Hepatitis C Virus Proteins Interact with the Endosomal Sorting Complex Required for Transport (ESCRT) Machinery via Ubiquitination To Facilitate Viral Envelopment

ABSTRACT Enveloped viruses commonly utilize late-domain motifs, sometimes cooperatively with ubiquitin, to hijack the endosomal sorting complex required for transport (ESCRT) machinery for budding at the plasma membrane. However, the mechanisms underlying budding of viruses lacking defined late-doma...

Descripción completa

Guardado en:

Detalles Bibliográficos
Autores principales:	Rina Barouch-Bentov, Gregory Neveu, Fei Xiao, Melanie Beer, Elena Bekerman, Stanford Schor, Joseph Campbell, Jim Boonyaratanakornkit, Brett Lindenbach, Albert Lu, Yves Jacob, Shirit Einav
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2016
Materias:	Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/7638b83bba084ee190baf218083848bc
Etiquetas:	Agregar Etiqueta Sin Etiquetas, Sea el primero en etiquetar este registro!

id	oai:doaj.org-article:7638b83bba084ee190baf218083848bc
record_format	dspace
spelling	oai:doaj.org-article:7638b83bba084ee190baf218083848bc2021-11-15T15:50:16ZHepatitis C Virus Proteins Interact with the Endosomal Sorting Complex Required for Transport (ESCRT) Machinery via Ubiquitination To Facilitate Viral Envelopment10.1128/mBio.01456-162150-7511https://doaj.org/article/7638b83bba084ee190baf218083848bc2016-12-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.01456-16https://doaj.org/toc/2150-7511ABSTRACT Enveloped viruses commonly utilize late-domain motifs, sometimes cooperatively with ubiquitin, to hijack the endosomal sorting complex required for transport (ESCRT) machinery for budding at the plasma membrane. However, the mechanisms underlying budding of viruses lacking defined late-domain motifs and budding into intracellular compartments are poorly characterized. Here, we map a network of hepatitis C virus (HCV) protein interactions with the ESCRT machinery using a mammalian-cell-based protein interaction screen and reveal nine novel interactions. We identify HRS (hepatocyte growth factor-regulated tyrosine kinase substrate), an ESCRT-0 complex component, as an important entry point for HCV into the ESCRT pathway and validate its interactions with the HCV nonstructural (NS) proteins NS2 and NS5A in HCV-infected cells. Infectivity assays indicate that HRS is an important factor for efficient HCV assembly. Specifically, by integrating capsid oligomerization assays, biophysical analysis of intracellular viral particles by continuous gradient centrifugations, proteolytic digestion protection, and RNase digestion protection assays, we show that HCV co-opts HRS to mediate a late assembly step, namely, envelopment. In the absence of defined late-domain motifs, K63-linked polyubiquitinated lysine residues in the HCV NS2 protein bind the HRS ubiquitin-interacting motif to facilitate assembly. Finally, ESCRT-III and VPS/VTA1 components are also recruited by HCV proteins to mediate assembly. These data uncover involvement of ESCRT proteins in intracellular budding of a virus lacking defined late-domain motifs and a novel mechanism by which HCV gains entry into the ESCRT network, with potential implications for other viruses. IMPORTANCE Viruses commonly bud at the plasma membrane by recruiting the host ESCRT machinery via conserved motifs termed late domains. The mechanism by which some viruses, such as HCV, bud intracellularly is, however, poorly characterized. Moreover, whether envelopment of HCV and other viruses lacking defined late domains is ESCRT mediated and, if so, what the entry points into the ESCRT pathway are remain unknown. Here, we report the interaction network of HCV with the ESCRT machinery and a critical role for HRS, an ESCRT-0 complex component, in HCV envelopment. Viral protein