Development of a versatile nuclease prime editor with upgraded precision.
Nat Commun
; 14(1): 305, 2023 01 19.
Article
em En
| MEDLINE
| ID: mdl-36658146
ABSTRACT
The applicability of nuclease-based form of prime editor (PEn) has been hindered by its complexed editing outcomes. A chemical inhibitor against DNA-PK, which mediates the nonhomologous end joining (NHEJ) pathway, was recently shown to promote precise insertions by PEn. Nevertheless, the intrinsic issues of specificity and toxicity for such a chemical approach necessitate development of alternative strategies. Here, we find that co-introduction of PEn and a NHEJ-restraining, 53BP1-inhibitory ubiquitin variant potently drives precise edits via mitigation of unintended edits, framing a high-activity editing platform (uPEn) apparently complementing the canonical PE. Further developments involve exploring the effective configuration of a homologous region-containing pegRNA (HR-pegRNA). Overall, uPEn can empower high-efficiency installation of insertions (38%), deletions (43%) and replacements (52%) in HEK293T cells. When compared with PE3/5max, uPEn demonstrates superior activities for typically refractory base substitutions, and for small-block edits. Collectively, this work establishes a highly efficient PE platform with broad application potential.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Quebras de DNA de Cadeia Dupla
/
Edição de Genes
Limite:
Humans
Idioma:
En
Ano de publicação:
2023
Tipo de documento:
Article