{"id":{"repo_id":"dundee","oai_identifier":"oai:discovery.dundee.ac.uk:studenttheses/1c55ab45-cd21-4475-a858-6b1bc46d8b92"},"canonical_url":"https://search.dev.ndltd.org/etd/dundee/oai:discovery.dundee.ac.uk:studenttheses/1c55ab45-cd21-4475-a858-6b1bc46d8b92","repository":{"repo_id":"dundee","name":"University of Dundee","base_url":"https://discovery.dundee.ac.uk/ws/oai"},"display":{"title":"Molecular Analysis of Hepatitis C Virus Envelope Glycoprotein E2 Binding and Entry","abstract":"The envelope of the hepatitis C virus (HCV) mediates entry into cells by binding directly to the cluster of differentiation 81 receptor (CD81) and to the scavenger receptor class B type 1 (SR-B1). The critical incorporation of claudin-1 receptor (CLDN-1) and occludin receptor (OCLN) (indirect interaction) has been reported. Then, fusion process is intiated between viral and cellular membranes following acidification of endosomes. All known HCV-receptor interactions are mediated by HCV envelope protein 2 (E2) but the manner in which E2 coordinates interactions with multiple entry receptors and cell surface co-factors in order to promote viral entry are only just beginning to be understood. Here, we have developed soluble recombinant forms of the E2 protein, which are fused to the Fc region of human IgG for use in the dissection of the E2 function and as molecular probes to interrogate the early events in the E2-dependent HCV entry pathway. These recombinant E2-immunoadhesins retain their immunological profile, as well as the cell surface and CD81-binding specificity that are typical of native E2. We have demonstrated that E2, in the complete absence of all other viral protein, is competent both for targeted binding and for internalisation into hepatoma cells. Rates of E2-Fc internalisation differ between cell types, being dependent on the density of the CD81 receptors displayed on the cell surface, to an extent. Internalised E2-immunoadhesin localises with endocytosis markers and tends to accumulate in early endosomes in target cells. Although binding to CD81 promotes and accelerates E2 internalisation rates, high-level expression, along with the display of CD81, is not sufficient to drive the internalisation of E2 in 293T cells, which confirms that the recruitment of CD81 to a multi-component entry complex is a critical event in the rapid attachment and endocytosis of E2. We have also identified important residues located somewhere within E2195 immunoadhesin, which undergo remarkably direct interactions with alternative host surface factors, when known HCV receptors are engaged. Our data enables us to speculate that the alternative factor has not defined yet and need further study.","abstract_html":"The envelope of the hepatitis C virus (HCV) mediates entry into cells by binding directly to the cluster of differentiation 81 receptor (CD81) and to the scavenger receptor class B type 1 (SR-B1). The critical incorporation of claudin-1 receptor (CLDN-1) and occludin receptor (OCLN) (indirect interaction) has been reported. Then, fusion process is intiated between viral and cellular membranes following acidification of endosomes. All known HCV-receptor interactions are mediated by HCV envelope protein 2 (E2) but the manner in which E2 coordinates interactions with multiple entry receptors and cell surface co-factors in order to promote viral entry are only just beginning to be understood. Here, we have developed soluble recombinant forms of the E2 protein, which are fused to the Fc region of human IgG for use in the dissection of the E2 function and as molecular probes to interrogate the early events in the E2-dependent HCV entry pathway. These recombinant E2-immunoadhesins retain their immunological profile, as well as the cell surface and CD81-binding specificity that are typical of native E2. We have demonstrated that E2, in the complete absence of all other viral protein, is competent both for targeted binding and for internalisation into hepatoma cells. Rates of E2-Fc internalisation differ between cell types, being dependent on the density of the CD81 receptors displayed on the cell surface, to an extent. Internalised E2-immunoadhesin localises with endocytosis markers and tends to accumulate in early endosomes in target cells. Although binding to CD81 promotes and accelerates E2 internalisation rates, high-level expression, along with the display of CD81, is not sufficient to drive the internalisation of E2 in 293T cells, which confirms that the recruitment of CD81 to a multi-component entry complex is a critical event in the rapid attachment and endocytosis of E2. We have also identified important residues located somewhere within E2195 immunoadhesin, which undergo remarkably direct interactions with alternative host surface factors, when known HCV receptors are engaged. Our data enables us to speculate that the alternative factor has not defined yet and need further study.","abstract_has_math":false,"creators":["Alshehri, Ahmad A."],"institution":"University of Dundee","degree_name":"Doctor of Philosophy","degree_level":"Doctoral Thesis","degree_discipline":null,"degree_department":null,"school":null,"contributors":[],"advisors":["Dillon, John"],"committee_chairs":[],"committee_members":[],"year":2017,"date_issued":"2017","date_published":"2017","updated_at":"2026-07-24T02:08:26Z","subjects":[],"languages":["eng"],"rights":[],"rights_urls":[],"identifier_entries":[{"key":"dc:identifier","label":"Identifier","values":["oai:discovery.dundee.ac.uk:studenttheses/1c55ab45-cd21-4475-a858-6b1bc46d8b92"],"render_values":[{"text":"oai:discovery.dundee.ac.uk:studenttheses/1c55ab45-cd21-4475-a858-6b1bc46d8b92","href":null,"code":true}]}]},"links":{"outbound_url":"https://discovery.dundee.ac.uk/en/studentTheses/1c55ab45-cd21-4475-a858-6b1bc46d8b92","outbound_label":"Repository record","outbound_source":"dc:identifier"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor.advisor","label":"Advisor","values":["Dillon, John"]},{"key":"dc:contributor.sponsor","label":"Sponsor","values":["Najran University"]},{"key":"dc:creator","label":"Author","values":["Alshehri, Ahmad A."]