The position of the target site for engineered nucleases improves the aberrant mRNA clearance in in vivo genome editing
Engineered nucleases are widely used for creating frameshift or nonsense mutations in the target genes to eliminate gene functions. The resulting mRNAs carrying premature termination codons can be eliminated by nonsense-mediated mRNA decay. However, it is unclear how effective this process would be...
Autores principales: | , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Nature Publishing Group UK
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7060192/ https://www.ncbi.nlm.nih.gov/pubmed/32144373 http://dx.doi.org/10.1038/s41598-020-61154-4 |
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author | Lee, Jae Hoon Yu, Sungsook Nam, Tae Wook Roh, Jae-il Jin, Young Han, Jeong Pil Cha, Ji-Young Kim, Yoon Ki Yeom, Su-Cheong Nam, Ki Taek Lee, Han-Woong |
author_facet | Lee, Jae Hoon Yu, Sungsook Nam, Tae Wook Roh, Jae-il Jin, Young Han, Jeong Pil Cha, Ji-Young Kim, Yoon Ki Yeom, Su-Cheong Nam, Ki Taek Lee, Han-Woong |
author_sort | Lee, Jae Hoon |
collection | PubMed |
description | Engineered nucleases are widely used for creating frameshift or nonsense mutations in the target genes to eliminate gene functions. The resulting mRNAs carrying premature termination codons can be eliminated by nonsense-mediated mRNA decay. However, it is unclear how effective this process would be in vivo. Here, we found that the nonsense-mediated decay was unable to remove the mutant mRNAs in twelve out of sixteen homozygous mutant mice with frameshift mutations generated using engineered nucleases, which is far beyond what we expected. The frameshift mutant proteins translated by a single nucleotide deletion within the coding region were also detected in the p53 mutant mice. Furthermore, we showed that targeting the exons present downstream of the exons with a start codon or distant from ATG is relatively effective for eliminating mutant mRNAs in vivo, whereas the exons with a start codon are targeted to express the mutant mRNAs. Of the sixteen mutant mice generated, only four mutant mice targeting the downstream exons exhibited over 80% clearance of mutant mRNAs. Since the abnormal products, either mutant RNAs or mutant proteins, expressed by the target alleles might obscure the outcome of genome editing, these findings will provide insights in the improved performance of engineered nucleases when they are applied in vivo. |
format | Online Article Text |
id | pubmed-7060192 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | Nature Publishing Group UK |
record_format | MEDLINE/PubMed |
spelling | pubmed-70601922020-03-18 The position of the target site for engineered nucleases improves the aberrant mRNA clearance in in vivo genome editing Lee, Jae Hoon Yu, Sungsook Nam, Tae Wook Roh, Jae-il Jin, Young Han, Jeong Pil Cha, Ji-Young Kim, Yoon Ki Yeom, Su-Cheong Nam, Ki Taek Lee, Han-Woong Sci Rep Article Engineered nucleases are widely used for creating frameshift or nonsense mutations in the target genes to eliminate gene functions. The resulting mRNAs carrying premature termination codons can be eliminated by nonsense-mediated mRNA decay. However, it is unclear how effective this process would be in vivo. Here, we found that the nonsense-mediated decay was unable to remove the mutant mRNAs in twelve out of sixteen homozygous mutant mice with frameshift mutations generated using engineered nucleases, which is far beyond what we expected. The frameshift mutant proteins translated by a single nucleotide deletion within the coding region were also detected in the p53 mutant mice. Furthermore, we showed that targeting the exons present downstream of the exons with a start codon or distant from ATG is relatively effective for eliminating mutant mRNAs in vivo, whereas the exons with a start codon are targeted to express the mutant mRNAs. Of the sixteen mutant mice generated, only four mutant mice targeting the downstream exons exhibited over 80% clearance of mutant mRNAs. Since the abnormal products, either mutant RNAs or mutant proteins, expressed by the target alleles might obscure the outcome of genome editing, these findings will provide insights in the improved performance of engineered nucleases when they are applied in vivo. Nature Publishing Group UK 2020-03-06 /pmc/articles/PMC7060192/ /pubmed/32144373 http://dx.doi.org/10.1038/s41598-020-61154-4 Text en © The Author(s) 2020 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. |
spellingShingle | Article Lee, Jae Hoon Yu, Sungsook Nam, Tae Wook Roh, Jae-il Jin, Young Han, Jeong Pil Cha, Ji-Young Kim, Yoon Ki Yeom, Su-Cheong Nam, Ki Taek Lee, Han-Woong The position of the target site for engineered nucleases improves the aberrant mRNA clearance in in vivo genome editing |
title | The position of the target site for engineered nucleases improves the aberrant mRNA clearance in in vivo genome editing |
title_full | The position of the target site for engineered nucleases improves the aberrant mRNA clearance in in vivo genome editing |
title_fullStr | The position of the target site for engineered nucleases improves the aberrant mRNA clearance in in vivo genome editing |
title_full_unstemmed | The position of the target site for engineered nucleases improves the aberrant mRNA clearance in in vivo genome editing |
title_short | The position of the target site for engineered nucleases improves the aberrant mRNA clearance in in vivo genome editing |
title_sort | position of the target site for engineered nucleases improves the aberrant mrna clearance in in vivo genome editing |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7060192/ https://www.ncbi.nlm.nih.gov/pubmed/32144373 http://dx.doi.org/10.1038/s41598-020-61154-4 |
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