ubiquitination was discovered to be a signal for HRS binding and HCV assembly, thereby functionally compensating for the absence of late domains. These findings characterize how a virus lacking defined late domains co-opts ESCRT to bud intracellularly. Since the ESCRT machinery is essential for the life cycle of multiple viruses, better understanding of this virus-host interplay may yield targets for broad-spectrum antiviral therapies.Rina Barouch-BentovGregory NeveuFei XiaoMelanie BeerElena BekermanStanford SchorJoseph CampbellJim BoonyaratanakornkitBrett LindenbachAlbert LuYves JacobShirit EinavAmerican Society for MicrobiologyarticleMicrobiologyQR1-502ENmBio, Vol 7, Iss 6 (2016)
institution	DOAJ
collection	DOAJ
language	EN
topic	Microbiology QR1-502
spellingShingle	Microbiology QR1-502 Rina Barouch-Bentov Gregory Neveu Fei Xiao Melanie Beer Elena Bekerman Stanford Schor Joseph Campbell Jim Boonyaratanakornkit Brett Lindenbach Albert Lu Yves Jacob Shirit Einav Hepatitis C Virus Proteins Interact with the Endosomal Sorting Complex Required for Transport (ESCRT) Machinery via Ubiquitination To Facilitate Viral Envelopment
description	ABSTRACT Enveloped viruses commonly utilize late-domain motifs, sometimes cooperatively with ubiquitin, to hijack the endosomal sorting complex required for transport (ESCRT) machinery for budding at the plasma membrane. However, the mechanisms underlying budding of viruses lacking defined late-domain motifs and budding into intracellular compartments are poorly characterized. Here, we map a network of hepatitis C virus (HCV) protein interactions with the ESCRT machinery using a mammalian-cell-based protein interaction screen and reveal nine novel interactions. We identify HRS (hepatocyte growth factor-regulated tyrosine kinase substrate), an ESCRT-0 complex component, as an important entry point for HCV into the ESCRT pathway and validate its interactions with the HCV nonstructural (NS) proteins NS2 and NS5A in HCV-infected cells. Infectivity assays indicate that HRS is an important factor for efficient HCV assembly. Specifically, by integrating capsid oligomerization assays, biophysical analysis of intracellular viral particles by continuous gradient centrifugations, proteolytic digestion protection, and RNase digestion protection assays, we show that HCV co-opts HRS to mediate a late assembly step, namely, envelopment. In the absence of defined late-domain motifs, K63-linked polyubiquitinated lysine residues in the HCV NS2 protein bind the HRS ubiquitin-interacting motif to facilitate assembly. Finally, ESCRT-III and VPS/VTA1 components are also recruited by HCV proteins to mediate assembly. These data uncover involvement of ESCRT proteins in intracellular budding of a virus lacking defined late-domain motifs and a novel mechanism by which HCV gains entry into the ESCRT network, with potential implications for other viruses. IMPORTANCE Viruses commonly bud at the plasma membrane by recruiting the host ESCRT machinery via conserved motifs termed late domains. The mechanism by which some viruses, such as HCV, bud intracellularly is, however, poorly characterized. Moreover, whether envelopment of HCV and other viruses lacking defined late domains is ESCRT mediated and, if so, what the entry points into the ESCRT pathway are remain unknown. Here, we report the interaction network of HCV with the ESCRT machinery and a critical role for HRS, an ESCRT-0 complex component, in HCV envelopment. Viral protein ubiquitination was discovered to be a signal for HRS binding and HCV assembly, thereby functionally compensating for the absence of late domains. These findings characterize how a virus lacking defined late domains co-opts ESCRT to bud intracellularly. Since the ESCRT machinery is essential for the life cycle of multiple viruses, better understanding of this virus-host interplay may yield targets for broad-spectrum antiviral therapies.
