Papa, G, Venditti, L, Arnoldi, F et al. (5 more authors) (2020) Recombinant rotaviruses rescued by reverse genetics reveal the role of NSP5 hyperphosphorylation in the assembly of viral factories. Journal of Virology, 94 (1). e01110-19. ISSN 0022-538X
Abstract
Rotavirus (RV) replicates in round-shaped cytoplasmic viral factories, although how they assemble remains unknown.
During RV infection, NSP5 undergoes hyperphosphorylation, which is primed by the phosphorylation of a single serine residue. The role of this post-translational modification in the formation of viroplasms and its impact on the virus replication remain obscure. Here we investigated the role of NSP5 during RV infection by taking advantage of a modified fully tractable reverse genetics system. A trans-complementing cell line stably producing NSP5 was used to generate and characterise several recombinant rotaviruses (rRVs) with mutations in NSP5. We demonstrate that an rRV lacking NSP5 was completely unable to assemble viroplasms and to replicate, confirming its pivotal role in rotavirus replication.
A number of mutants with impaired NSP5 phosphorylation were generated to further interrogate the function of this post-translational modification in the assembly of replication-competent viroplasms. We showed that the rRV mutant strains exhibited impaired viral replication and the ability to assemble round-shaped viroplasms in MA104 cells. Furthermore, we have investigated the mechanism of NSP5 hyper-phosphorylation during RV infection using NSP5 phosphorylation-negative rRV strains, as well as MA104-derived stable transfectant cell lines expressing either wt NSP5 or selected NSP5 deletion mutants. Our results indicate that NSP5 hyper-phosphorylation is a crucial step for the assembly of round-shaped viroplasms, highlighting the key role of the C-terminal tail of NSP5 in the formation of replication-competent viral factories. Such a complex NSP5 phosphorylation cascade may serve as a paradigm for the assembly of functional viral factories in other RNA viruses.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | © 2019 Papa et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license. |
Dates: |
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Institution: | The University of Leeds |
Academic Units: | The University of Leeds > Faculty of Biological Sciences (Leeds) > School of Molecular and Cellular Biology (Leeds) > Bioinformatics (Leeds) |
Funding Information: | Funder Grant number Wellcome Trust 103068/Z/13/Z |
Depositing User: | Symplectic Publications |
Date Deposited: | 04 Oct 2019 09:59 |
Last Modified: | 25 Jun 2023 22:00 |
Status: | Published |
Publisher: | American Society for Microbiology |
Identification Number: | 10.1128/JVI.01110-19 |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:151681 |
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