Cargando…

Point Mutations in the Glycoprotein Ectodomain of Field Rabies Viruses Mediate Cell Culture Adaptation through Improved Virus Release in a Host Cell Dependent and Independent Manner

Molecular details of field rabies virus (RABV) adaptation to cell culture replication are insufficiently understood. A better understanding of adaptation may not only reveal requirements for efficient RABV replication in cell lines, but may also provide novel insights into RABV biology and adaptatio...

Descripción completa

Detalles Bibliográficos
Autores principales: Nitschel, Sabine, Zaeck, Luca M., Potratz, Madlin, Nolden, Tobias, te Kamp, Verena, Franzke, Kati, Höper, Dirk, Pfaff, Florian, Finke, Stefan
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8538267/
https://www.ncbi.nlm.nih.gov/pubmed/34696419
http://dx.doi.org/10.3390/v13101989
_version_ 1784588464531963904
author Nitschel, Sabine
Zaeck, Luca M.
Potratz, Madlin
Nolden, Tobias
te Kamp, Verena
Franzke, Kati
Höper, Dirk
Pfaff, Florian
Finke, Stefan
author_facet Nitschel, Sabine
Zaeck, Luca M.
Potratz, Madlin
Nolden, Tobias
te Kamp, Verena
Franzke, Kati
Höper, Dirk
Pfaff, Florian
Finke, Stefan
author_sort Nitschel, Sabine
collection PubMed
description Molecular details of field rabies virus (RABV) adaptation to cell culture replication are insufficiently understood. A better understanding of adaptation may not only reveal requirements for efficient RABV replication in cell lines, but may also provide novel insights into RABV biology and adaptation-related loss of virulence and pathogenicity. Using two recombinant field rabies virus clones (rRABV Dog and rRABV Fox), we performed virus passages in three different cell lines to identify cell culture adaptive mutations. Ten passages were sufficient for the acquisition of adaptive mutations in the glycoprotein G and in the C-terminus of phosphoprotein P. Apart from the insertion of a glycosylation sequon via the mutation D247N in either virus, both acquired additional and cell line-specific mutations after passages on BHK (K425N) and MDCK-II (R346S or R350G) cells. As determined by virus replication kinetics, complementation, and immunofluorescence analysis, the major bottleneck in cell culture replication was the intracellular accumulation of field virus G protein, which was overcome after the acquisition of the adaptive mutations. Our data indicate that limited release of extracellular infectious virus at the plasma membrane is a defined characteristic of highly virulent field rabies viruses and we hypothesize that the observed suboptimal release of infectious virions is due to the inverse correlation of virus release and virulence in vivo.
format Online
Article
Text
id pubmed-8538267
institution National Center for Biotechnology Information
language English
publishDate 2021
publisher MDPI
record_format MEDLINE/PubMed
spelling pubmed-85382672021-10-24 Point Mutations in the Glycoprotein Ectodomain of Field Rabies Viruses Mediate Cell Culture Adaptation through Improved Virus Release in a Host Cell Dependent and Independent Manner Nitschel, Sabine Zaeck, Luca M. Potratz, Madlin Nolden, Tobias te Kamp, Verena Franzke, Kati Höper, Dirk Pfaff, Florian Finke, Stefan Viruses Article Molecular details of field rabies virus (RABV) adaptation to cell culture replication are insufficiently understood. A better understanding of adaptation may not only reveal requirements for efficient RABV replication in cell lines, but may also provide novel insights into RABV biology and adaptation-related loss of virulence and pathogenicity. Using two recombinant field rabies virus clones (rRABV Dog and rRABV Fox), we performed virus passages in three different cell lines to identify cell culture adaptive mutations. Ten passages were sufficient for the acquisition of adaptive mutations in the glycoprotein G and in the C-terminus of phosphoprotein P. Apart from the insertion of a glycosylation sequon via the mutation D247N in either virus, both acquired additional and cell