CRISPR-Cas13a mediated targeting of hepatitis C virus internal-ribosomal entry site (IRES) as an effective antiviral strategy
Hepatitis C is an inflammatory liver disease caused by the single-stranded RNA (ssRNA) hepatitis C virus (HCV). The genetic diversity of the virus and quasispecies produced during replication have resulted in viral resistance to direct-acting antivirals (DAAs) as well as impediments in vaccine devel...
Main Authors: | , , , , , , , |
---|---|
Format: | Article |
Language: | English |
Published: |
Elsevier
2021-04-01
|
Series: | Biomedicine & Pharmacotherapy |
Subjects: | |
Online Access: | http://www.sciencedirect.com/science/article/pii/S075333222100024X |
id |
doaj-29e8d4c53eda4b47b6a3658c7d4bd7a7 |
---|---|
record_format |
Article |
spelling |
doaj-29e8d4c53eda4b47b6a3658c7d4bd7a72021-07-17T04:32:20ZengElsevierBiomedicine & Pharmacotherapy0753-33222021-04-01136111239CRISPR-Cas13a mediated targeting of hepatitis C virus internal-ribosomal entry site (IRES) as an effective antiviral strategyMuhammad Usman Ashraf0Hafiz Muhammad Salman1Muhammad Farhan Khalid2Muhammad Haider Farooq Khan3Saima Anwar4Samia Afzal5Muhammad Idrees6Safee Ullah Chaudhary7Biomedical Informatics Research Laboratory, Department of Biology, Lahore University of Management Sciences, Lahore, Pakistan; Center of Excellence in Molecular Biology, University of the Punjab, Lahore, PakistanBiomedical Informatics Research Laboratory, Department of Biology, Lahore University of Management Sciences, Lahore, PakistanBiomedical Informatics Research Laboratory, Department of Biology, Lahore University of Management Sciences, Lahore, PakistanBiomedical Informatics Research Laboratory, Department of Biology, Lahore University of Management Sciences, Lahore, PakistanDepartment of Biomedical Engineering, University of Engineering and Technology, Lahore, PakistanCenter of Excellence in Molecular Biology, University of the Punjab, Lahore, PakistanCenter of Excellence in Molecular Biology, University of the Punjab, Lahore, PakistanBiomedical Informatics Research Laboratory, Department of Biology, Lahore University of Management Sciences, Lahore, Pakistan; Corresponding author.Hepatitis C is an inflammatory liver disease caused by the single-stranded RNA (ssRNA) hepatitis C virus (HCV). The genetic diversity of the virus and quasispecies produced during replication have resulted in viral resistance to direct-acting antivirals (DAAs) as well as impediments in vaccine development. The recent adaptation of CRISPR-Cas as an alternative antiviral approach has demonstrated degradation of viral nucleic acids in eukaryotes. In particular, the CRISPR-effector Cas13 enzyme has been shown to target ssRNA viruses effectively. In this work, we have employed Cas13a to knockdown HCV in mammalian cells. Using a computational screen, we identified several potential Cas13a target sites within highly conserved regions of the HCV internal ribosomal entry site (IRES). Our results demonstrate significant inhibition of HCV replication as well as translation in huh-7.5 cells with minimal effects on cell viability. These findings were validated using a multi-modality approach involving qRT-PCR, luciferase assay, and MTT cell viability assay. In conclusion, the CRISPR-Cas13a system efficiently targets HCV in vitro, suggesting its potential as a programmable therapeutic antiviral strategy.http://www.sciencedirect.com/science/article/pii/S075333222100024XHCV IRES inhibitionCRISPR Cas13CRISPRiAntiviral treatment |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Muhammad Usman Ashraf Hafiz Muhammad Salman Muhammad Farhan Khalid Muhammad Haider Farooq Khan Saima Anwar Samia Afzal Muhammad Idrees Safee Ullah Chaudhary |
spellingShingle |
