title: Mice Expressing Minimally Humanized CD81 and Occludin Genes Support Hepatitis C Virus Uptake In Vivo.
abstract: Hepatitis C virus (HCV) causes chronic infections in at least 150 million individuals worldwide. HCV has a narrow host range and robustly infects only humans and chimpanzees. The underlying mechanisms for this narrow host range are incompletely understood. At the level of entry, differences in the amino acid sequences between the human and mouse orthologues of two essential host factors, the tetraspanin CD81 and the tight junction protein occludin (OCLN), explain, at least in part, HCV's limited ability to enter mouse hepatocytes. We have previously shown that adenoviral or transgenic overexpression of human CD81 and OCLN facilitates HCV uptake into mouse hepatocytes in vitro and in vivo In efforts to refine these models, we constructed knock-in mice in which the second extracellular loops of CD81 and OCLN were replaced with the respective human sequences, which contain the determinants that are critical for HCV uptake. We demonstrate that the humanized CD81 and OCLN were expressed at physiological levels in a tissue-appropriate fashion. Mice bearing the humanized alleles formed normal tight junctions and did not exhibit any immunologic abnormalities, indicating that interactions with their physiological ligands were intact. HCV entry factor knock-in mice take up HCV with an efficiency similar to that in mice expressing HCV entry factors transgenically or adenovirally, demonstrating the utility of this model for studying HCV infection in vivo IMPORTANCE: At least 150 million individuals are chronically infected with hepatitis C virus (HCV). Chronic hepatitis C can result in progressive liver disease and liver cancer. New antiviral treatments can cure HCV in the majority of patients, but a vaccine remains elusive. To gain a better understanding of the processes culminating in liver failure and cancer and to prioritize vaccine candidates more efficiently, small-animal models are needed. Here, we describe the characterization of a new mouse model in which the parts of two host factors that are essential for HCV uptake, CD81 and occludin (OCLN), which differ between mice and humans, were humanized. We demonstrate that such minimally humanized mice develop normally, express the modified genes at physiological levels, and support HCV uptake. This model is of considerable utility for studying viral entry in the three-dimensional context of the liver and to test approaches aimed at preventing HCV entry.
authors.affiliation: Princeton University, Department of Molecular Biology, Lewis Thomas Laboratory, Princeton New Jersey, USA.
authors.forename: Qiang
authors.initials: Q
authors.lastname: Ding
authors.affiliation: Princeton University, Department of Molecular Biology, Lewis Thomas Laboratory, Princeton New Jersey, USA.
authors.forename: Markus
authors.initials: M
authors.lastname: von Schaewen
authors.affiliation: Princeton University, Department of Molecular Biology, Lewis Thomas Laboratory, Princeton New Jersey, USA.
authors.forename: Gabriela
authors.initials: G
authors.lastname: Hrebikova
authors.affiliation: Princeton University, Department of Molecular Biology, Lewis Thomas Laboratory, Princeton New Jersey, USA.
authors.forename: Brigitte
authors.initials: B
authors.lastname: Heller
authors.affiliation: Hannover Medical School, Hannover, Germany.
authors.forename: Lisa
authors.initials: L
authors.lastname: Sandmann
authors.affiliation: University of Tartu, Faculty of Medicine, Institute of Biomedicine and Translational Medicine, Laboratory Animal Centre, Tartu, Estonia.
authors.forename: Mario
authors.initials: M
authors.lastname: Plaas
authors.affiliation: Princeton University, Department of Molecular Biology, Lewis Thomas Laboratory, Princeton New Jersey, USA aploss@princeton.edu.
authors.forename: Alexander
authors.initials: A
authors.lastname: Ploss
mesh_terms: D000483:Alleles; D000595:Amino Acid Sequence; D000818:Animals; D004195:Disease Models, Animal; D015870:Gene Expression; D018390:Gene Targeting; D017930:Genes, Reporter; D056426:Genetic Loci; D016174:Hepacivirus / Q000502:physiology*; D006526:Hepatitis C / Q000235:genetics* / Q000821:virology*; D054884:Host-Pathogen Interactions; D006801:Humans; D051379:Mice; D008822:Mice, Transgenic; D062793:Occludin / Q000235:genetics* / Q000378:metabolism; D009928:Organ Specificity / Q000235:genetics; D010641:Phenotype; D060225:Tetraspanin 28 / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic; D053586:Virus Internalization
chemical_list: D062793:Occludin; D060225:Tetraspanin 28
keywords: animal models; hepatitis C virus; viral entry; viral hepatitis