Cargando…
Ubiquitination of Ebola virus VP35 at lysine 309 regulates viral transcription and assembly
Ebola virus (EBOV) VP35 is a polyfunctional protein involved in viral genome packaging, viral polymerase function, and host immune antagonism. The mechanisms regulating VP35’s engagement in different functions are not well-understood. We previously showed that the host E3 ubiquitin ligase TRIM6 ubiq...
Autores principales: | , , , , , , , , , , , , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2022
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9119628/ https://www.ncbi.nlm.nih.gov/pubmed/35533195 http://dx.doi.org/10.1371/journal.ppat.1010532 |
_version_ | 1784710734412775424 |
---|---|
author | van Tol, Sarah Kalveram, Birte Ilinykh, Philipp A. Ronk, Adam Huang, Kai Aguilera-Aguirre, Leopoldo Bharaj, Preeti Hage, Adam Atkins, Colm Giraldo, Maria I. Wakamiya, Maki Gonzalez-Orozco, Maria Warren, Abbey N. Bukreyev, Alexander Freiberg, Alexander N. Rajsbaum, Ricardo |
author_facet | van Tol, Sarah Kalveram, Birte Ilinykh, Philipp A. Ronk, Adam Huang, Kai Aguilera-Aguirre, Leopoldo Bharaj, Preeti Hage, Adam Atkins, Colm Giraldo, Maria I. Wakamiya, Maki Gonzalez-Orozco, Maria Warren, Abbey N. Bukreyev, Alexander Freiberg, Alexander N. Rajsbaum, Ricardo |
author_sort | van Tol, Sarah |
collection | PubMed |
description | Ebola virus (EBOV) VP35 is a polyfunctional protein involved in viral genome packaging, viral polymerase function, and host immune antagonism. The mechanisms regulating VP35’s engagement in different functions are not well-understood. We previously showed that the host E3 ubiquitin ligase TRIM6 ubiquitinates VP35 at lysine 309 (K309) to facilitate virus replication. However, how K309 ubiquitination regulates the function of VP35 as the viral polymerase co-factor and the precise stage(s) of the EBOV replication cycle that require VP35 ubiquitination are not known. Here, we generated recombinant EBOVs encoding glycine (G) or arginine (R) mutations at VP35/K309 (rEBOV-VP35/K309G/-R) and show that both mutations prohibit VP35/K309 ubiquitination. The K309R mutant retains dsRNA binding and efficient type-I Interferon (IFN-I) antagonism due to the basic residue conservation. The rEBOV-VP35/K309G mutant loses the ability to efficiently antagonize the IFN-I response, while the rEBOV-VP35/K309R mutant’s suppression is enhanced. The replication of both mutants was significantly attenuated in both IFN-competent and -deficient cells due to impaired interactions with the viral polymerase. The lack of ubiquitination on VP35/K309 or TRIM6 deficiency disrupts viral transcription with increasing severity along the transcriptional gradient. This disruption of the transcriptional gradient results in unbalanced viral protein production, including reduced synthesis of the viral transcription factor VP30. In addition, lack of ubiquitination on K309 results in enhanced interactions with the viral nucleoprotein and premature nucleocapsid packaging, leading to dysregulation of virus assembly. Overall, we identified a novel role of VP35 ubiquitination in coordinating viral transcription and assembly. |
format | Online Article Text |
id | pubmed-9119628 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-91196282022-05-20 Ubiquitination of Ebola virus VP35 at lysine 309 regulates viral transcription and assembly van Tol, Sarah Kalveram, Birte Ilinykh, Philipp A. Ronk, Adam Huang, Kai Aguilera-Aguirre, Leopoldo Bharaj, Preeti Hage, Adam Atkins, Colm Giraldo, Maria I. Wakamiya, Maki Gonzalez-Orozco, Maria Warren, Abbey N. Bukreyev, Alexander Freiberg, Alexander N. Rajsbaum, Ricardo PLoS Pathog Research Article Ebola virus (EBOV) VP35 is a polyfunctional protein involved in viral genome packaging, viral polymerase function, and host immune antagonism. The mechanisms regulating VP35’s engagement in different functions are not well-understood. We previously showed that the host E3 ubiquitin ligase TRIM6 ubiquitinates VP35 at lysine 309 (K309) to facilitate virus replication. However, how K309 ubiquitination regulates the function of VP35 as the viral polymerase co-factor and the precise stage(s) of the EBOV replication cycle that require VP35 ubiquitination are not known. Here, we generated