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Nat Commun. 2018 Jun 5;9(1):2184. doi: 10.1038/s41467-018-04580-3.

In vivo base editing of post-mitotic sensory cells.

Yeh WH1,2,3,4,5, Chiang H4,6, Rees HA1,2,3, Edge ASB7,8,9,10, Liu DR11,12,13.

Author information

1
Department of Chemistry and Chemical Biology, Harvard University, Cambridge, MA, 02138, USA.
2
Howard Hughes Medical Institute, Harvard University, Cambridge, MA, 02138, USA.
3
Merkin Institute of Transformative Technologies in Healthcare, Broad Institute of MIT and Harvard, Cambridge, MA, 02142, USA.
4
Eaton-Peabody Laboratory, Massachusetts Eye and Ear, Boston, MA, 02114, USA.
5
Program in Speech and Hearing Bioscience and Technology, Harvard Medical School, Boston, MA, 02115, USA.
6
Department of Otolaryngology, Harvard Medical School, Boston, MA, 02115, USA.
7
Eaton-Peabody Laboratory, Massachusetts Eye and Ear, Boston, MA, 02114, USA. albert_edge@meei.harvard.edu.
8
Program in Speech and Hearing Bioscience and Technology, Harvard Medical School, Boston, MA, 02115, USA. albert_edge@meei.harvard.edu.
9
Department of Otolaryngology, Harvard Medical School, Boston, MA, 02115, USA. albert_edge@meei.harvard.edu.
10
Harvard Stem Cell Institute, Cambridge, MA, 02138, USA. albert_edge@meei.harvard.edu.
11
Department of Chemistry and Chemical Biology, Harvard University, Cambridge, MA, 02138, USA. drliu@fas.harvard.edu.
12
Howard Hughes Medical Institute, Harvard University, Cambridge, MA, 02138, USA. drliu@fas.harvard.edu.
13
Merkin Institute of Transformative Technologies in Healthcare, Broad Institute of MIT and Harvard, Cambridge, MA, 02142, USA. drliu@fas.harvard.edu.

Abstract

Programmable nucleases can introduce precise changes to genomic DNA through homology-directed repair (HDR). Unfortunately, HDR is largely restricted to mitotic cells, and is typically accompanied by an excess of stochastic insertions and deletions (indels). Here we present an in vivo base editing strategy that addresses these limitations. We use nuclease-free base editing to install a S33F mutation in β-catenin that blocks β-catenin phosphorylation, impedes β-catenin degradation, and upregulates Wnt signaling. In vitro, base editing installs the S33F mutation with a 200-fold higher editing:indel ratio than HDR. In post-mitotic cells in mouse inner ear, injection of base editor protein:RNA:lipid installs this mutation, resulting in Wnt activation that induces mitosis of cochlear supporting cells and cellular reprogramming. In contrast, injection of HDR agents does not induce Wnt upregulation. These results establish a strategy for modifying posttranslational states in signaling pathways, and an approach to precision editing in post-mitotic tissues.

PMID:
29872041
PMCID:
PMC5988727
DOI:
10.1038/s41467-018-04580-3
[Indexed for MEDLINE]
Free PMC Article

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