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Suppression of HBV replication by the expression of nickase- and nuclease dead-Cas9

Complete removal of hepatitis B virus (HBV) DNA from nuclei is difficult by the current therapies. Recent reports have shown that a novel genome-editing tool using Cas9 with a single-guide RNA (sgRNA) system can cleave the HBV genome in vitro and in vivo. However, induction of a double-strand break...

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Autores principales: Kurihara, Takeshi, Fukuhara, Takasuke, Ono, Chikako, Yamamoto, Satomi, Uemura, Kentaro, Okamoto, Toru, Sugiyama, Masaya, Motooka, Daisuke, Nakamura, Shota, Ikawa, Masato, Mizokami, Masashi, Maehara, Yoshihiko, Matsuura, Yoshiharu
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Nature Publishing Group UK 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5522428/
https://www.ncbi.nlm.nih.gov/pubmed/28733609
http://dx.doi.org/10.1038/s41598-017-05905-w
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author Kurihara, Takeshi
Fukuhara, Takasuke
Ono, Chikako
Yamamoto, Satomi
Uemura, Kentaro
Okamoto, Toru
Sugiyama, Masaya
Motooka, Daisuke
Nakamura, Shota
Ikawa, Masato
Mizokami, Masashi
Maehara, Yoshihiko
Matsuura, Yoshiharu
author_facet Kurihara, Takeshi
Fukuhara, Takasuke
Ono, Chikako
Yamamoto, Satomi
Uemura, Kentaro
Okamoto, Toru
Sugiyama, Masaya
Motooka, Daisuke
Nakamura, Shota
Ikawa, Masato
Mizokami, Masashi
Maehara, Yoshihiko
Matsuura, Yoshiharu
author_sort Kurihara, Takeshi
collection PubMed
description Complete removal of hepatitis B virus (HBV) DNA from nuclei is difficult by the current therapies. Recent reports have shown that a novel genome-editing tool using Cas9 with a single-guide RNA (sgRNA) system can cleave the HBV genome in vitro and in vivo. However, induction of a double-strand break (DSB) on the targeted genome by Cas9 risks undesirable off-target cleavage on the host genome. Nickase-Cas9 cleaves a single strand of DNA, and thereby two sgRNAs are required for inducing DSBs. To avoid Cas9-induced off-target mutagenesis, we examined the effects of the expressions of nickase-Cas9 and nuclease dead Cas9 (d-Cas9) with sgRNAs on HBV replication. The expression of nickase-Cas9 with a pair of sgRNAs cleaved the target HBV genome and suppressed the viral-protein expression and HBV replication in vitro. Moreover, nickase-Cas9 with the sgRNA pair cleaved the targeted HBV genome in mouse liver. Interestingly, d-Cas9 expression with the sgRNAs also suppressed HBV replication in vitro without cleaving the HBV genome. These results suggest the possible use of nickase-Cas9 and d-Cas9 with a pair of sgRNAs for eliminating HBV DNA from the livers of chronic hepatitis B patients with low risk of undesirable off-target mutation on the host genome.
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spelling pubmed-55224282017-07-26 Suppression of HBV replication by the expression of nickase- and nuclease dead-Cas9 Kurihara, Takeshi Fukuhara, Takasuke Ono, Chikako Yamamoto, Satomi Uemura, Kentaro Okamoto, Toru Sugiyama, Masaya Motooka, Daisuke Nakamura, Shota Ikawa, Masato Mizokami, Masashi Maehara, Yoshihiko Matsuura, Yoshiharu Sci Rep Article Complete removal of hepatitis B virus (HBV) DNA from nuclei is difficult by the current therapies. Recent reports have shown that a novel genome-editing tool using Cas9 with a single-guide RNA (sgRNA) system can cleave the HBV genome in vitro and in vivo. However, induction of a double-strand break (DSB) on the targeted genome by Cas9 risks undesirable off-target cleavage on the host genome. Nickase-Cas9 cleaves a single strand of DNA, and thereby two sgRNAs are required for inducing DSBs. To avoid Cas9-induced off-target mutagenesis, we examined the effects of the expressions of nickase-Cas9 and nuclease dead Cas9 (d-Cas9) with sgRNAs on HBV replication. The expression of nickase-Cas9 with a pair of sgRNAs cleaved the target HBV genome and suppressed the viral-protein expression and HBV replication in vitro. Moreover, nickase-Cas9 with the sgRNA pair cleaved the targeted HBV genome in mouse liver. Interestingly, d-Cas9 expression with the sgRNAs also suppressed HBV replication in vitro without cleaving the HBV genome. These results suggest the possible use of nickase-Cas9 and d-Cas9 with a pair of sgRNAs for eliminating HBV DNA from the livers of chronic hepatitis B patients with low risk of undesirable off-target mutation on the host genome. Nature Publishing Group UK 2017-07-21 /pmc/articles/PMC5522428/ /pubmed/28733609 http://dx.doi.org/10.1038/s41598-017-05905-w Text en © The Author(s) 2017 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
Kurihara, Takeshi
Fukuhara, Takasuke
Ono, Chikako
Yamamoto, Satomi
Uemura, Kentaro
Okamoto, Toru
Sugiyama, Masaya
Motooka, Daisuke
Nakamura, Shota
Ikawa, Masato
Mizokami, Masashi
Maehara, Yoshihiko
Matsuura, Yoshiharu
Suppression of HBV replication by the expression of nickase- and nuclease dead-Cas9
title Suppression of HBV replication by the expression of nickase- and nuclease dead-Cas9
title_full Suppression of HBV replication by the expression of nickase- and nuclease dead-Cas9
title_fullStr Suppression of HBV replication by the expression of nickase- and nuclease dead-Cas9
title_full_unstemmed Suppression of HBV replication by the expression of nickase- and nuclease dead-Cas9
title_short Suppression of HBV replication by the expression of nickase- and nuclease dead-Cas9
title_sort suppression of hbv replication by the expression of nickase- and nuclease dead-cas9
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5522428/
https://www.ncbi.nlm.nih.gov/pubmed/28733609
http://dx.doi.org/10.1038/s41598-017-05905-w
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