format	article
author	Rina Barouch-Bentov Gregory Neveu Fei Xiao Melanie Beer Elena Bekerman Stanford Schor Joseph Campbell Jim Boonyaratanakornkit Brett Lindenbach Albert Lu Yves Jacob Shirit Einav
author_facet	Rina Barouch-Bentov Gregory Neveu Fei Xiao Melanie Beer Elena Bekerman Stanford Schor Joseph Campbell Jim Boonyaratanakornkit Brett Lindenbach Albert Lu Yves Jacob Shirit Einav
author_sort	Rina Barouch-Bentov
title	Hepatitis C Virus Proteins Interact with the Endosomal Sorting Complex Required for Transport (ESCRT) Machinery via Ubiquitination To Facilitate Viral Envelopment
title_short	Hepatitis C Virus Proteins Interact with the Endosomal Sorting Complex Required for Transport (ESCRT) Machinery via Ubiquitination To Facilitate Viral Envelopment
title_full	Hepatitis C Virus Proteins Interact with the Endosomal Sorting Complex Required for Transport (ESCRT) Machinery via Ubiquitination To Facilitate Viral Envelopment
title_fullStr	Hepatitis C Virus Proteins Interact with the Endosomal Sorting Complex Required for Transport (ESCRT) Machinery via Ubiquitination To Facilitate Viral Envelopment
title_full_unstemmed	Hepatitis C Virus Proteins Interact with the Endosomal Sorting Complex Required for Transport (ESCRT) Machinery via Ubiquitination To Facilitate Viral Envelopment
title_sort	hepatitis c virus proteins interact with the endosomal sorting complex required for transport (escrt) machinery via ubiquitination to facilitate viral envelopment
publisher	American Society for Microbiology
publishDate	2016
url	https://doaj.org/article/7638b83bba084ee190baf218083848bc
work_keys_str_mv	AT rinabarouchbentov hepatitiscvirusproteinsinteractwiththeendosomalsortingcomplexrequiredfortransportescrtmachineryviaubiquitinationtofacilitateviralenvelopment AT gregoryneveu hepatitiscvirusproteinsinteractwiththeendosomalsortingcomplexrequiredfortransportescrtmachineryviaubiquitinationtofacilitateviralenvelopment AT feixiao hepatitiscvirusproteinsinteractwiththeendosomalsortingcomplexrequiredfortransportescrtmachineryviaubiquitinationtofacilitateviralenvelopment AT melaniebeer hepatitiscvirusproteinsinteractwiththeendosomalsortingcomplexrequiredfortransportescrtmachineryviaubiquitinationtofacilitateviralenvelopment AT elenabekerman hepatitiscvirusproteinsinteractwiththeendosomalsortingcomplexrequiredfortransportescrtmachineryviaubiquitinationtofacilitateviralenvelopment AT stanfordschor hepatitiscvirusproteinsinteractwiththeendosomalsortingcomplexrequiredfortransportescrtmachineryviaubiquitinationtofacilitateviralenvelopment AT josephcampbell hepatitiscvirusproteinsinteractwiththeendosomalsortingcomplexrequiredfortransportescrtmachineryviaubiquitinationtofacilitateviralenvelopment AT jimboonyaratanakornkit hepatitiscvirusproteinsinteractwiththeendosomalsortingcomplexrequiredfortransportescrtmachineryviaubiquitinationtofacilitateviralenvelopment AT brettlindenbach hepatitiscvirusproteinsinteractwiththeendosomalsortingcomplexrequiredfortransportescrtmachineryviaubiquitinationtofacilitateviralenvelopment AT albertlu hepatitiscvirusproteinsinteractwiththeendosomalsortingcomplexrequiredfortransportescrtmachineryviaubiquitinationtofacilitateviralenvelopment AT yvesjacob hepatitiscvirusproteinsinteractwiththeendosomalsortingcomplexrequiredfortransportescrtmachineryviaubiquitinationtofacilitateviralenvelopment AT shiriteinav hepatitiscvirusproteinsinteractwiththeendosomalsortingcomplexrequiredfortransportescrtmachineryviaubiquitinationtofacilitateviralenvelopment
_version_	1718427465248931840

Hepatitis C Virus Proteins Interact with the Endosomal Sorting Complex Required for Transport (ESCRT) Machinery via Ubiquitination To Facilitate Viral Envelopment

Ejemplares similares