Establishment of a Rapid Detection System for ISG20-Dependent SARS-CoV-2 Subreplicon RNA Degradation Induced by Interferon-α

Inhaled nebulized interferon (IFN)-α and IFN-β have been shown to be effective in the management of coronavirus disease 2019 (COVID-19). We aimed to construct a virus-free rapid detection system for high-throughput screening of IFN-like compounds that induce viral RNA degradation and suppress the re...

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Autores principales:	Yutaka Furutani, Mariko Toguchi, Shoko Higuchi, Kaori Yanaka, Luc Gailhouste, Xian-Yang Qin, Takahiro Masaki, Sae Ochi, Tomokazu Matsuura
Formato:	article
Lenguaje:	EN
Publicado:	MDPI AG 2021
Materias:	COVID-19 replicon interferon interferon-stimulated genes ISG20 CDM-3008 Biology (General) QH301-705.5 Chemistry QD1-999
Acceso en línea:	https://doaj.org/article/66259d5985bd428b9b4e9acf53472578
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spelling	oai:doaj.org-article:66259d5985bd428b9b4e9acf534725782021-11-11T17:06:55ZEstablishment of a Rapid Detection System for ISG20-Dependent SARS-CoV-2 Subreplicon RNA Degradation Induced by Interferon-α10.3390/ijms2221116411422-00671661-6596https://doaj.org/article/66259d5985bd428b9b4e9acf534725782021-10-01T00:00:00Zhttps://www.mdpi.com/1422-0067/22/21/11641https://doaj.org/toc/1661-6596https://doaj.org/toc/1422-0067Inhaled nebulized interferon (IFN)-α and IFN-β have been shown to be effective in the management of coronavirus disease 2019 (COVID-19). We aimed to construct a virus-free rapid detection system for high-throughput screening of IFN-like compounds that induce viral RNA degradation and suppress the replication of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We prepared a SARS-CoV-2 subreplicon RNA expression vector which contained the SARS-CoV-2 5′-UTR, the partial sequence of ORF1a, luciferase, nucleocapsid, ORF10, and 3′-UTR under the control of the cytomegalovirus promoter. The expression vector was transfected into Calu-3 cells and treated with IFN-α and the IFNAR2 agonist CDM-3008 (RO8191) for 3 days. SARS-CoV-2 subreplicon RNA degradation was subsequently evaluated based on luciferase levels. IFN-α and CDM-3008 suppressed SARS-CoV-2 subreplicon RNA in a dose-dependent manner, with IC50 values of 193 IU/mL and 2.54 μM, respectively. HeLa cells stably expressing SARS-CoV-2 subreplicon RNA were prepared and treated with the IFN-α and pan-JAK inhibitor Pyridone 6 or siRNA-targeting ISG20. IFN-α activity was canceled with Pyridone 6. The knockdown of ISG20 partially canceled IFN-α activity. Collectively, we constructed a virus-free rapid detection system to measure SARS-CoV-2 RNA suppression. Our data suggest that the SARS-CoV-2 subreplicon RNA was degraded by IFN-α-induced ISG20 exonuclease activity.Yutaka FurutaniMariko ToguchiShoko HiguchiKaori YanakaLuc GailhousteXian-Yang QinTakahiro MasakiSae OchiTomokazu MatsuuraMDPI AGarticleCOVID-19repliconinterferoninterferon-stimulated genesISG20CDM-3008Biology (General)QH301-705.5ChemistryQD1-999ENInternational Journal of Molecular Sciences, Vol 22, Iss 11641, p 11641 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	COVID-19 replicon interferon interferon-stimulated genes ISG20 CDM-3008 Biology (General) QH301-705.5 Chemistry QD1-999
spellingShingle	COVID-19 replicon interferon interferon-stimulated genes ISG20 CDM-3008 Biology (General) QH301-705.5 Chemistry QD1-999 Yutaka Furutani Mariko Toguchi Shoko Higuchi Kaori Yanaka Luc Gailhouste Xian-Yang Qin Takahiro Masaki Sae Ochi Tomokazu Matsuura Establishment of a Rapid Detection System for ISG20-Dependent SARS-CoV-2 Subreplicon RNA Degradation Induced by Interferon-α
description	Inhaled nebulized interferon (IFN)-α and IFN-β have been shown to be effective in the management of coronavirus disease 2019 (COVID-19). We aimed to construct a virus-free rapid detection system for high-throughput screening of IFN-like compounds that induce viral RNA degradation and suppress the replication of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We prepared a SARS-CoV-2 subreplicon RNA expression vector which contained the SARS-CoV-2 5′-UTR, the partial sequence of ORF1a, luciferase, nucleocapsid, ORF10, and 3′-UTR under the control of the cytomegalovirus promoter. The expression vector was transfected into Calu-3 cells and treated with IFN-α and the IFNAR2 agonist CDM-3008 (RO8191) for 3 days. SARS-CoV-2 subreplicon RNA degradation was subsequently evaluated based on luciferase levels. IFN-α and CDM-3008 suppressed SARS-CoV-2 subreplicon RNA in a dose-dependent manner, with IC50 values of 193 IU/mL and 2.54 μM, respectively. HeLa cells stably expressing SARS-CoV-2 subreplicon RNA were prepared and treated with the IFN-α and pan-JAK inhibitor Pyridone 6 or siRNA-targeting ISG20. IFN-α activity was canceled with Pyridone 6. The knockdown of ISG20 partially canceled IFN-α activity. Collectively, we constructed a virus-free rapid detection system to measure SARS-CoV-2 RNA suppression. Our data suggest that the SARS-CoV-2 subreplicon RNA was degraded by IFN-α-induced ISG20 exonuclease activity.
format	article
author	Yutaka Furutani Mariko Toguchi Shoko Higuchi Kaori Yanaka Luc Gailhouste Xian-Yang Qin Takahiro Masaki Sae Ochi Tomokazu Matsuura
author_facet	Yutaka Furutani Mariko Toguchi Shoko Higuchi Kaori Yanaka Luc Gailhouste Xian-Yang Qin Takahiro Masaki Sae Ochi Tomokazu Matsuura
author_sort	Yutaka Furutani
title	Establishment of a Rapid Detection System for ISG20-Dependent SARS-CoV-2 Subreplicon RNA Degradation Induced by Interferon-α
title_short	Establishment of a Rapid Detection System for ISG20-Dependent SARS-CoV-2 Subreplicon RNA Degradation Induced by Interferon-α
title_full	Establishment of a Rapid Detection System for ISG20-Dependent SARS-CoV-2 Subreplicon RNA Degradation Induced by Interferon-α
title_fullStr	Establishment of a Rapid Detection System for ISG20-Dependent SARS-CoV-2 Subreplicon RNA Degradation Induced by Interferon-α
title_full_unstemmed	Establishment of a Rapid Detection System for ISG20-Dependent SARS-CoV-2 Subreplicon RNA Degradation Induced by Interferon-α
title_sort	establishment of a rapid detection system for isg20-dependent sars-cov-2 subreplicon rna degradation induced by interferon-α
publisher	MDPI AG
publishDate	2021
url	https://doaj.org/article/66259d5985bd428b9b4e9acf53472578
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Establishment of a Rapid Detection System for ISG20-Dependent SARS-CoV-2 Subreplicon RNA Degradation Induced by Interferon-α

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