In vitro adaptation and characterization of attenuated hypervariable region 1 swap chimeras of hepatitis C virus
Hepatitis C virus (HCV) chronically infects 70 million people worldwide with an estimated annual disease-related mortality of 400,000. A vaccine could prevent spread of this pervasive human pathogen, but has proven difficult to develop, partly due to neutralizing antibody evasion mechanisms that are...
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| description | Hepatitis C virus (HCV) chronically infects 70 million people worldwide with an estimated annual disease-related mortality of 400,000. A vaccine could prevent spread of this pervasive human pathogen, but has proven difficult to develop, partly due to neutralizing antibody evasion mechanisms that are inherent features of the virus envelope glycoproteins, E1 and E2. A central actor is the E2 motif, hypervariable region 1 (HVR1), which protects several non-overlapping neutralization epitopes through an incompletely understood mechanism. Here, we show that introducing different HVR1-isolate sequences into cell-culture infectious JFH1-based H77 (genotype 1a) and J4 (genotype 1b) Core-NS2 recombinants can lead to severe viral attenuation. Culture adaptation of attenuated HVR1-swapped recombinants permitted us to identify E1/E2 substitutions at conserved positions both within and outside HVR1 that increased the infectivity of attenuated HVR1-swapped recombinants but were not adaptive for original recombinants. H77 recombinants with HVR1 from multiple other isolates consistently acquired substitutions at position 348 in E1 and position 385 in HVR1 of E2. Interestingly, HVR1-swapped J4 recombinants primarily acquired other substitutions: F291I (E1), F438V (E2), F447L/V/I (E2) and V710L (E2), indicating a different adaptation pathway. For H77 recombinants, the adaptive E1/E2 substitutions increased sensitivity to the neutralizing monoclonal antibodies AR3A and AR4A, whereas for J4 recombinants, they increased sensitivity to AR3A, while having no effect on sensitivity to AR4A. To evaluate effects of the substitutions on AR3A and AR4A binding, we performed ELISAs on extracted E1/E2 protein and performed immunoprecipitation of relevant viruses. However, extracted E1/E2 protein and immunoprecipitation of HCV particles only reproduced the neutralization phenotypes of the J4 recombinants. Finally, we found that the HVR1-swap E1/E2 substitutions decrease virus entry dependency on co-receptor SR-BI. Our study identifies E1/E2 positions that could be critical for intra-complex HVR1 interactions while emphasizing the need for developing novel tools for molecular studies of E1/E2 interactions. |
| doi_str_mv | 10.1371/journal.ppat.1009720 |
| format | Article |
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A vaccine could prevent spread of this pervasive human pathogen, but has proven difficult to develop, partly due to neutralizing antibody evasion mechanisms that are inherent features of the virus envelope glycoproteins, E1 and E2. A central actor is the E2 motif, hypervariable region 1 (HVR1), which protects several non-overlapping neutralization epitopes through an incompletely understood mechanism. Here, we show that introducing different HVR1-isolate sequences into cell-culture infectious JFH1-based H77 (genotype 1a) and J4 (genotype 1b) Core-NS2 recombinants can lead to severe viral attenuation. Culture adaptation of attenuated HVR1-swapped recombinants permitted us to identify E1/E2 substitutions at conserved positions both within and outside HVR1 that increased the infectivity of attenuated HVR1-swapped recombinants but were not adaptive for original recombinants. H77 recombinants with HVR1 from multiple other isolates consistently acquired substitutions at position 348 in E1 and position 385 in HVR1 of E2. Interestingly, HVR1-swapped J4 recombinants primarily acquired other substitutions: F291I (E1), F438V (E2), F447L/V/I (E2) and V710L (E2), indicating a different adaptation pathway. For H77 recombinants, the adaptive E1/E2 substitutions increased sensitivity to the neutralizing monoclonal antibodies AR3A and AR4A, whereas for J4 recombinants, they increased sensitivity to AR3A, while having no effect on sensitivity to AR4A. To evaluate effects of the substitutions on AR3A and AR4A binding, we performed ELISAs on extracted E1/E2 protein and performed immunoprecipitation of relevant viruses. However, extracted E1/E2 protein and immunoprecipitation of HCV particles only reproduced the neutralization phenotypes of the J4 recombinants. Finally, we found that the HVR1-swap E1/E2 substitutions decrease virus entry dependency on co-receptor SR-BI. Our study identifies E1/E2 positions that could be critical for intra-complex HVR1 interactions while emphasizing the need for developing novel tools for molecular studies of E1/E2 interactions.