR-Cas Systems
/
Gene Editing
Limits:
Animals
/
Humans
Language:
En
Journal:
Mol Cell
/
Mol. cell
/
Molecular cell
Journal subject:
BIOLOGIA MOLECULAR
Year:
2024
Document type:
Article
Affiliation country:
China
Country of publication:
Estados Unidos