Programmable Base Editing in Mycobacterium tuberculosis Using an Engineered CRISPR RNA-Guided Cytidine Deaminase
Multidrug-resistant Mycobacterium tuberculosis (Mtb) infection seriously endangers global human health, creating an urgent need for new treatment strategies. Efficient genome editing tools can facilitate identification of key genes and pathways involved in bacterial physiology, pathogenesis, and dru...
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Frontiers Media S.A.
2021-12-01
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Series: | Frontiers in Genome Editing |
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Online Access: | https://www.frontiersin.org/articles/10.3389/fgeed.2021.734436/full |
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author | Xin-Yuan Ding Si-Shang Li Yi-Man Geng Mei-Yi Yan Guo-Bao Li Guo-Liang Zhang Yi-Cheng Sun |
author_facet | Xin-Yuan Ding Si-Shang Li Yi-Man Geng Mei-Yi Yan Guo-Bao Li Guo-Liang Zhang Yi-Cheng Sun |
author_sort | Xin-Yuan Ding |
collection | DOAJ |
description | Multidrug-resistant Mycobacterium tuberculosis (Mtb) infection seriously endangers global human health, creating an urgent need for new treatment strategies. Efficient genome editing tools can facilitate identification of key genes and pathways involved in bacterial physiology, pathogenesis, and drug resistance mechanisms, and thus contribute to the development of novel treatments for drug-resistant tuberculosis. Here, we report a two-plasmid system, MtbCBE, used to inactivate genes and introduce point mutations in Mtb. In this system, the assistant plasmid pRecX-NucSE107A expresses RecX and NucSE107A to repress RecA-dependent and NucS-dependent DNA repair systems, and the base editor plasmid pCBE expresses a fusion protein combining cytidine deaminase APOBEC1, Cas9 nickase (nCas9), and uracil DNA glycosylase inhibitor (UGI). Together, the two plasmids enabled efficient G:C to A:T base pair conversion at desired sites in the Mtb genome. The successful development of a base editing system will facilitate elucidation of the molecular mechanisms underlying Mtb pathogenesis and drug resistance and provide critical inspiration for the development of base editing tools in other microbes. |
first_indexed | 2024-12-17T19:26:53Z |
format | Article |
id | doaj.art-75994ec9eeba4a85a9eb8d6c2919e600 |
institution | Directory Open Access Journal |
issn | 2673-3439 |
language | English |
last_indexed | 2024-12-17T19:26:53Z |
publishDate | 2021-12-01 |
publisher | Frontiers Media S.A. |
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series | Frontiers in Genome Editing |
spelling | doaj.art-75994ec9eeba4a85a9eb8d6c2919e6002022-12-21T21:35:22ZengFrontiers Media S.A.Frontiers in Genome Editing2673-34392021-12-01310.3389/fgeed.2021.734436734436Programmable Base Editing in Mycobacterium tuberculosis Using an Engineered CRISPR RNA-Guided Cytidine DeaminaseXin-Yuan Ding0Si-Shang Li1Yi-Man Geng2Mei-Yi Yan3Guo-Bao Li4Guo-Liang Zhang5Yi-Cheng Sun6NHC Key Laboratory of Systems Biology of Pathogens, Institute of Pathogen Biology, Center for Tuberculosis Research, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, ChinaNHC Key Laboratory of Systems Biology of Pathogens, Institute of Pathogen Biology, Center for Tuberculosis Research, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, ChinaDepartment of Clinical Laboratory, Henan Provincial People’s Hospital, People’s Hospital of Zhengzhou University, Zhengzhou, ChinaNHC Key Laboratory of Systems Biology of Pathogens, Institute of Pathogen Biology, Center for Tuberculosis Research, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, ChinaNational Clinical Research Center for Infectious Diseases, Shenzhen Third People’s Hospital, Southern University of Science and Technology, Shenzhen, ChinaNational Clinical Research Center for Infectious Diseases, Shenzhen Third People’s Hospital, Southern University of Science and Technology, Shenzhen, ChinaNHC Key Laboratory of Systems Biology of Pathogens, Institute of Pathogen Biology, Center for Tuberculosis Research, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, ChinaMultidrug-resistant Mycobacterium tuberculosis (Mtb) infection seriously endangers global human health, creating an urgent need for new treatment strategies. Efficient genome editing tools can facilitate identification of key genes and pathways involved in bacterial physiology, pathogenesis, and drug resistance mechanisms, and thus contribute to the development of novel treatments for drug-resistant tuberculosis. Here, we report a two-plasmid system, MtbCBE, used to inactivate genes and introduce point mutations in Mtb. In this system, the assistant plasmid pRecX-NucSE107A expresses RecX and NucSE107A to repress RecA-dependent and NucS-dependent DNA repair systems, and the base editor plasmid pCBE expresses a fusion protein combining cytidine deaminase APOBEC1, Cas9 nickase (nCas9), and uracil DNA glycosylase inhibitor (UGI). Together, the two plasmids enabled efficient G:C to A:T base pair conversion at desired sites in the Mtb genome. The successful development of a base editing system will facilitate elucidation of the molecular mechanisms underlying Mtb pathogenesis and drug resistance and provide critical inspiration for the development of base editing tools in other microbes.https://www.frontiersin.org/articles/10.3389/fgeed.2021.734436/fullmycobacteriacytidine deaminasegenome editingCRISPRbase editing |
spellingShingle | Xin-Yuan Ding Si-Shang Li Yi-Man Geng Mei-Yi Yan Guo-Bao Li Guo-Liang Zhang Yi-Cheng Sun Programmable Base Editing in Mycobacterium tuberculosis Using an Engineered CRISPR RNA-Guided Cytidine Deaminase Frontiers in Genome Editing mycobacteria cytidine deaminase genome editing CRISPR base editing |
title | Programmable Base Editing in Mycobacterium tuberculosis Using an Engineered CRISPR RNA-Guided Cytidine Deaminase |
title_full | Programmable Base Editing in Mycobacterium tuberculosis Using an Engineered CRISPR RNA-Guided Cytidine Deaminase |
title_fullStr | Programmable Base Editing in Mycobacterium tuberculosis Using an Engineered CRISPR RNA-Guided Cytidine Deaminase |
title_full_unstemmed | Programmable Base Editing in Mycobacterium tuberculosis Using an Engineered CRISPR RNA-Guided Cytidine Deaminase |
title_short | Programmable Base Editing in Mycobacterium tuberculosis Using an Engineered CRISPR RNA-Guided Cytidine Deaminase |
title_sort | programmable base editing in mycobacterium tuberculosis using an engineered crispr rna guided cytidine deaminase |
topic | mycobacteria cytidine deaminase genome editing CRISPR base editing |
url | https://www.frontiersin.org/articles/10.3389/fgeed.2021.734436/full |
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