Cargando…
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...
Autores principales: | , , , , , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2021
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8583800/ https://www.ncbi.nlm.nih.gov/pubmed/34769072 http://dx.doi.org/10.3390/ijms222111641 |
_version_ | 1784597291020058624 |
---|---|
author | Furutani, Yutaka Toguchi, Mariko Higuchi, Shoko Yanaka, Kaori Gailhouste, Luc Qin, Xian-Yang Masaki, Takahiro Ochi, Sae Matsuura, Tomokazu |
author_facet | Furutani, Yutaka Toguchi, Mariko Higuchi, Shoko Yanaka, Kaori Gailhouste, Luc Qin, Xian-Yang Masaki, Takahiro Ochi, Sae Matsuura, Tomokazu |
author_sort | Furutani, Yutaka |
collection | PubMed |
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 | Online Article Text |
id | pubmed-8583800 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-85838002021-11-12 Establishment of a Rapid Detection System for ISG20-Dependent SARS-CoV-2 Subreplicon RNA Degradation Induced by Interferon-α Furutani, Yutaka Toguchi, Mariko Higuchi, Shoko Yanaka, Kaori Gailhouste, Luc Qin, Xian-Yang Masaki, Takahiro Ochi, Sae Matsuura, Tomokazu Int J Mol Sci Article 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. MDPI 2021-10-28 /pmc/articles/PMC8583800/ /pubmed/34769072 http://dx.doi.org/10.3390/ijms222111641 Text en © 2021 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Furutani, Yutaka Toguchi, Mariko Higuchi, Shoko Yanaka, Kaori Gailhouste, Luc Qin, Xian-Yang Masaki, Takahiro Ochi, Sae Matsuura, Tomokazu Establishment of a Rapid Detection System for ISG20-Dependent SARS-CoV-2 Subreplicon RNA Degradation Induced by Interferon-α |
title | 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_short | 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 |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8583800/ https://www.ncbi.nlm.nih.gov/pubmed/34769072 http://dx.doi.org/10.3390/ijms222111641 |
work_keys_str_mv | AT furutaniyutaka establishmentofarapiddetectionsystemforisg20dependentsarscov2subrepliconrnadegradationinducedbyinterferona AT toguchimariko establishmentofarapiddetectionsystemforisg20dependentsarscov2subrepliconrnadegradationinducedbyinterferona AT higuchishoko establishmentofarapiddetectionsystemforisg20dependentsarscov2subrepliconrnadegradationinducedbyinterferona AT yanakakaori establishmentofarapiddetectionsystemforisg20dependentsarscov2subrepliconrnadegradationinducedbyinterferona AT gailhousteluc establishmentofarapiddetectionsystemforisg20dependentsarscov2subrepliconrnadegradationinducedbyinterferona AT qinxianyang establishmentofarapiddetectionsystemforisg20dependentsarscov2subrepliconrnadegradationinducedbyinterferona AT masakitakahiro establishmentofarapiddetectionsystemforisg20dependentsarscov2subrepliconrnadegradationinducedbyinterferona AT ochisae establishmentofarapiddetectionsystemforisg20dependentsarscov2subrepliconrnadegradationinducedbyinterferona AT matsuuratomokazu establishmentofarapiddetectionsystemforisg20dependentsarscov2subrepliconrnadegradationinducedbyinterferona |