</description><identifier>ISSN: 1553-7374</identifier><identifier>ISSN: 1553-7366</identifier><identifier>EISSN: 1553-7374</identifier><identifier>DOI: 10.1371/journal.ppat.1009720</identifier><identifier>PMID: 34280245</identifier><language>eng</language><publisher>United States: Public Library of Science</publisher><subject>Adaptation ; Adaptation, Physiological - physiology ; Analysis ; Attenuation ; Biology and Life Sciences ; Cell culture ; Cell Line ; Chimera ; Chimeras ; Diagnosis ; Drug resistance ; E2 protein ; Epitopes ; Genetic aspects ; Genotype & phenotype ; Genotypes ; Glycoproteins ; HEK293 Cells ; Hepacivirus - pathogenicity ; Hepacivirus - physiology ; Hepatitis ; Hepatitis C ; Hepatitis C virus ; Humans ; Immune Evasion - physiology ; Immunoprecipitation ; Infections ; Infectivity ; Medicine and health sciences ; Monoclonal antibodies ; Mutation ; Neutralization ; Neutralizing ; Phenotypes ; Proteins ; Recombinants ; Research and Analysis Methods ; Sensitivity analysis ; Vaccines ; Viral Envelope Proteins - physiology ; Virus Internalization ; Viruses</subject><ispartof>PLoS pathogens, 2021-07, Vol.17 (7), p.e1009720-e1009720</ispartof><rights>COPYRIGHT 2021 Public Library of Science</rights><rights>2021 Olesen et al. This is an open access article distributed under the terms of the Creative Commons Attribution License: http://creativecommons.org/licenses/by/4.0/ (the “License”), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License.</rights><rights>2021 Olesen et al 2021 Olesen et al</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c661t-cacfedd8bbb4b76764a4ee6231a052f5ad420effd2945b5e6b4a74d74bf7de573</citedby><cites>FETCH-LOGICAL-c661t-cacfedd8bbb4b76764a4ee6231a052f5ad420effd2945b5e6b4a74d74bf7de573</cites><orcidid>0000-0002-7815-4806 ; 0000-0002-0575-1597</orcidid></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC8321405/pdf/$$EPDF$$P50$$Gpubmedcentral$$Hfree_for_read</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC8321405/$$EHTML$$P50$$Gpubmedcentral$$Hfree_for_read</linktohtml><link.rule.ids>230,314,727,780,784,864,885,2102,2928,23866,27924,27925,53791,53793,79600,79601</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/34280245$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><contributor>Ploss, Alexander</contributor><creatorcontrib>Olesen, Christina Holmboe</creatorcontrib><creatorcontrib>Augestad, Elias H</creatorcontrib><creatorcontrib>Troise, Fulvia</creatorcontrib><creatorcontrib>Bukh, Jens</creatorcontrib><creatorcontrib>Prentoe, Jannick</creatorcontrib><title>In vitro adaptation and characterization of attenuated hypervariable region 1 swap chimeras of hepatitis C virus</title><title>PLoS pathogens</title><addtitle>PLoS Pathog</addtitle><description>Hepatitis C virus (HCV) chronically infects 70 million people worldwide with an estimated annual disease-related mortality of 400,000. A vaccine could prevent spread of this pervasive human pathogen, but has proven difficult to develop, partly due to neutralizing antibody evasion mechanisms that are inherent features of the virus envelope glycoproteins, E1 and E2. A central actor is the E2 motif, hypervariable region 1 (HVR1), which protects several non-overlapping neutralization epitopes through an incompletely understood mechanism. Here, we show that introducing different HVR1-isolate sequences into cell-culture infectious JFH1-based H77 (genotype 1a) and J4 (genotype 1b) Core-NS2 recombinants can lead to severe viral attenuation. Culture adaptation of attenuated HVR1-swapped recombinants permitted us to identify E1/E2 substitutions at conserved positions both within and outside HVR1 that increased the infectivity of attenuated HVR1-swapped recombinants but were not adaptive for original recombinants. H77 recombinants with HVR1 from multiple other isolates consistently acquired substitutions at position 348 in E1 and position 385 in HVR1 of E2. Interestingly, HVR1-swapped J4 