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Engineered IscB-ωRNA system with expanded target range for base editing.
Engineered IscB-ωRNA system with expanded target range for base editing.
- Source :
-
Nature chemical biology [Nat Chem Biol] 2025 Jan; Vol. 21 (1), pp. 100-108. Date of Electronic Publication: 2024 Aug 15. - Publication Year :
- 2025
-
Abstract
- As the evolutionary ancestor of Cas9 nuclease, IscB proteins serve as compact RNA-guided DNA endonucleases and nickases, making them strong candidates for base editing. Nevertheless, the narrow targeting scope limits the application of IscB systems; thus, it is necessary to find more IscBs that recognize different target-adjacent motifs (TAMs). Here, we identified 10 of 19 uncharacterized IscB proteins from uncultured microbes with activity in mammalian cells. Through protein and ωRNA engineering, we further enhanced the activity of IscB ortholog IscB.m16 and expanded its TAM scope from MRNRAA to NNNGNA, resulting in a variant named IscB.m16*. By fusing the deaminase domains with IscB.m16* nickase, we generated IscB.m16*-derived base editors that exhibited robust base-editing efficiency in mammalian cells and effectively restored Duchenne muscular dystrophy proteins in diseased mice through single adeno-associated virus delivery. Thus, this study establishes a set of compact base-editing tools for basic research and therapeutic applications.<br />Competing Interests: Competing interests: H.Z. and Q.X. disclose a patent application (PCT/CN2024/071744) related to the proteins described in this manuscript. H.Y. and L.S. are cofounders of HUIDAGENE Therapeutics. The other authors declare no competing interests.<br /> (© 2024. The Author(s).)
- Subjects :
- Animals
Mice
Humans
HEK293 Cells
RNA, Guide, CRISPR-Cas Systems genetics
RNA, Guide, CRISPR-Cas Systems metabolism
RNA metabolism
RNA chemistry
RNA genetics
CRISPR-Cas Systems
Bacterial Proteins genetics
Bacterial Proteins metabolism
Bacterial Proteins chemistry
Dependovirus genetics
Gene Editing methods
Muscular Dystrophy, Duchenne genetics
Muscular Dystrophy, Duchenne metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 1552-4469
- Volume :
- 21
- Issue :
- 1
- Database :
- MEDLINE
- Journal :
- Nature chemical biology
- Publication Type :
- Academic Journal
- Accession number :
- 39147927
- Full Text :
- https://doi.org/10.1038/s41589-024-01706-1