Determinants for Membrane Association of the Hepatitis C Virus RNA-dependent RNA Polymerase
The hepatitis C virus (HCV) RNA-dependent RNA polymerase (RdRp), represented by nonstructural protein 5B (NS5B), is believed to form a membrane-associated RNA replication complex together with other nonstructural proteins and as yet unidentified host components. However, the determinants for membran...
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| Veröffentlicht in: | The Journal of biological chemistry 2001-11, Vol.276 (47), p.44052-44063 |
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| creator | Schmidt-Mende, Juliane Bieck, Elke Hügle, Thomas Penin, François Rice, Charles M. Blum, Hubert E. Moradpour, Darius |
| description | The hepatitis C virus (HCV) RNA-dependent RNA polymerase (RdRp), represented by nonstructural protein 5B (NS5B), is believed to form a membrane-associated RNA replication complex together with other nonstructural proteins and as yet unidentified host components. However, the determinants for membrane association of this essential viral enzyme have not been defined. By double label immunofluorescence analyses, NS5B was found in the endoplasmic reticulum (ER) or an ER-like modified compartment both when expressed alone or in the context of the entire HCV polyprotein. The carboxyl-terminal 21 amino acid residues were necessary and sufficient to target NS5B or a heterologous protein to the cytosolic side of the ER membrane. This hydrophobic domain is highly conserved among 269 HCV isolates analyzed and predicted to form a transmembrane α-helix. Association of NS5B with the ER membrane occurred by a posttranslational mechanism that was ATP-independent. These features define the HCV RdRp as a new member of the tail-anchored protein family, a class of integral membrane proteins that are membrane-targeted posttranslationally via a carboxyl-terminal insertion sequence. Formation of the HCV replication complex, therefore, involves specific determinants for membrane association that represent potential targets for antiviral intervention. |
| doi_str_mv | 10.1074/jbc.M103358200 |
| format | Article |
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However, the determinants for membrane association of this essential viral enzyme have not been defined. By double label immunofluorescence analyses, NS5B was found in the endoplasmic reticulum (ER) or an ER-like modified compartment both when expressed alone or in the context of the entire HCV polyprotein. The carboxyl-terminal 21 amino acid residues were necessary and sufficient to target NS5B or a heterologous protein to the cytosolic side of the ER membrane. This hydrophobic domain is highly conserved among 269 HCV isolates analyzed and predicted to form a transmembrane α-helix. Association of NS5B with the ER membrane occurred by a posttranslational mechanism that was ATP-independent. These features define the HCV RdRp as a new member of the tail-anchored protein family, a class of integral membrane proteins that are membrane-targeted posttranslationally via a carboxyl-terminal insertion sequence. Formation of the HCV replication complex, therefore, involves specific determinants for membrane association that represent potential targets for antiviral intervention.</description><identifier>ISSN: 0021-9258</identifier><identifier>EISSN: 1083-351X</identifier><identifier>DOI: 10.1074/jbc.M103358200</identifier><identifier>PMID: 11557752</identifier><language>eng</language><publisher>United States: Elsevier Inc</publisher><subject>Amino Acid Sequence ; Base Sequence ; Biochemistry, Molecular Biology ; Cell Line ; Cell Membrane - metabolism ; Cell Membrane - virology ; DNA Primers ; Endoplasmic Reticulum - metabolism ; Fluorescent Antibody Technique ; Hepacivirus - enzymology ; Hepatitis C virus ; Humans ; Life Sciences ; Molecular Sequence Data ; NS5B protein ; Protein Biosynthesis ; RNA-Dependent RNA Polymerase - metabolism ; Sequence Homology, Amino Acid ; Tetracycline - pharmacology ; Transcription, Genetic ; Viral Nonstructural Proteins - chemistry ; Viral Nonstructural Proteins - metabolism</subject><ispartof>The Journal of biological chemistry, 2001-11, Vol.276 (47), p.44052-44063</ispartof><rights>2001 © 2001 ASBMB. Currently published by Elsevier Inc; originally published by American Society for Biochemistry and Molecular Biology.