Cargando…
Engineering Nme2Cas9 Adenine Base Editors with Improved Activity and Targeting Scope
Nme2Cas9 has been established as a genome editing platform with compact size, high accuracy, and broad targeting range, including single-AAV-deliverable adenine base editors. Here, we have engineered Nme2Cas9 to further increase the activity and targeting scope of compact Nme2Cas9 base editors. We f...
Autores principales: | , , , , , , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Cold Spring Harbor Laboratory
2023
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10153126/ https://www.ncbi.nlm.nih.gov/pubmed/37131611 http://dx.doi.org/10.1101/2023.04.14.536905 |
_version_ | 1785035875622584320 |
---|---|
author | Bamidele, Nathan Zhang, Han Dong, Xiaolong Gaston, Nicholas Cheng, Haoyang Kelly, Karen Watts, Jonathan K. Xie, Jun Gao, Guangping Sontheimer, Erik J. |
author_facet | Bamidele, Nathan Zhang, Han Dong, Xiaolong Gaston, Nicholas Cheng, Haoyang Kelly, Karen Watts, Jonathan K. Xie, Jun Gao, Guangping Sontheimer, Erik J. |
author_sort | Bamidele, Nathan |
collection | PubMed |
description | Nme2Cas9 has been established as a genome editing platform with compact size, high accuracy, and broad targeting range, including single-AAV-deliverable adenine base editors. Here, we have engineered Nme2Cas9 to further increase the activity and targeting scope of compact Nme2Cas9 base editors. We first used domain insertion to position the deaminase domain nearer the displaced DNA strand in the target-bound complex. These domain-inlaid Nme2Cas9 variants exhibited shifted editing windows and increased activity in comparison to the N-terminally fused Nme2-ABE. We next expanded the editing scope by swapping the Nme2Cas9 PAM-interacting domain with that of SmuCas9, which we had previously defined as recognizing a single-cytidine PAM. We used these enhancements to correct two common MECP2 mutations associated with Rett syndrome with little or no bystander editing. Finally, we validated domain-inlaid Nme2-ABEs for single-AAV delivery in vivo. |
format | Online Article Text |
id | pubmed-10153126 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | Cold Spring Harbor Laboratory |
record_format | MEDLINE/PubMed |
spelling | pubmed-101531262023-05-03 Engineering Nme2Cas9 Adenine Base Editors with Improved Activity and Targeting Scope Bamidele, Nathan Zhang, Han Dong, Xiaolong Gaston, Nicholas Cheng, Haoyang Kelly, Karen Watts, Jonathan K. Xie, Jun Gao, Guangping Sontheimer, Erik J. bioRxiv Article Nme2Cas9 has been established as a genome editing platform with compact size, high accuracy, and broad targeting range, including single-AAV-deliverable adenine base editors. Here, we have engineered Nme2Cas9 to further increase the activity and targeting scope of compact Nme2Cas9 base editors. We first used domain insertion to position the deaminase domain nearer the displaced DNA strand in the target-bound complex. These domain-inlaid Nme2Cas9 variants exhibited shifted editing windows and increased activity in comparison to the N-terminally fused Nme2-ABE. We next expanded the editing scope by swapping the Nme2Cas9 PAM-interacting domain with that of SmuCas9, which we had previously defined as recognizing a single-cytidine PAM. We used these enhancements to correct two common MECP2 mutations associated with Rett syndrome with little or no bystander editing. Finally, we validated domain-inlaid Nme2-ABEs for single-AAV delivery in vivo. Cold Spring Harbor Laboratory 2023-04-18 /pmc/articles/PMC10153126/ /pubmed/37131611 http://dx.doi.org/10.1101/2023.04.14.536905 Text en https://creativecommons.org/licenses/by-nc-nd/4.0/This work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License (https://creativecommons.org/licenses/by-nc-nd/4.0/) , which allows reusers to copy and distribute the material in any medium or format in unadapted form only, for noncommercial purposes only, and only so long as attribution is given to the creator. |
spellingShingle | Article Bamidele, Nathan Zhang, Han Dong, Xiaolong Gaston, Nicholas Cheng, Haoyang Kelly, Karen Watts, Jonathan K. Xie, Jun Gao, Guangping Sontheimer, Erik J. Engineering Nme2Cas9 Adenine Base Editors with Improved Activity and Targeting Scope |
title | Engineering Nme2Cas9 Adenine Base Editors with Improved Activity and Targeting Scope |
title_full | Engineering Nme2Cas9 Adenine Base Editors with Improved Activity and Targeting Scope |
title_fullStr | Engineering Nme2Cas9 Adenine Base Editors with Improved Activity and Targeting Scope |
title_full_unstemmed | Engineering Nme2Cas9 Adenine Base Editors with Improved Activity and Targeting Scope |
title_short | Engineering Nme2Cas9 Adenine Base Editors with Improved Activity and Targeting Scope |
title_sort | engineering nme2cas9 adenine base editors with improved activity and targeting scope |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10153126/ https://www.ncbi.nlm.nih.gov/pubmed/37131611 http://dx.doi.org/10.1101/2023.04.14.536905 |
work_keys_str_mv | AT bamidelenathan engineeringnme2cas9adeninebaseeditorswithimprovedactivityandtargetingscope AT zhanghan engineeringnme2cas9adeninebaseeditorswithimprovedactivityandtargetingscope AT dongxiaolong engineeringnme2cas9adeninebaseeditorswithimprovedactivityandtargetingscope AT gastonnicholas engineeringnme2cas9adeninebaseeditorswithimprovedactivityandtargetingscope AT chenghaoyang engineeringnme2cas9adeninebaseeditorswithimprovedactivityandtargetingscope AT kellykaren engineeringnme2cas9adeninebaseeditorswithimprovedactivityandtargetingscope AT wattsjonathank engineeringnme2cas9adeninebaseeditorswithimprovedactivityandtargetingscope AT xiejun engineeringnme2cas9adeninebaseeditorswithimprovedactivityandtargetingscope AT gaoguangping engineeringnme2cas9adeninebaseeditorswithimprovedactivityandtargetingscope AT sontheimererikj engineeringnme2cas9adeninebaseeditorswithimprovedactivityandtargetingscope |