recombinants primarily acquired other substitutions: F291I (E1), F438V (E2), F447L/V/I (E2) and V710L (E2), indicating a different adaptation pathway. For H77 recombinants, the adaptive E1/E2 substitutions increased sensitivity to the neutralizing monoclonal antibodies AR3A and AR4A, whereas for J4 recombinants, they increased sensitivity to AR3A, while having no effect on sensitivity to AR4A. To evaluate effects of the substitutions on AR3A and AR4A binding, we performed ELISAs on extracted E1/E2 protein and performed immunoprecipitation of relevant viruses. However, extracted E1/E2 protein and immunoprecipitation of HCV particles only reproduced the neutralization phenotypes of the J4 recombinants. Finally, we found that the HVR1-swap E1/E2 substitutions decrease virus entry dependency on co-receptor SR-BI. Our study identifies E1/E2 positions that could be critical for intra-complex HVR1 interactions while emphasizing the need for developing novel tools for molecular studies of E1/E2 interactions.</description><subject>Adaptation</subject><subject>Adaptation, Physiological - physiology</subject><subject>Analysis</subject><subject>Attenuation</subject><subject>Biology and Life Sciences</subject><subject>Cell culture</subject><subject>Cell Line</subject><subject>Chimera</subject><subject>Chimeras</subject><subject>Diagnosis</subject><subject>Drug resistance</subject><subject>E2 protein</subject><subject>Epitopes</subject><subject>Genetic aspects</subject><subject>Genotype & phenotype</subject><subject>Genotypes</subject><subject>Glycoproteins</subject><subject>HEK293 Cells</subject><subject>Hepacivirus - pathogenicity</subject><subject>Hepacivirus - physiology</subject><subject>Hepatitis</subject><subject>Hepatitis C</subject><subject>Hepatitis C virus</subject><subject>Humans</subject><subject>Immune Evasion - physiology</subject><subject>Immunoprecipitation</subject><subject>Infections</subject><subject>Infectivity</subject><subject>Medicine and health sciences</subject><subject>Monoclonal antibodies</subject><subject>Mutation</subject><subject>Neutralization</subject><subject>Neutralizing</subject><subject>Phenotypes</subject><subject>Proteins</subject><subject>Recombinants</subject><subject>Research and Analysis Methods</subject><subject>Sensitivity analysis</subject><subject>Vaccines</subject><subject>Viral Envelope Proteins - physiology</subject><subject>Virus Internalization</subject><subject>Viruses</subject><issn>1553-7374</issn><issn>1553-7366</issn><issn>1553-7374</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2021</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>ABUWG</sourceid><sourceid>AFKRA</sourceid><sourceid>AZQEC</sourceid><sourceid>BENPR</sourceid><sourceid>CCPQU</sourceid><sourceid>DWQXO</sourceid><sourceid>GNUQQ</sourceid><sourceid>DOA</sourceid><recordid>eNqVkltv1DAQhSMEoqXwDxBE4gUedrEdXzYvlaoVl5UqkLg8WxN7sutVNg62Uyi_HofdVl3UF5SHRJPvnDkzmqJ4TsmcVoq-3fox9NDNhwHSnBJSK0YeFKdUiGqmKsUf3vk-KZ7EuCWE04rKx8VJxdmCMC5Oi2HVl1cuBV-ChSFBcr4vobel2UAAkzC43_uib0tICfsREtpycz1guILgoOmwDLieEFrGnzBkqdthgDhJNpjjueRiucx9whifFo9a6CI-O7zPiu_v331bfpxdfv6wWl5czoyUNM0MmBatXTRNwxslleTAESWrKBDBWgGWM4Jta1nNRSNQNhwUt4o3rbIoVHVWvNz7Dp2P-rCsqJmQtOY0e2RitSesh60egttBuNYenP5b8GGtISRnOtQGGiukYmi45DXDmjU1l1IRU1kua8xe54duY7NDa7BPAboj0-M_vdvotb_Si4pRTkQ2eH0wCP7HiDHpnYsGuw569OOUW1Q5M6VT7lf_oPdPd6DWkAdwfetzXzOZ6oscvM6NFzJT83uo_FjcOeN7bF2uHwneHAkyk_BXWsMYo159_fIf7Kdjlu9ZE3yMAdvb3VGip3u_GVJP964P955lL-7u_VZ0c-DVH_nz_Xo</recordid><startdate>20210701</startdate><enddate>20210701</enddate><creator>Olesen, Christina Holmboe</creator><creator>Augestad, Elias H</creator><creator>Troise, Fulvia</creator><creator>Bukh, Jens</creator><creator>Prentoe, Jannick</creator><general>Public Library of Science</general><general>Public Library of Science (PLoS)</general><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>ISN</scope><scope>ISR</scope><scope>3V.</scope><scope>7QL</scope><scope>7U9</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>8FE</scope><scope>8FH</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABUWG</scope><scope>AFKRA</scope><scope>AZQEC</scope><scope>BBNVY</scope><scope>BENPR</scope><scope>BHPHI</scope><scope>C1K</scope><scope>CCPQU</scope><scope>DWQXO</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>H94</scope><scope>HCIFZ</scope><scope>K9.