line-specific mutations after passages on BHK (K425N) and MDCK-II (R346S or R350G) cells. As determined by virus replication kinetics, complementation, and immunofluorescence analysis, the major bottleneck in cell culture replication was the intracellular accumulation of field virus G protein, which was overcome after the acquisition of the adaptive mutations. Our data indicate that limited release of extracellular infectious virus at the plasma membrane is a defined characteristic of highly virulent field rabies viruses and we hypothesize that the observed suboptimal release of infectious virions is due to the inverse correlation of virus release and virulence in vivo. MDPI 2021-10-03 /pmc/articles/PMC8538267/ /pubmed/34696419 http://dx.doi.org/10.3390/v13101989 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
Nitschel, Sabine
Zaeck, Luca M.
Potratz, Madlin
Nolden, Tobias
te Kamp, Verena
Franzke, Kati
Höper, Dirk
Pfaff, Florian
Finke, Stefan
Point Mutations in the Glycoprotein Ectodomain of Field Rabies Viruses Mediate Cell Culture Adaptation through Improved Virus Release in a Host Cell Dependent and Independent Manner
title Point Mutations in the Glycoprotein Ectodomain of Field Rabies Viruses Mediate Cell Culture Adaptation through Improved Virus Release in a Host Cell Dependent and Independent Manner
title_full Point Mutations in the Glycoprotein Ectodomain of Field Rabies Viruses Mediate Cell Culture Adaptation through Improved Virus Release in a Host Cell Dependent and Independent Manner
title_fullStr Point Mutations in the Glycoprotein Ectodomain of Field Rabies Viruses Mediate Cell Culture Adaptation through Improved Virus Release in a Host Cell Dependent and Independent Manner
title_full_unstemmed Point Mutations in the Glycoprotein Ectodomain of Field Rabies Viruses Mediate Cell Culture Adaptation through Improved Virus Release in a Host Cell Dependent and Independent Manner
title_short Point Mutations in the Glycoprotein Ectodomain of Field Rabies Viruses Mediate Cell Culture Adaptation through Improved Virus Release in a Host Cell Dependent and Independent Manner
title_sort point mutations in the glycoprotein ectodomain of field rabies viruses mediate cell culture adaptation through improved virus release in a host cell dependent and independent manner
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8538267/
https://www.ncbi.nlm.nih.gov/pubmed/34696419
http://dx.doi.org/10.3390/v13101989
work_keys_str_mv AT nitschelsabine pointmutationsintheglycoproteinectodomainoffieldrabiesvirusesmediatecellcultureadaptationthroughimprovedvirusreleaseinahostcelldependentandindependentmanner
AT zaecklucam pointmutationsintheglycoproteinectodomainoffieldrabiesvirusesmediatecellcultureadaptationthroughimprovedvirusreleaseinahostcelldependentandindependentmanner
AT potratzmadlin pointmutationsintheglycoproteinectodomainoffieldrabiesvirusesmediatecellcultureadaptationthroughimprovedvirusreleaseinahostcelldependentandindependentmanner
AT noldentobias pointmutationsintheglycoproteinectodomainoffieldrabiesvirusesmediatecellcultureadaptationthroughimprovedvirusreleaseinahostcelldependentandindependentmanner
AT tekampverena pointmutationsintheglycoproteinectodomainoffieldrabiesvirusesmediatecellcultureadaptationthroughimprovedvirusreleaseinahostcelldependentandindependentmanner
AT franzkekati pointmutationsintheglycoproteinectodomainoffieldrabiesvirusesmediatecellcultureadaptationthroughimprovedvirusreleaseinahostcelldependentandindependentmanner
AT hoperdirk pointmutationsintheglycoproteinectodomainoffieldrabiesvirusesmediatecellcultureadaptationthroughimprovedvirusreleaseinahostcelldependentandindependentmanner
AT pfaffflorian pointmutationsintheglycoproteinectodomainoffieldrabiesvirusesmediatecellcultureadaptationthroughimprovedvirusreleaseinahostcelldependentandindependentmanner
AT finkestefan pointmutationsintheglycoproteinectodomainoffieldrabiesvirusesmediatecellcultureadaptationthroughimprovedvirusreleaseinahostcelldependentandindependentmanner