title: Live Cell Imaging Confocal Microscopy Analysis of HBV Myr-PreS1 Peptide Binding and Uptake in NTCP-GFP Expressing HepG2 Cells.
abstract: To obtain basic knowledge about specific molecular mechanisms involved in the entry of pathogens into cells is the basis for establishing pharmacologic substances blocking initial viral binding, infection, and subsequent viral spread. Lack of information about key cellular factors involved in the initial steps of HBV infection has hampered the characterization of HBV binding and entry for decades. However, recently, the liver-specific sodium-dependent taurocholate cotransporting polypeptide (NTCP) has been discovered as a functional receptor for HBV and HDV, thus opening the field for new concepts of basic binding and entry of HBV and HDV. Here, we describe practical issues of a basic in vitro assay system to examine kinetics and mechanisms of receptor-dependent HBV binding, uptake, and intracellular trafficking by live-cell imaging confocal microscopy. The assay system is comprised of HepG2 cells expressing a NTCP-GFP fusion-protein and chemically synthesized, fluorophore-labeled part of HBV surface protein, spanning the first N-terminal 48 amino acids of preS1 of the large hepatitis B virus surface protein.
authors.affiliation: Institute of Medical Virology, Justus Liebig University Giessen, National Reference Center for Hepatitis B and D Viruses, Biomedical Research Center Seltersberg, Schubertstr. 81, 35392, Giessen, Germany.|German Center for Infection Research (DZIF), partner site Giessen, Germany.
authors.forename: Alexander
authors.initials: A
authors.lastname: König
authors.affiliation: Institute of Medical Virology, Justus Liebig University Giessen, National Reference Center for Hepatitis B and D Viruses, Biomedical Research Center Seltersberg, Schubertstr. 81, 35392, Giessen, Germany. dieter.glebe@viro.med.uni-giessen.de.|German Center for Infection Research (DZIF), partner site Giessen, Germany. dieter.glebe@viro.med.uni-giessen.de.
authors.forename: Dieter
authors.initials: D
authors.lastname: Glebe
mesh_terms: D002478:Cells, Cultured; D015870:Gene Expression; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D056945:Hep G2 Cells; D006509:Hepatitis B / Q000235:genetics / Q000378:metabolism* / Q000821:virology*; D006514:Hepatitis B Surface Antigens / Q000378:metabolism*; D006515:Hepatitis B virus / Q000502:physiology*; D006801:Humans; D018613:Microscopy, Confocal; D057054:Molecular Imaging / Q000379:methods; D029363:Organic Anion Transporters, Sodium-Dependent / Q000235:genetics / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics; D011498:Protein Precursors / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D027981:Symporters / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D053585:Virus Attachment; D053586:Virus Internalization
chemical_list: D006514:Hepatitis B Surface Antigens; D029363:Organic Anion Transporters, Sodium-Dependent; D011498:Protein Precursors; D011993:Recombinant Fusion Proteins; D027981:Symporters; C050806:presurface protein 1, hepatitis B surface antigen; C071908:sodium-bile acid cotransporter; D049452:Green Fluorescent Proteins
keywords: Endocytosis; HBV entry; HBV preS1-domain; Live-cell imaging confocal microscopy; NTCP