</scope><scope>LK8</scope><scope>M0S</scope><scope>M1P</scope><scope>M7P</scope><scope>PIMPY</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>7X8</scope><scope>5PM</scope><scope>DOA</scope><orcidid>https://orcid.org/0000-0002-7815-4806</orcidid><orcidid>https://orcid.org/0000-0002-0575-1597</orcidid></search><sort><creationdate>20210701</creationdate><title>In vitro adaptation and characterization of attenuated hypervariable region 1 swap chimeras of hepatitis C virus</title><author>Olesen, Christina Holmboe ; Augestad, Elias H ; Troise, Fulvia ; Bukh, Jens ; Prentoe, Jannick</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c661t-cacfedd8bbb4b76764a4ee6231a052f5ad420effd2945b5e6b4a74d74bf7de573</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2021</creationdate><topic>Adaptation</topic><topic>Adaptation, Physiological - 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physiology</topic><topic>Virus Internalization</topic><topic>Viruses</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Olesen, Christina Holmboe</creatorcontrib><creatorcontrib>Augestad, Elias H</creatorcontrib><creatorcontrib>Troise, Fulvia</creatorcontrib><creatorcontrib>Bukh, Jens</creatorcontrib><creatorcontrib>Prentoe, Jannick</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Gale In Context: Canada</collection><collection>Gale In Context: Science</collection><collection>ProQuest Central (Corporate)</collection><collection>Bacteriology Abstracts (Microbiology B)</collection><collection>Virology and AIDS Abstracts</collection><collection>Health & Medical Collection</collection><collection>ProQuest Central (purchase pre-March 2016)</collection><collection>Medical Database (Alumni Edition)</collection><collection>ProQuest SciTech Collection</collection><collection>ProQuest Natural Science Collection</collection><collection>Hospital Premium Collection</collection><collection>Hospital Premium Collection (Alumni Edition)</collection><collection>ProQuest Central (Alumni) (purchase pre-March 2016)</collection><collection>ProQuest Central (Alumni Edition)</collection><collection>ProQuest Central UK/Ireland</collection><collection>ProQuest Central Essentials</collection><collection>Biological Science Collection</collection><collection>ProQuest Central</collection><collection>Natural Science Collection</collection><collection>Environmental Sciences and Pollution Management</collection><collection>ProQuest One Community College</collection><collection>ProQuest Central Korea</collection><collection>Health Research Premium Collection</collection><collection>Health Research Premium Collection (Alumni)</collection><collection>ProQuest Central Student</collection><collection>AIDS and Cancer Research Abstracts</collection><collection>SciTech Premium Collection</collection><collection>ProQuest Health & Medical Complete (Alumni)</collection><collection>ProQuest Biological Science Collection</collection><collection>Health & Medical Collection (Alumni Edition)</collection><collection>Medical Database</collection><collection>Biological Science Database</collection><collection>Publicly Available Content Database</collection><collection>ProQuest One Academic Eastern Edition (DO NOT USE)</collection><collection>ProQuest One Academic</collection><collection>ProQuest One Academic UKI Edition</collection><collection>MEDLINE - Academic</collection><collection>PubMed Central (Full Participant titles)</collection><collection>DOAJ Directory of Open Access Journals</collection><jtitle>PLoS pathogens</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Olesen, Christina Holmboe</au><au>Augestad, Elias H</au><au>Troise, Fulvia</au><au>Bukh, Jens</au><au>Prentoe, Jannick</au><au>Ploss, Alexander</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>In vitro adaptation and characterization of attenuated hypervariable region 1 swap chimeras of hepatitis C virus</atitle><jtitle>PLoS pathogens</jtitle><addtitle>PLoS Pathog</addtitle><date>2021-07-01</date><risdate>2021</risdate><volume>17</volume><issue>7</issue><spage>e1009720</spage><epage>e1009720</epage><pages>e1009720-e1009720</pages><issn>1553-7374</issn><issn>1553-7366</issn><eissn>1553-7374</eissn><abstract>Hepatitis C virus (HCV) chronically infects 70 million people worldwide with an estimated annual disease-related mortality of 400,000. A vaccine could prevent spread of this pervasive human pathogen, but has proven difficult to develop, partly due to neutralizing antibody evasion mechanisms that are inherent features of