PE-STOP: A versatile tool for installing nonsense substitutions amenable for precise reversion.
PE-STOP: A versatile tool for installing nonsense substitutions amenable for precise reversion.
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DOI：
10.1016/j.jbc.2023.104942
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发表时间：
2023-08
影响因子：
4.8
通讯作者：
Wang, Xiaolong
中科院分区：
文献类型：
--
作者：
Song, Ziguo;Zhang, Guiquan;Huang, Shuhong;Liu, Yao;Li, Guanglei;Zhou, Xianhui;Sun, Jiayuan;Gao, Pengfei;Chen, Yulin;Huang, Xingxu;Liu, Jianghuai;Wang, Xiaolong
The rapid advances in genome editing technologies have revolutionized the study of gene functions in cell or animal models. The recent generation of double-stranded DNA cleavage–independent base editors has been suitably adapted for interrogation of protein-coding genes on the basis of introducing premature stop codons or disabling the start codons. However, such versions of stop/start codon–oriented genetic tools still present limitations on their versatility, base-level precision, and target specificity. Here, we exploit a newly developed prime editor (PE) that differs from base editors by its adoption of a reverse transcriptase activity, which enables incorporation of various types of precise edits templated by a specialized prime editing guide RNA. Based on such a versatile platform, we established a prime editing–empowered method (PE-STOP) for installation of nonsense substitutions, providing a complementary approach to the present gene-targeting tools. PE-STOP is bioinformatically predicted to feature substantially expanded coverage in the genome space. In practice, PE-STOP introduces stop codons with good efficiencies in human embryonic kidney 293T and N2a cells (with medians of 29% [ten sites] and 25% [four sites] editing efficiencies, respectively), while exhibiting minimal off-target effects and high on-target precision. Furthermore, given the fact that PE installs prime editing guide RNA–templated mutations, we introduce a unique strategy for precise genetic rescue of PE-STOP-dependent nonsense mutation via the same PE platform. Altogether, the present work demonstrates a versatile and specific tool for gene inactivation and for functional interrogation of nonsense mutations.
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影响因子：
9.8
作者：
Coban-Akdemir, Zeynep;White, Janson J.;Carvalho, Claudia M. B.
通讯作者：
Carvalho, Claudia M. B.
影响因子：
64.5
作者：
Hsu PD;Lander ES;Zhang F
通讯作者：
Zhang F
影响因子：
--
作者：
Giuliano, Christopher J;Lin, Ann;Sheltzer, Jason M
通讯作者：
Sheltzer, Jason M
影响因子：
5.3
作者：
DUBRIDGE, RB;TANG, P;CALOS, MP
通讯作者：
CALOS, MP
影响因子：
64.8
作者：
Anzalone, Andrew V.;Randolph, Peyton B.;Liu, David R.
通讯作者：
Liu, David R.