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Targeted activation of diverse CRISPR-Cas systems for mammalian genome editing via proximal CRISPR targeting

Bacterial CRISPR–Cas systems comprise diverse effector endonucleases with different targeting ranges, specificities and enzymatic properties, but many of them are inactive in mammalian cells and are thus precluded from genome-editing applications. Here we show that the type II-B FnCas9 from Francise...

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Autores principales: Chen, Fuqiang, Ding, Xiao, Feng, Yongmei, Seebeck, Timothy, Jiang, Yanfang, Davis, Gregory D.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Nature Publishing Group 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5385574/
https://www.ncbi.nlm.nih.gov/pubmed/28387220
http://dx.doi.org/10.1038/ncomms14958
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author Chen, Fuqiang
Ding, Xiao
Feng, Yongmei
Seebeck, Timothy
Jiang, Yanfang
Davis, Gregory D.
author_facet Chen, Fuqiang
Ding, Xiao
Feng, Yongmei
Seebeck, Timothy
Jiang, Yanfang
Davis, Gregory D.
author_sort Chen, Fuqiang
collection PubMed
description Bacterial CRISPR–Cas systems comprise diverse effector endonucleases with different targeting ranges, specificities and enzymatic properties, but many of them are inactive in mammalian cells and are thus precluded from genome-editing applications. Here we show that the type II-B FnCas9 from Francisella novicida possesses novel properties, but its nuclease function is frequently inhibited at many genomic loci in living human cells. Moreover, we develop a proximal CRISPR (termed proxy-CRISPR) targeting method that restores FnCas9 nuclease activity in a target-specific manner. We further demonstrate that this proxy-CRISPR strategy is applicable to diverse CRISPR–Cas systems, including type II-C Cas9 and type V Cpf1 systems, and can facilitate precise gene editing even between identical genomic sites within the same genome. Our findings provide a novel strategy to enable use of diverse otherwise inactive CRISPR–Cas systems for genome-editing applications and a potential path to modulate the impact of chromatin microenvironments on genome modification.
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spelling pubmed-53855742017-04-26 Targeted activation of diverse CRISPR-Cas systems for mammalian genome editing via proximal CRISPR targeting Chen, Fuqiang Ding, Xiao Feng, Yongmei Seebeck, Timothy Jiang, Yanfang Davis, Gregory D. Nat Commun Article Bacterial CRISPR–Cas systems comprise diverse effector endonucleases with different targeting ranges, specificities and enzymatic properties, but many of them are inactive in mammalian cells and are thus precluded from genome-editing applications. Here we show that the type II-B FnCas9 from Francisella novicida possesses novel properties, but its nuclease function is frequently inhibited at many genomic loci in living human cells. Moreover, we develop a proximal CRISPR (termed proxy-CRISPR) targeting method that restores FnCas9 nuclease activity in a target-specific manner. We further demonstrate that this proxy-CRISPR strategy is applicable to diverse CRISPR–Cas systems, including type II-C Cas9 and type V Cpf1 systems, and can facilitate precise gene editing even between identical genomic sites within the same genome. Our findings provide a novel strategy to enable use of diverse otherwise inactive CRISPR–Cas systems for genome-editing applications and a potential path to modulate the impact of chromatin microenvironments on genome modification. Nature Publishing Group 2017-04-07 /pmc/articles/PMC5385574/ /pubmed/28387220 http://dx.doi.org/10.1038/ncomms14958 Text en Copyright © 2017, The Author(s) http://creativecommons.org/licenses/by/4.0/ This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article's Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/
spellingShingle Article
Chen, Fuqiang
Ding, Xiao
Feng, Yongmei
Seebeck, Timothy
Jiang, Yanfang
Davis, Gregory D.
Targeted activation of diverse CRISPR-Cas systems for mammalian genome editing via proximal CRISPR targeting
title Targeted activation of diverse CRISPR-Cas systems for mammalian genome editing via proximal CRISPR targeting
title_full Targeted activation of diverse CRISPR-Cas systems for mammalian genome editing via proximal CRISPR targeting
title_fullStr Targeted activation of diverse CRISPR-Cas systems for mammalian genome editing via proximal CRISPR targeting
title_full_unstemmed Targeted activation of diverse CRISPR-Cas systems for mammalian genome editing via proximal CRISPR targeting
title_short Targeted activation of diverse CRISPR-Cas systems for mammalian genome editing via proximal CRISPR targeting
title_sort targeted activation of diverse crispr-cas systems for mammalian genome editing via proximal crispr targeting
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5385574/
https://www.ncbi.nlm.nih.gov/pubmed/28387220
http://dx.doi.org/10.1038/ncomms14958
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