</rights><rights>Distributed under a Creative Commons Attribution 4.0 International License</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c540t-f9afc548403b714cb0792a3370633feca8825cad1864162d157fb8b69304b94e3</citedby><cites>FETCH-LOGICAL-c540t-f9afc548403b714cb0792a3370633feca8825cad1864162d157fb8b69304b94e3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>230,314,780,784,885,27924,27925</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/11557752$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink><backlink>$$Uhttps://hal.science/hal-00313707$$DView record in HAL$$Hfree_for_read</backlink></links><search><creatorcontrib>Schmidt-Mende, Juliane</creatorcontrib><creatorcontrib>Bieck, Elke</creatorcontrib><creatorcontrib>Hügle, Thomas</creatorcontrib><creatorcontrib>Penin, François</creatorcontrib><creatorcontrib>Rice, Charles M.</creatorcontrib><creatorcontrib>Blum, Hubert E.</creatorcontrib><creatorcontrib>Moradpour, Darius</creatorcontrib><title>Determinants for Membrane Association of the Hepatitis C Virus RNA-dependent RNA Polymerase</title><title>The Journal of biological chemistry</title><addtitle>J Biol Chem</addtitle><description>The hepatitis C virus (HCV) RNA-dependent RNA polymerase (RdRp), represented by nonstructural protein 5B (NS5B), is believed to form a membrane-associated RNA replication complex together with other nonstructural proteins and as yet unidentified host components. However, the determinants for membrane association of this essential viral enzyme have not been defined. By double label immunofluorescence analyses, NS5B was found in the endoplasmic reticulum (ER) or an ER-like modified compartment both when expressed alone or in the context of the entire HCV polyprotein. The carboxyl-terminal 21 amino acid residues were necessary and sufficient to target NS5B or a heterologous protein to the cytosolic side of the ER membrane. This hydrophobic domain is highly conserved among 269 HCV isolates analyzed and predicted to form a transmembrane α-helix. Association of NS5B with the ER membrane occurred by a posttranslational mechanism that was ATP-independent. These features define the HCV RdRp as a new member of the tail-anchored protein family, a class of integral membrane proteins that are membrane-targeted posttranslationally via a carboxyl-terminal insertion sequence. Formation of the HCV replication complex, therefore, involves specific determinants for membrane association that represent potential targets for antiviral intervention.</description><subject>Amino Acid Sequence</subject><subject>Base Sequence</subject><subject>Biochemistry, Molecular Biology</subject><subject>Cell Line</subject><subject>Cell Membrane - metabolism</subject><subject>Cell Membrane - virology</subject><subject>DNA Primers</subject><subject>Endoplasmic Reticulum - metabolism</subject><subject>Fluorescent Antibody Technique</subject><subject>Hepacivirus - enzymology</subject><subject>Hepatitis C virus</subject><subject>Humans</subject><subject>Life Sciences</subject><subject>Molecular Sequence Data</subject><subject>NS5B protein</subject><subject>Protein Biosynthesis</subject><subject>RNA-Dependent RNA Polymerase - metabolism</subject><subject>Sequence Homology, Amino Acid</subject><subject>Tetracycline - pharmacology</subject><subject>Transcription, Genetic</subject><subject>Viral Nonstructural Proteins - chemistry</subject><subject>Viral Nonstructural Proteins - metabolism</subject><issn>0021-9258</issn><issn>1083-351X</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2001</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkU1vEzEQhi0EomnhyhH5gJB62DD-2LX3GIWWIKUFIUBIHCyvd5a42l2n9qao_x6HRO0J4ct4Rs-8mpmXkFcM5gyUfHfTuPkVAyFKzQGekBkDLQpRsh9PyQyAs6LmpT4hpyndQH6yZs_JCWNlqVTJZ-Tne5wwDn6045RoFyK9wqGJdkS6SCk4bycfRho6Om2QrnCb88knuqTffdwl-uV6UbS4xbHFcdpn9HPo7weMNuEL8qyzfcKXx3hGvl1efF2uivWnDx-Xi3XhSglT0dW2yz8tQTSKSdeAqrkVQkElRIfOas1LZ1umK8kq3rJSdY1uqlqAbGqJ4oycH3Q3tjfb6Acb702w3qwWa7OvAQiW5dQdy-zbA7uN4XaHaTKDTw77Pm8cdskozlXNhP4vyDTjNdR7cH4AXQwpReweRmBg9h6Z7JF59Cg3vD4q75oB20f8aEoG3hz38b82v31E0_jgNjgYriojlZES_mL6gGG-7Z3HaJLzODpsc4ubTBv8v0b4A1oxqQY</recordid><startdate>20011123</startdate><enddate>20011123</enddate><creator>Schmidt-Mende, Juliane</creator><creator>Bieck, Elke</creator><creator>Hügle, Thomas</creator><creator>Penin, François</creator><creator>Rice, Charles M.</creator><creator>Blum, Hubert E.</creator><creator>Moradpour, Darius</creator><general>Elsevier Inc</general><general>American Society for Biochemistry and Molecular Biology</general><scope>6I.