Cargando…

Genomic stability of self-inactivating rabies

Transsynaptic viral vectors provide means to gain genetic access to neurons based on synaptic connectivity and are essential tools for the dissection of neural circuit function. Among them, the retrograde monosynaptic ΔG-Rabies has been widely used in neuroscience research. A recently developed engi...

Descripción completa

Detalles Bibliográficos
Autores principales: Ciabatti, Ernesto, González-Rueda, Ana, de Malmazet, Daniel, Lee, Hassal, Morgese, Fabio, Tripodi, Marco
Formato: Online Artículo Texto
Lenguaje:English
Publicado: eLife Sciences Publications, Ltd 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10666929/
https://www.ncbi.nlm.nih.gov/pubmed/37921437
http://dx.doi.org/10.7554/eLife.83459
_version_ 1785139148014747648
author Ciabatti, Ernesto
González-Rueda, Ana
de Malmazet, Daniel
Lee, Hassal
Morgese, Fabio
Tripodi, Marco
author_facet Ciabatti, Ernesto
González-Rueda, Ana
de Malmazet, Daniel
Lee, Hassal
Morgese, Fabio
Tripodi, Marco
author_sort Ciabatti, Ernesto
collection PubMed
description Transsynaptic viral vectors provide means to gain genetic access to neurons based on synaptic connectivity and are essential tools for the dissection of neural circuit function. Among them, the retrograde monosynaptic ΔG-Rabies has been widely used in neuroscience research. A recently developed engineered version of the ΔG-Rabies, the non-toxic self-inactivating (SiR) virus, allows the long term genetic manipulation of neural circuits. However, the high mutational rate of the rabies virus poses a risk that mutations targeting the key genetic regulatory element in the SiR genome could emerge and revert it to a canonical ΔG-Rabies. Such revertant mutations have recently been identified in a SiR batch. To address the origin, incidence and relevance of these mutations, we investigated the genomic stability of SiR in vitro and in vivo. We found that “revertant” mutations are rare and accumulate only when SiR is extensively amplified in vitro, particularly in suboptimal production cell lines that have insufficient levels of TEV protease activity. Moreover, we confirmed that SiR-CRE, unlike canonical ΔG-Rab-CRE or revertant-SiR-CRE, is non-toxic and that revertant mutations do not emerge in vivo during long-term experiments.
format Online
Article
Text
id pubmed-10666929
institution National Center for Biotechnology Information
language English
publishDate 2023
publisher eLife Sciences Publications, Ltd
record_format MEDLINE/PubMed
spelling pubmed-106669292023-11-03 Genomic stability of self-inactivating rabies Ciabatti, Ernesto González-Rueda, Ana de Malmazet, Daniel Lee, Hassal Morgese, Fabio Tripodi, Marco eLife Genetics and Genomics Transsynaptic viral vectors provide means to gain genetic access to neurons based on synaptic connectivity and are essential tools for the dissection of neural circuit function. Among them, the retrograde monosynaptic ΔG-Rabies has been widely used in neuroscience research. A recently developed engineered version of the ΔG-Rabies, the non-toxic self-inactivating (SiR) virus, allows the long term genetic manipulation of neural circuits. However, the high mutational rate of the rabies virus poses a risk that mutations targeting the key genetic regulatory element in the SiR genome could emerge and revert it to a canonical ΔG-Rabies. Such revertant mutations have recently been identified in a SiR batch. To address the origin, incidence and relevance of these mutations, we investigated the genomic stability of SiR in vitro and in vivo. We found that “revertant” mutations are rare and accumulate only when SiR is extensively amplified in vitro, particularly in suboptimal production cell lines that have insufficient levels of TEV protease activity. Moreover, we confirmed that SiR-CRE, unlike canonical ΔG-Rab-CRE or revertant-SiR-CRE, is non-toxic and that revertant mutations do not emerge in vivo during long-term experiments. eLife Sciences Publications, Ltd 2023-11-03 /pmc/articles/PMC10666929/ /pubmed/37921437 http://dx.doi.org/10.7554/eLife.83459 Text en © 2023, Ciabatti et al https://creativecommons.org/licenses/by/4.0/This article is distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use and redistribution provided that the original author and source are credited.
spellingShingle Genetics and Genomics
Ciabatti, Ernesto
González-Rueda, Ana
de Malmazet, Daniel
Lee, Hassal
Morgese, Fabio
Tripodi, Marco
Genomic stability of self-inactivating rabies
title Genomic stability of self-inactivating rabies
title_full Genomic stability of self-inactivating rabies
title_fullStr Genomic stability of self-inactivating rabies
title_full_unstemmed Genomic stability of self-inactivating rabies
title_short Genomic stability of self-inactivating rabies
title_sort genomic stability of self-inactivating rabies
topic Genetics and Genomics
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10666929/
https://www.ncbi.nlm.nih.gov/pubmed/37921437
http://dx.doi.org/10.7554/eLife.83459
work_keys_str_mv AT ciabattiernesto genomicstabilityofselfinactivatingrabies
AT gonzalezruedaana genomicstabilityofselfinactivatingrabies
AT demalmazetdaniel genomicstabilityofselfinactivatingrabies
AT leehassal genomicstabilityofselfinactivatingrabies
AT morgesefabio genomicstabilityofselfinactivatingrabies
AT tripodimarco genomicstabilityofselfinactivatingrabies