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date","label":"Dc Date","values":["2017"]},{"key":"dc:date.issued","label":"Date","values":["2017"]},{"key":"dc:publisher.department","label":"Dc Publisher Department","values":["Postgraduate Medicine"]},{"key":"dc:publisher.institution","label":"Dc Publisher Institution","values":["University of Dundee"]},{"key":"dc:relation.isreferencedby","label":"Dc Relation Isreferencedby","values":["https://discovery.dundee.ac.uk/en/studentTheses/1c55ab45-cd21-4475-a858-6b1bc46d8b92"]},{"key":"dc:type","label":"Dc Type","values":["Thesis"]},{"key":"dc:type.qualificationlevel","label":"Dc Type Qualificationlevel","values":["Doctoral Thesis"]},{"key":"dc:type.qualificationname","label":"Dc Type Qualificationname","values":["Doctor of Philosophy"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:language","label":"Dc Language","values":["eng"]},{"key":"dc:rights.embargodate","label":"Dc Rights Embargodate","values":["2020-06-30"]},{"key":"dc:rights.embargoreason","label":"Dc Rights Embargoreason","values":["/dk/atira/pure/core/document/studentthesisembargoreason/commercialexploitation"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier","label":"Identifier","values":["oai:discovery.dundee.ac.uk:studenttheses/1c55ab45-cd21-4475-a858-6b1bc46d8b92","https://discovery.dundee.ac.uk/en/studentTheses/1c55ab45-cd21-4475-a858-6b1bc46d8b92"]},{"key":"dc:identifier.uri","label":"Identifier URI","values":["https://discovery.dundee.ac.uk/files/37113995/MY_MAIN_CORRECTED_PhD_degree_Thesis.pdf"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description.abstract","label":"Abstract","values":["The envelope of the hepatitis C virus (HCV) mediates entry into cells by binding directly to the cluster of differentiation 81 receptor (CD81) and to the scavenger receptor class B type 1 (SR-B1). The critical incorporation of claudin-1 receptor (CLDN-1) and occludin receptor (OCLN) (indirect interaction) has been reported. Then, fusion process is intiated between viral and cellular membranes following acidification of endosomes. All known HCV-receptor interactions are mediated by HCV envelope protein 2 (E2) but the manner in which E2 coordinates interactions with multiple entry receptors and cell surface co-factors in order to promote viral entry are only just beginning to be understood. Here, we have developed soluble recombinant forms of the E2 protein, which are fused to the Fc region of human IgG for use in the dissection of the E2 function and as molecular probes to interrogate the early events in the E2-dependent HCV entry pathway. These recombinant E2-immunoadhesins retain their immunological profile, as well as the cell surface and CD81-binding specificity that are typical of native E2. We have demonstrated that E2, in the complete absence of all other viral protein, is competent both for targeted binding and for internalisation into hepatoma cells. Rates of E2-Fc internalisation differ between cell types, being dependent on the density of the CD81 receptors displayed on the cell surface, to an extent. Internalised E2-immunoadhesin localises with endocytosis markers and tends to accumulate in early endosomes in target cells. Although binding to CD81 promotes and accelerates E2 internalisation rates, high-level expression, along with the display of CD81, is not sufficient to drive the internalisation of E2 in 293T cells, which confirms that the recruitment of CD81 to a multi-component entry complex is a critical event in the rapid attachment and endocytosis of E2. 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Then, fusion process is intiated between viral and cellular membranes following acidification of endosomes. All known HCV-receptor interactions are mediated by HCV envelope protein 2 (E2) but the manner in which E2 coordinates interactions with multiple entry receptors and cell surface co-factors in order to promote viral entry are only just beginning to be understood. Here, we have developed soluble recombinant forms of the E2 protein, which are fused to the Fc region of human IgG for use in the dissection of the E2 function and as molecular probes to interrogate the early events in the E2-dependent HCV entry pathway. These recombinant E2-immunoadhesins retain their immunological profile, as well as the cell surface and CD81-binding specificity that are typical of native E2. We have demonstrated that E2, in the complete absence of all other viral protein, is competent both for targeted binding and for internalisation into hepatoma cells. 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