</scope><scope>AAFTH</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7TM</scope><scope>7U9</scope><scope>H94</scope><scope>7X8</scope><scope>1XC</scope></search><sort><creationdate>20011123</creationdate><title>Determinants for Membrane Association of the Hepatitis C Virus RNA-dependent RNA Polymerase</title><author>Schmidt-Mende, Juliane ; Bieck, Elke ; Hügle, Thomas ; Penin, François ; Rice, Charles M. ; Blum, Hubert E. ; Moradpour, Darius</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c540t-f9afc548403b714cb0792a3370633feca8825cad1864162d157fb8b69304b94e3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2001</creationdate><topic>Amino Acid Sequence</topic><topic>Base Sequence</topic><topic>Biochemistry, Molecular Biology</topic><topic>Cell Line</topic><topic>Cell Membrane - metabolism</topic><topic>Cell Membrane - virology</topic><topic>DNA Primers</topic><topic>Endoplasmic Reticulum - metabolism</topic><topic>Fluorescent Antibody Technique</topic><topic>Hepacivirus - enzymology</topic><topic>Hepatitis C virus</topic><topic>Humans</topic><topic>Life Sciences</topic><topic>Molecular Sequence Data</topic><topic>NS5B protein</topic><topic>Protein Biosynthesis</topic><topic>RNA-Dependent RNA Polymerase - metabolism</topic><topic>Sequence Homology, Amino Acid</topic><topic>Tetracycline - pharmacology</topic><topic>Transcription, Genetic</topic><topic>Viral Nonstructural Proteins - chemistry</topic><topic>Viral Nonstructural Proteins - metabolism</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Schmidt-Mende, Juliane</creatorcontrib><creatorcontrib>Bieck, Elke</creatorcontrib><creatorcontrib>Hügle, Thomas</creatorcontrib><creatorcontrib>Penin, François</creatorcontrib><creatorcontrib>Rice, Charles M.</creatorcontrib><creatorcontrib>Blum, Hubert E.</creatorcontrib><creatorcontrib>Moradpour, Darius</creatorcontrib><collection>ScienceDirect Open Access Titles</collection><collection>Elsevier:ScienceDirect:Open Access</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Nucleic Acids Abstracts</collection><collection>Virology and AIDS Abstracts</collection><collection>AIDS and Cancer Research Abstracts</collection><collection>MEDLINE - Academic</collection><collection>Hyper Article en Ligne (HAL)</collection><jtitle>The Journal of biological chemistry</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Schmidt-Mende, Juliane</au><au>Bieck, Elke</au><au>Hügle, Thomas</au><au>Penin, François</au><au>Rice, Charles M.</au><au>Blum, Hubert E.</au><au>Moradpour, Darius</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Determinants for Membrane Association of the Hepatitis C Virus RNA-dependent RNA Polymerase</atitle><jtitle>The Journal of biological chemistry</jtitle><addtitle>J Biol Chem</addtitle><date>2001-11-23</date><risdate>2001</risdate><volume>276</volume><issue>47</issue><spage>44052</spage><epage>44063</epage><pages>44052-44063</pages><issn>0021-9258</issn><eissn>1083-351X</eissn><abstract>The hepatitis C virus (HCV) RNA-dependent RNA polymerase (RdRp), represented by nonstructural protein 5B (NS5B), is believed to form a membrane-associated RNA replication complex together with other nonstructural proteins and as yet unidentified host components. However, the determinants for membrane association of this essential viral enzyme have not been defined. By double label immunofluorescence analyses, NS5B was found in the endoplasmic reticulum (ER) or an ER-like modified compartment both when expressed alone or in the context of the entire HCV polyprotein. The carboxyl-terminal 21 amino acid residues were necessary and sufficient to target NS5B or a heterologous protein to the cytosolic side of the ER membrane. This hydrophobic domain is highly conserved among 269 HCV isolates analyzed and predicted to form a transmembrane α-helix. Association of NS5B with the ER membrane occurred by a posttranslational mechanism that was ATP-independent. These features define the HCV RdRp as a new member of the tail-anchored protein family, a class of integral membrane proteins that are membrane-targeted posttranslationally via a carboxyl-terminal insertion sequence. 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| subjects | Amino Acid Sequence Base Sequence Biochemistry, Molecular Biology Cell Line Cell Membrane - metabolism Cell Membrane - virology DNA Primers Endoplasmic Reticulum - metabolism Fluorescent Antibody Technique Hepacivirus - enzymology Hepatitis C virus Humans Life Sciences Molecular Sequence Data NS5B protein Protein Biosynthesis RNA-Dependent RNA Polymerase - metabolism Sequence Homology, Amino Acid Tetracycline - pharmacology Transcription, Genetic Viral Nonstructural Proteins - chemistry Viral Nonstructural Proteins - metabolism |
| title | Determinants for Membrane Association of the Hepatitis C Virus RNA-dependent RNA Polymerase |
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