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BESST: a novel LncRNA knockout strategy with less genome perturbance

Long noncoding RNAs (lncRNAs) are >200 nt RNA transcripts without protein-coding potential. LncRNAs can be categorized into intergenic, intronic, bidirectional, sense, and antisense lncRNAs based on the genomic localization to nearby protein-coding genes. The current CRISPR-based lncRNA knockout...

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Autores principales: Zhang, Shikuan, Chen, Yue, Dong, Kunzhe, Zhao, Yiwan, Wang, Yanzhi, Wang, Songmao, Qu, Chen, Xu, Naihan, Xie, Weidong, Zeng, Chunyu, Lyu, Qing Rex, Zhang, Yaou
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Oxford University Press 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10201427/
https://www.ncbi.nlm.nih.gov/pubmed/36938886
http://dx.doi.org/10.1093/nar/gkad197
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author Zhang, Shikuan
Chen, Yue
Dong, Kunzhe
Zhao, Yiwan
Wang, Yanzhi
Wang, Songmao
Qu, Chen
Xu, Naihan
Xie, Weidong
Zeng, Chunyu
Lyu, Qing Rex
Zhang, Yaou
author_facet Zhang, Shikuan
Chen, Yue
Dong, Kunzhe
Zhao, Yiwan
Wang, Yanzhi
Wang, Songmao
Qu, Chen
Xu, Naihan
Xie, Weidong
Zeng, Chunyu
Lyu, Qing Rex
Zhang, Yaou
author_sort Zhang, Shikuan
collection PubMed
description Long noncoding RNAs (lncRNAs) are >200 nt RNA transcripts without protein-coding potential. LncRNAs can be categorized into intergenic, intronic, bidirectional, sense, and antisense lncRNAs based on the genomic localization to nearby protein-coding genes. The current CRISPR-based lncRNA knockout strategy works efficiently for lncRNAs distant from the protein-coding gene, whereas it causes genomic perturbance inevitably due to technical limitations. In this study, we introduce a novel lncRNA knockout strategy, BESST, by deleting the genomic DNA fragment from the branch point to the 3′ splicing site in the last intron of the target lncRNA. The BESST knockout exhibited comparable or superior repressive efficiency to RNA silencing or conventional promoter-exon1 deletion. Significantly, the BESST knockout strategy minimized the intervention of adjacent/overlap protein-coding genes by removing an average of ∼130 bp from genomic DNA. Our data also found that the BESST knockout strategy causes lncRNA nuclear retention, resulting in decapping and deadenylation of the lncRNA poly(A) tail. Further study revealed that PABPN1 is essential for the BESST-mediated decay and subsequent poly(A) deadenylation and decapping. Together, the BESST knockout strategy provides a versatile tool for investigating gene function by generating knockout cells or animals with high specificity and efficiency.
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spelling pubmed-102014272023-05-23 BESST: a novel LncRNA knockout strategy with less genome perturbance Zhang, Shikuan Chen, Yue Dong, Kunzhe Zhao, Yiwan Wang, Yanzhi Wang, Songmao Qu, Chen Xu, Naihan Xie, Weidong Zeng, Chunyu Lyu, Qing Rex Zhang, Yaou Nucleic Acids Res Methods Online Long noncoding RNAs (lncRNAs) are >200 nt RNA transcripts without protein-coding potential. LncRNAs can be categorized into intergenic, intronic, bidirectional, sense, and antisense lncRNAs based on the genomic localization to nearby protein-coding genes. The current CRISPR-based lncRNA knockout strategy works efficiently for lncRNAs distant from the protein-coding gene, whereas it causes genomic perturbance inevitably due to technical limitations. In this study, we introduce a novel lncRNA knockout strategy, BESST, by deleting the genomic DNA fragment from the branch point to the 3′ splicing site in the last intron of the target lncRNA. The BESST knockout exhibited comparable or superior repressive efficiency to RNA silencing or conventional promoter-exon1 deletion. Significantly, the BESST knockout strategy minimized the intervention of adjacent/overlap protein-coding genes by removing an average of ∼130 bp from genomic DNA. Our data also found that the BESST knockout strategy causes lncRNA nuclear retention, resulting in decapping and deadenylation of the lncRNA poly(A) tail. Further study revealed that PABPN1 is essential for the BESST-mediated decay and subsequent poly(A) deadenylation and decapping. Together, the BESST knockout strategy provides a versatile tool for investigating gene function by generating knockout cells or animals with high specificity and efficiency. Oxford University Press 2023-03-20 /pmc/articles/PMC10201427/ /pubmed/36938886 http://dx.doi.org/10.1093/nar/gkad197 Text en © The Author(s) 2023. Published by Oxford University Press on behalf of Nucleic Acids Research. https://creativecommons.org/licenses/by-nc/4.0/This is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial License (https://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com
spellingShingle Methods Online
Zhang, Shikuan
Chen, Yue
Dong, Kunzhe
Zhao, Yiwan
Wang, Yanzhi
Wang, Songmao
Qu, Chen
Xu, Naihan
Xie, Weidong
Zeng, Chunyu
Lyu, Qing Rex
Zhang, Yaou
BESST: a novel LncRNA knockout strategy with less genome perturbance
title BESST: a novel LncRNA knockout strategy with less genome perturbance
title_full BESST: a novel LncRNA knockout strategy with less genome perturbance
title_fullStr BESST: a novel LncRNA knockout strategy with less genome perturbance
title_full_unstemmed BESST: a novel LncRNA knockout strategy with less genome perturbance
title_short BESST: a novel LncRNA knockout strategy with less genome perturbance
title_sort besst: a novel lncrna knockout strategy with less genome perturbance
topic Methods Online
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10201427/
https://www.ncbi.nlm.nih.gov/pubmed/36938886
http://dx.doi.org/10.1093/nar/gkad197
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