Muhammad Usman Ashraf Hafiz Muhammad Salman Muhammad Farhan Khalid Muhammad Haider Farooq Khan Saima Anwar Samia Afzal Muhammad Idrees Safee Ullah Chaudhary CRISPR-Cas13a mediated targeting of hepatitis C virus internal-ribosomal entry site (IRES) as an effective antiviral strategy Biomedicine & Pharmacotherapy HCV IRES inhibition CRISPR Cas13 CRISPRi Antiviral treatment |
author_facet |
Muhammad Usman Ashraf Hafiz Muhammad Salman Muhammad Farhan Khalid Muhammad Haider Farooq Khan Saima Anwar Samia Afzal Muhammad Idrees Safee Ullah Chaudhary |
author_sort |
Muhammad Usman Ashraf |
title |
CRISPR-Cas13a mediated targeting of hepatitis C virus internal-ribosomal entry site (IRES) as an effective antiviral strategy |
title_short |
CRISPR-Cas13a mediated targeting of hepatitis C virus internal-ribosomal entry site (IRES) as an effective antiviral strategy |
title_full |
CRISPR-Cas13a mediated targeting of hepatitis C virus internal-ribosomal entry site (IRES) as an effective antiviral strategy |
title_fullStr |
CRISPR-Cas13a mediated targeting of hepatitis C virus internal-ribosomal entry site (IRES) as an effective antiviral strategy |
title_full_unstemmed |
CRISPR-Cas13a mediated targeting of hepatitis C virus internal-ribosomal entry site (IRES) as an effective antiviral strategy |
title_sort |
crispr-cas13a mediated targeting of hepatitis c virus internal-ribosomal entry site (ires) as an effective antiviral strategy |
publisher |
Elsevier |
series |
Biomedicine & Pharmacotherapy |
issn |
0753-3322 |
publishDate |
2021-04-01 |
description |
Hepatitis C is an inflammatory liver disease caused by the single-stranded RNA (ssRNA) hepatitis C virus (HCV). The genetic diversity of the virus and quasispecies produced during replication have resulted in viral resistance to direct-acting antivirals (DAAs) as well as impediments in vaccine development. The recent adaptation of CRISPR-Cas as an alternative antiviral approach has demonstrated degradation of viral nucleic acids in eukaryotes. In particular, the CRISPR-effector Cas13 enzyme has been shown to target ssRNA viruses effectively. In this work, we have employed Cas13a to knockdown HCV in mammalian cells. Using a computational screen, we identified several potential Cas13a target sites within highly conserved regions of the HCV internal ribosomal entry site (IRES). Our results demonstrate significant inhibition of HCV replication as well as translation in huh-7.5 cells with minimal effects on cell viability. These findings were validated using a multi-modality approach involving qRT-PCR, luciferase assay, and MTT cell viability assay. In conclusion, the CRISPR-Cas13a system efficiently targets HCV in vitro, suggesting its potential as a programmable therapeutic antiviral strategy. |
topic |
HCV IRES inhibition CRISPR Cas13 CRISPRi Antiviral treatment |
url |
http://www.sciencedirect.com/science/article/pii/S075333222100024X |
work_keys_str_mv |
AT muhammadusmanashraf crisprcas13amediatedtargetingofhepatitiscvirusinternalribosomalentrysiteiresasaneffectiveantiviralstrategy AT hafizmuhammadsalman crisprcas13amediatedtargetingofhepatitiscvirusinternalribosomalentrysiteiresasaneffectiveantiviralstrategy AT muhammadfarhankhalid crisprcas13amediatedtargetingofhepatitiscvirusinternalribosomalentrysiteiresasaneffectiveantiviralstrategy AT muhammadhaiderfarooqkhan crisprcas13amediatedtargetingofhepatitiscvirusinternalribosomalentrysiteiresasaneffectiveantiviralstrategy AT saimaanwar crisprcas13amediatedtargetingofhepatitiscvirusinternalribosomalentrysiteiresasaneffectiveantiviralstrategy AT samiaafzal crisprcas13amediatedtargetingofhepatitiscvirusinternalribosomalentrysiteiresasaneffectiveantiviralstrategy AT muhammadidrees crisprcas13amediatedtargetingofhepatitiscvirusinternalribosomalentrysiteiresasaneffectiveantiviralstrategy AT safeeullahchaudhary crisprcas13amediatedtargetingofhepatitiscvirusinternalribosomalentrysiteiresasaneffectiveantiviralstrategy |
_version_ |
1721297060852924416 |