recombinant EBOVs encoding glycine (G) or arginine (R) mutations at VP35/K309 (rEBOV-VP35/K309G/-R) and show that both mutations prohibit VP35/K309 ubiquitination. The K309R mutant retains dsRNA binding and efficient type-I Interferon (IFN-I) antagonism due to the basic residue conservation. The rEBOV-VP35/K309G mutant loses the ability to efficiently antagonize the IFN-I response, while the rEBOV-VP35/K309R mutant’s suppression is enhanced. The replication of both mutants was significantly attenuated in both IFN-competent and -deficient cells due to impaired interactions with the viral polymerase. The lack of ubiquitination on VP35/K309 or TRIM6 deficiency disrupts viral transcription with increasing severity along the transcriptional gradient. This disruption of the transcriptional gradient results in unbalanced viral protein production, including reduced synthesis of the viral transcription factor VP30. In addition, lack of ubiquitination on K309 results in enhanced interactions with the viral nucleoprotein and premature nucleocapsid packaging, leading to dysregulation of virus assembly. Overall, we identified a novel role of VP35 ubiquitination in coordinating viral transcription and assembly. Public Library of Science 2022-05-09 /pmc/articles/PMC9119628/ /pubmed/35533195 http://dx.doi.org/10.1371/journal.ppat.1010532 Text en © 2022 van Tol et al https://creativecommons.org/licenses/by/4.0/This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. |
spellingShingle | Research Article van Tol, Sarah Kalveram, Birte Ilinykh, Philipp A. Ronk, Adam Huang, Kai Aguilera-Aguirre, Leopoldo Bharaj, Preeti Hage, Adam Atkins, Colm Giraldo, Maria I. Wakamiya, Maki Gonzalez-Orozco, Maria Warren, Abbey N. Bukreyev, Alexander Freiberg, Alexander N. Rajsbaum, Ricardo Ubiquitination of Ebola virus VP35 at lysine 309 regulates viral transcription and assembly |
title | Ubiquitination of Ebola virus VP35 at lysine 309 regulates viral transcription and assembly |
title_full | Ubiquitination of Ebola virus VP35 at lysine 309 regulates viral transcription and assembly |
title_fullStr | Ubiquitination of Ebola virus VP35 at lysine 309 regulates viral transcription and assembly |
title_full_unstemmed | Ubiquitination of Ebola virus VP35 at lysine 309 regulates viral transcription and assembly |
title_short | Ubiquitination of Ebola virus VP35 at lysine 309 regulates viral transcription and assembly |
title_sort | ubiquitination of ebola virus vp35 at lysine 309 regulates viral transcription and assembly |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9119628/ https://www.ncbi.nlm.nih.gov/pubmed/35533195 http://dx.doi.org/10.1371/journal.ppat.1010532 |
work_keys_str_mv | AT vantolsarah ubiquitinationofebolavirusvp35atlysine309regulatesviraltranscriptionandassembly AT kalverambirte ubiquitinationofebolavirusvp35atlysine309regulatesviraltranscriptionandassembly AT ilinykhphilippa ubiquitinationofebolavirusvp35atlysine309regulatesviraltranscriptionandassembly AT ronkadam ubiquitinationofebolavirusvp35atlysine309regulatesviraltranscriptionandassembly AT huangkai ubiquitinationofebolavirusvp35atlysine309regulatesviraltranscriptionandassembly AT aguileraaguirreleopoldo ubiquitinationofebolavirusvp35atlysine309regulatesviraltranscriptionandassembly AT bharajpreeti ubiquitinationofebolavirusvp35atlysine309regulatesviraltranscriptionandassembly AT hageadam ubiquitinationofebolavirusvp35atlysine309regulatesviraltranscriptionandassembly AT atkinscolm ubiquitinationofebolavirusvp35atlysine309regulatesviraltranscriptionandassembly AT giraldomariai ubiquitinationofebolavirusvp35atlysine309regulatesviraltranscriptionandassembly AT wakamiyamaki ubiquitinationofebolavirusvp35atlysine309regulatesviraltranscriptionandassembly AT gonzalezorozcomaria ubiquitinationofebolavirusvp35atlysine309regulatesviraltranscriptionandassembly AT warrenabbeyn ubiquitinationofebolavirusvp35atlysine309regulatesviraltranscriptionandassembly AT bukreyevalexander ubiquitinationofebolavirusvp35atlysine309regulatesviraltranscriptionandassembly AT freibergalexandern ubiquitinationofebolavirusvp35atlysine309regulatesviraltranscriptionandassembly AT rajsbaumricardo ubiquitinationofebolavirusvp35atlysine309regulatesviraltranscriptionandassembly |