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Palmitoylation mediates membrane association of hepatitis E virus ORF3 protein and is required for infectious particle secretion

Hepatitis E virus (HEV) is a positive-strand RNA virus encoding 3 open reading frames (ORF). HEV ORF3 protein is a small, hitherto poorly characterized protein involved in viral particle secretion and possibly other functions. Here, we show that HEV ORF3 protein forms membrane-associated oligomers....

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Autores principales: Gouttenoire, Jérôme, Pollán, Angela, Abrami, Laurence, Oechslin, Noémie, Mauron, Johann, Matter, Maxime, Oppliger, Joël, Szkolnicka, Dagmara, Dao Thi, Viet Loan, van der Goot, F. Gisou, Moradpour, Darius
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6307819/
https://www.ncbi.nlm.nih.gov/pubmed/30532200
http://dx.doi.org/10.1371/journal.ppat.1007471
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author Gouttenoire, Jérôme
Pollán, Angela
Abrami, Laurence
Oechslin, Noémie
Mauron, Johann
Matter, Maxime
Oppliger, Joël
Szkolnicka, Dagmara
Dao Thi, Viet Loan
van der Goot, F. Gisou
Moradpour, Darius
author_facet Gouttenoire, Jérôme
Pollán, Angela
Abrami, Laurence
Oechslin, Noémie
Mauron, Johann
Matter, Maxime
Oppliger, Joël
Szkolnicka, Dagmara
Dao Thi, Viet Loan
van der Goot, F. Gisou
Moradpour, Darius
author_sort Gouttenoire, Jérôme
collection PubMed
description Hepatitis E virus (HEV) is a positive-strand RNA virus encoding 3 open reading frames (ORF). HEV ORF3 protein is a small, hitherto poorly characterized protein involved in viral particle secretion and possibly other functions. Here, we show that HEV ORF3 protein forms membrane-associated oligomers. Immunoblot analyses of ORF3 protein expressed in cell-free vs. cellular systems suggested a posttranslational modification. Further analyses revealed that HEV ORF3 protein is palmitoylated at cysteine residues in its N-terminal region, as corroborated by (3)H-palmitate labeling, the investigation of cysteine-to-alanine substitution mutants and treatment with the palmitoylation inhibitor 2-bromopalmitate (2-BP). Abrogation of palmitoylation by site-directed mutagenesis or 2-BP treatment altered the subcellular localization of ORF3 protein, reduced the stability of the protein and strongly impaired the secretion of infectious particles. Moreover, selective membrane permeabilization coupled with immunofluorescence microscopy revealed that HEV ORF3 protein is entirely exposed to the cytosolic side of the membrane, allowing to propose a model for its membrane topology and interactions required in the viral life cycle. In conclusion, palmitoylation determines the subcellular localization, membrane topology and function of HEV ORF3 protein in the HEV life cycle.
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spelling pubmed-63078192019-01-08 Palmitoylation mediates membrane association of hepatitis E virus ORF3 protein and is required for infectious particle secretion Gouttenoire, Jérôme Pollán, Angela Abrami, Laurence Oechslin, Noémie Mauron, Johann Matter, Maxime Oppliger, Joël Szkolnicka, Dagmara Dao Thi, Viet Loan van der Goot, F. Gisou Moradpour, Darius PLoS Pathog Research Article Hepatitis E virus (HEV) is a positive-strand RNA virus encoding 3 open reading frames (ORF). HEV ORF3 protein is a small, hitherto poorly characterized protein involved in viral particle secretion and possibly other functions. Here, we show that HEV ORF3 protein forms membrane-associated oligomers. Immunoblot analyses of ORF3 protein expressed in cell-free vs. cellular systems suggested a posttranslational modification. Further analyses revealed that HEV ORF3 protein is palmitoylated at cysteine residues in its N-terminal region, as corroborated by (3)H-palmitate labeling, the investigation of cysteine-to-alanine substitution mutants and treatment with the palmitoylation inhibitor 2-bromopalmitate (2-BP). Abrogation of palmitoylation by site-directed mutagenesis or 2-BP treatment altered the subcellular localization of ORF3 protein, reduced the stability of the protein and strongly impaired the secretion of infectious particles. Moreover, selective membrane permeabilization coupled with immunofluorescence microscopy revealed that HEV ORF3 protein is entirely exposed to the cytosolic side of the membrane, allowing to propose a model for its membrane topology and interactions required in the viral life cycle. In conclusion, palmitoylation determines the subcellular localization, membrane topology and function of HEV ORF3 protein in the HEV life cycle. Public Library of Science 2018-12-10 /pmc/articles/PMC6307819/ /pubmed/30532200 http://dx.doi.org/10.1371/journal.ppat.1007471 Text en © 2018 Gouttenoire et al http://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the terms of the Creative Commons Attribution License (http://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
Gouttenoire, Jérôme
Pollán, Angela
Abrami, Laurence
Oechslin, Noémie
Mauron, Johann
Matter, Maxime
Oppliger, Joël
Szkolnicka, Dagmara
Dao Thi, Viet Loan
van der Goot, F. Gisou
Moradpour, Darius
Palmitoylation mediates membrane association of hepatitis E virus ORF3 protein and is required for infectious particle secretion
title Palmitoylation mediates membrane association of hepatitis E virus ORF3 protein and is required for infectious particle secretion
title_full Palmitoylation mediates membrane association of hepatitis E virus ORF3 protein and is required for infectious particle secretion
title_fullStr Palmitoylation mediates membrane association of hepatitis E virus ORF3 protein and is required for infectious particle secretion
title_full_unstemmed Palmitoylation mediates membrane association of hepatitis E virus ORF3 protein and is required for infectious particle secretion
title_short Palmitoylation mediates membrane association of hepatitis E virus ORF3 protein and is required for infectious particle secretion
title_sort palmitoylation mediates membrane association of hepatitis e virus orf3 protein and is required for infectious particle secretion
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6307819/
https://www.ncbi.nlm.nih.gov/pubmed/30532200
http://dx.doi.org/10.1371/journal.ppat.1007471
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