the virus envelope glycoproteins, E1 and E2. A central actor is the E2 motif, hypervariable region 1 (HVR1), which protects several non-overlapping neutralization epitopes through an incompletely understood mechanism. Here, we show that introducing different HVR1-isolate sequences into cell-culture infectious JFH1-based H77 (genotype 1a) and J4 (genotype 1b) Core-NS2 recombinants can lead to severe viral attenuation. Culture adaptation of attenuated HVR1-swapped recombinants permitted us to identify E1/E2 substitutions at conserved positions both within and outside HVR1 that increased the infectivity of attenuated HVR1-swapped recombinants but were not adaptive for original recombinants. H77 recombinants with HVR1 from multiple other isolates consistently acquired substitutions at position 348 in E1 and position 385 in HVR1 of E2. Interestingly, HVR1-swapped J4 recombinants primarily acquired other substitutions: F291I (E1), F438V (E2), F447L/V/I (E2) and V710L (E2), indicating a different adaptation pathway. For H77 recombinants, the adaptive E1/E2 substitutions increased sensitivity to the neutralizing monoclonal antibodies AR3A and AR4A, whereas for J4 recombinants, they increased sensitivity to AR3A, while having no effect on sensitivity to AR4A. To evaluate effects of the substitutions on AR3A and AR4A binding, we performed ELISAs on extracted E1/E2 protein and performed immunoprecipitation of relevant viruses. However, extracted E1/E2 protein and immunoprecipitation of HCV particles only reproduced the neutralization phenotypes of the J4 recombinants. Finally, we found that the HVR1-swap E1/E2 substitutions decrease virus entry dependency on co-receptor SR-BI. Our study identifies E1/E2 positions that could be critical for intra-complex HVR1 interactions while emphasizing the need for developing novel tools for molecular studies of E1/E2 interactions.</abstract><cop>United States</cop><pub>Public Library of Science</pub><pmid>34280245</pmid><doi>10.1371/journal.ppat.1009720</doi><orcidid>https://orcid.org/0000-0002-7815-4806</orcidid><orcidid>https://orcid.org/0000-0002-0575-1597</orcidid><oa>free_for_read</oa></addata></record> |
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| ispartof | PLoS pathogens, 2021-07, Vol.17 (7), p.e1009720-e1009720 |
| issn | 1553-7374 1553-7366 1553-7374 |
| language | eng |
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| source | MEDLINE; DOAJ Directory of Open Access Journals; PubMed Central Open Access; Public Library of Science (PLoS); EZB-FREE-00999 freely available EZB journals; PubMed Central |
| subjects | Adaptation Adaptation, Physiological - physiology Analysis Attenuation Biology and Life Sciences Cell culture Cell Line Chimera Chimeras Diagnosis Drug resistance E2 protein Epitopes Genetic aspects Genotype & phenotype Genotypes Glycoproteins HEK293 Cells Hepacivirus - pathogenicity Hepacivirus - physiology Hepatitis Hepatitis C Hepatitis C virus Humans Immune Evasion - physiology Immunoprecipitation Infections Infectivity Medicine and health sciences Monoclonal antibodies Mutation Neutralization Neutralizing Phenotypes Proteins Recombinants Research and Analysis Methods Sensitivity analysis Vaccines Viral Envelope Proteins - physiology Virus Internalization Viruses |
| title | In vitro adaptation and characterization of attenuated hypervariable region 1 swap chimeras of hepatitis C virus |
| url | https://sfx.bib-bvb.de/sfx_tum?ctx_ver=Z39.88-2004&ctx_enc=info:ofi/enc:UTF-8&ctx_tim=2025-01-06T12%3A34%3A04IST&url_ver=Z39.88-2004&url_ctx_fmt=infofi/fmt:kev:mtx:ctx&rfr_id=info:sid/primo.exlibrisgroup.com:primo3-Article-gale_plos_&rft_val_fmt=info:ofi/fmt:kev:mtx:journal&rft.genre=article&rft.atitle=In%20vitro%20adaptation%20and%20characterization%20of%20attenuated%20hypervariable%20region%201%20swap%20chimeras%20of%20hepatitis%20C%20virus&rft.jtitle=PLoS%20pathogens&rft.au=Olesen,%20Christina%20Holmboe&rft.date=2021-07-01&rft.volume=17&rft.issue=7&rft.spage=e1009720&rft.epage=e1009720&rft.pages=e1009720-e1009720&rft.issn=1553-7374&rft.eissn=1553-7374&rft_id=info:doi/10.1371/journal.ppat.1009720&rft_dat=%3Cgale_plos_%3EA670983286%3C/gale_plos_%3E%3Curl%3E%3C/url%3E&disable_directlink=true&sfx.directlink=off&sfx.report_link=0&rft_id=info:oai/&rft_pqid=2561941052&rft_id=info:pmid/34280245&rft_galeid=A670983286&rft_doaj_id=oai_doaj_org_article_cabd5672ec46492e92b946670c3d469e&rfr_iscdi=true |