Cargando…
Computational Design of gRNAs Targeting Genetic Variants Across HIV-1 Subtypes for CRISPR-Mediated Antiviral Therapy
Clustered regularly interspaced short palindromic repeats (CRISPR)-based HIV-1 genome editing has shown promising outcomes in in vitro and in vivo viral infection models. However, existing HIV-1 sequence variants have been shown to reduce CRISPR-mediated efficiency and induce viral escape. Two metri...
Autores principales: | , , , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Frontiers Media S.A.
2021
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7985454/ https://www.ncbi.nlm.nih.gov/pubmed/33768011 http://dx.doi.org/10.3389/fcimb.2021.593077 |
_version_ | 1783668249604390912 |
---|---|
author | Chung, Cheng-Han Allen, Alexander G. Atkins, Andrew Link, Robert W. Nonnemacher, Michael R. Dampier, Will Wigdahl, Brian |
author_facet | Chung, Cheng-Han Allen, Alexander G. Atkins, Andrew Link, Robert W. Nonnemacher, Michael R. Dampier, Will Wigdahl, Brian |
author_sort | Chung, Cheng-Han |
collection | PubMed |
description | Clustered regularly interspaced short palindromic repeats (CRISPR)-based HIV-1 genome editing has shown promising outcomes in in vitro and in vivo viral infection models. However, existing HIV-1 sequence variants have been shown to reduce CRISPR-mediated efficiency and induce viral escape. Two metrics, global patient coverage and global subtype coverage, were used to identify guide RNA (gRNA) sequences that account for this viral diversity from the perspectives of cross-patient and cross-subtype gRNA design, respectively. Computational evaluation using these parameters and over 3.6 million possible 20-bp sequences resulted in nine lead gRNAs, two of which were previously published. This analysis revealed the benefit and necessity of considering all sequence variants for gRNA design. Of the other seven identified novel gRNAs, two were of note as they targeted interesting functional regions. One was a gRNA predicted to induce structural disruption in the nucleocapsid binding site (Ψ), which holds the potential to stop HIV-1 replication during the viral genome packaging process. The other was a reverse transcriptase (RT)-targeting gRNA that was predicted to cleave the subdomain responsible for dNTP incorporation. CRISPR-mediated sequence edits were predicted to occur on critical residues where HIV-1 has been shown to develop resistance against antiretroviral therapy (ART), which may provide additional evolutionary pressure at the DNA level. Given these observations, consideration of broad-spectrum gRNAs and cross-subtype diversity for gRNA design is not only required for the development of generalizable CRISPR-based HIV-1 therapy, but also helps identify optimal target sites. |
format | Online Article Text |
id | pubmed-7985454 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | Frontiers Media S.A. |
record_format | MEDLINE/PubMed |
spelling | pubmed-79854542021-03-24 Computational Design of gRNAs Targeting Genetic Variants Across HIV-1 Subtypes for CRISPR-Mediated Antiviral Therapy Chung, Cheng-Han Allen, Alexander G. Atkins, Andrew Link, Robert W. Nonnemacher, Michael R. Dampier, Will Wigdahl, Brian Front Cell Infect Microbiol Cellular and Infection Microbiology Clustered regularly interspaced short palindromic repeats (CRISPR)-based HIV-1 genome editing has shown promising outcomes in in vitro and in vivo viral infection models. However, existing HIV-1 sequence variants have been shown to reduce CRISPR-mediated efficiency and induce viral escape. Two metrics, global patient coverage and global subtype coverage, were used to identify guide RNA (gRNA) sequences that account for this viral diversity from the perspectives of cross-patient and cross-subtype gRNA design, respectively. Computational evaluation using these parameters and over 3.6 million possible 20-bp sequences resulted in nine lead gRNAs, two of which were previously published. This analysis revealed the benefit and necessity of considering all sequence variants for gRNA design. Of the other seven identified novel gRNAs, two were of note as they targeted interesting functional regions. One was a gRNA predicted to induce structural disruption in the nucleocapsid binding site (Ψ), which holds the potential to stop HIV-1 replication during the viral genome packaging process. The other was a reverse transcriptase (RT)-targeting gRNA that was predicted to cleave the subdomain responsible for dNTP incorporation. CRISPR-mediated sequence edits were predicted to occur on critical residues where HIV-1 has been shown to develop resistance against antiretroviral therapy (ART), which may provide additional evolutionary pressure at the DNA level. Given these observations, consideration of broad-spectrum gRNAs and cross-subtype diversity for gRNA design is not only required for the development of generalizable CRISPR-based HIV-1 therapy, but also helps identify optimal target sites. Frontiers Media S.A. 2021-03-09 /pmc/articles/PMC7985454/ /pubmed/33768011 http://dx.doi.org/10.3389/fcimb.2021.593077 Text en Copyright © 2021 Chung, Allen, Atkins, Link, Nonnemacher, Dampier and Wigdahl http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. |
spellingShingle | Cellular and Infection Microbiology Chung, Cheng-Han Allen, Alexander G. Atkins, Andrew Link, Robert W. Nonnemacher, Michael R. Dampier, Will Wigdahl, Brian Computational Design of gRNAs Targeting Genetic Variants Across HIV-1 Subtypes for CRISPR-Mediated Antiviral Therapy |
title | Computational Design of gRNAs Targeting Genetic Variants Across HIV-1 Subtypes for CRISPR-Mediated Antiviral Therapy |
title_full | Computational Design of gRNAs Targeting Genetic Variants Across HIV-1 Subtypes for CRISPR-Mediated Antiviral Therapy |
title_fullStr | Computational Design of gRNAs Targeting Genetic Variants Across HIV-1 Subtypes for CRISPR-Mediated Antiviral Therapy |
title_full_unstemmed | Computational Design of gRNAs Targeting Genetic Variants Across HIV-1 Subtypes for CRISPR-Mediated Antiviral Therapy |
title_short | Computational Design of gRNAs Targeting Genetic Variants Across HIV-1 Subtypes for CRISPR-Mediated Antiviral Therapy |
title_sort | computational design of grnas targeting genetic variants across hiv-1 subtypes for crispr-mediated antiviral therapy |
topic | Cellular and Infection Microbiology |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7985454/ https://www.ncbi.nlm.nih.gov/pubmed/33768011 http://dx.doi.org/10.3389/fcimb.2021.593077 |
work_keys_str_mv | AT chungchenghan computationaldesignofgrnastargetinggeneticvariantsacrosshiv1subtypesforcrisprmediatedantiviraltherapy AT allenalexanderg computationaldesignofgrnastargetinggeneticvariantsacrosshiv1subtypesforcrisprmediatedantiviraltherapy AT atkinsandrew computationaldesignofgrnastargetinggeneticvariantsacrosshiv1subtypesforcrisprmediatedantiviraltherapy AT linkrobertw computationaldesignofgrnastargetinggeneticvariantsacrosshiv1subtypesforcrisprmediatedantiviraltherapy AT nonnemachermichaelr computationaldesignofgrnastargetinggeneticvariantsacrosshiv1subtypesforcrisprmediatedantiviraltherapy AT dampierwill computationaldesignofgrnastargetinggeneticvariantsacrosshiv1subtypesforcrisprmediatedantiviraltherapy AT wigdahlbrian computationaldesignofgrnastargetinggeneticvariantsacrosshiv1subtypesforcrisprmediatedantiviraltherapy |