Yi Ni1, Florian A Lempp1, Stefan Mehrle1, Shirin Nkongolo1, Christina Kaufman1, Maria Fälth2, Jan Stindt3, Christian Königer4, Michael Nassal4, Ralf Kubitz3, Holger Sültmann2, Stephan Urban5. 1. Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, Germany. 2. German Cancer Research Center and National Center for Tumor Diseases, Unit Cancer Genome Research, Heidelberg, Germany. 3. Clinic for Gastroenterology, Hepatology and Infectiology, University Hospital Düsseldorf, Düsseldorf, Germany. 4. Department of Internal Medicine II, University Hospital Freiburg, Freiburg, Germany. 5. Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, Germany. Electronic address: Stephan.Urban@med.uni-heidelberg.de.
Abstract
BACKGROUND & AIMS: Hepatitis B and D viruses (HBV and HDV) are human pathogens with restricted host ranges and high selectivity for hepatocytes; the HBV L-envelope protein interacts specifically with a receptor on these cells. We aimed to identify this receptor and analyze whether it is the recently described sodium-taurocholate co-transporter polypeptide (NTCP), encoded by the SLC10A1 gene. METHODS: To identify receptor candidates, we compared gene expression patterns between differentiated HepaRG cells, which express the receptor, and naïve cells, which do not. Receptor candidates were evaluated by small hairpin RNA silencing in HepaRG cells; the ability of receptor expression to confer binding and infection were tested in transduced hepatoma cell lines. We used interspecies domain swapping to identify motifs for receptor-mediated host discrimination of HBV and HDV binding and infection. RESULTS: Bioinformatic analyses of comparative expression arrays confirmed that NTCP, which was previously identified through a biochemical approach is a bona fide receptor for HBV and HDV. NTCPs from rat, mouse, and human bound Myrcludex B, a peptide ligand derived from the HBV L-protein. Myrcludex B blocked NTCP transport of bile salts; small hairpin RNA-mediated knockdown of NTCP in HepaRG cells prevented their infection by HBV or HDV. Expression of human but not mouse NTCP in HepG2 and HuH7 cells conferred a limited cell-type-related and virus-dependent susceptibility to infection; these limitations were overcome when cells were cultured with dimethyl sulfoxide. We identified 2 short-sequence motifs in human NTCP that were required for species-specific binding and infection by HBV and HDV. CONCLUSIONS: Human NTCP is a specific receptor for HBV and HDV. NTCP-expressing cell lines can be efficiently infected with these viruses, and might be used in basic research and high-throughput screening studies. Mapping of motifs in NTCPs have increased our understanding of the species specificities of HBV and HDV, and could lead to small animal models for studies of viral infection and replication.
BACKGROUND & AIMS:Hepatitis B and D viruses (HBV and HDV) are human pathogens with restricted host ranges and high selectivity for hepatocytes; the HBV L-envelope protein interacts specifically with a receptor on these cells. We aimed to identify this receptor and analyze whether it is the recently described sodium-taurocholate co-transporter polypeptide (NTCP), encoded by the SLC10A1 gene. METHODS: To identify receptor candidates, we compared gene expression patterns between differentiated HepaRG cells, which express the receptor, and naïve cells, which do not. Receptor candidates were evaluated by small hairpin RNA silencing in HepaRG cells; the ability of receptor expression to confer binding and infection were tested in transduced hepatoma cell lines. We used interspecies domain swapping to identify motifs for receptor-mediated host discrimination of HBV and HDV binding and infection. RESULTS: Bioinformatic analyses of comparative expression arrays confirmed that NTCP, which was previously identified through a biochemical approach is a bona fide receptor for HBV and HDV. NTCPs from rat, mouse, and human bound Myrcludex B, a peptide ligand derived from the HBV L-protein. Myrcludex B blocked NTCP transport of bile salts; small hairpin RNA-mediated knockdown of NTCP in HepaRG cells prevented their infection by HBV or HDV. Expression of human but not mouseNTCP in HepG2 and HuH7 cells conferred a limited cell-type-related and virus-dependent susceptibility to infection; these limitations were overcome when cells were cultured with dimethyl sulfoxide. We identified 2 short-sequence motifs in humanNTCP that were required for species-specific binding and infection by HBV and HDV. CONCLUSIONS:HumanNTCP is a specific receptor for HBV and HDV. NTCP-expressing cell lines can be efficiently infected with these viruses, and might be used in basic research and high-throughput screening studies. Mapping of motifs in NTCPs have increased our understanding of the species specificities of HBV and HDV, and could lead to small animal models for studies of viral infection and replication.
Authors: T Jake Liang; Timothy M Block; Brian J McMahon; Marc G Ghany; Stephan Urban; Ju-Tao Guo; Stephen Locarnini; Fabien Zoulim; Kyong-Mi Chang; Anna S Lok Journal: Hepatology Date: 2015-10-27 Impact factor: 17.425
Authors: Benjamin Y Winer; Elham Shirvani-Dastgerdi; Yaron Bram; Julie Sellau; Benjamin E Low; Heath Johnson; Tiffany Huang; Gabriela Hrebikova; Brigitte Heller; Yael Sharon; Katja Giersch; Sherif Gerges; Kathleen Seneca; Mihai-Alexandru Pais; Angela S Frankel; Luis Chiriboga; John Cullen; Ronald G Nahass; Marc Lutgehetmann; Jared E Toettcher; Michael V Wiles; Robert E Schwartz; Alexander Ploss Journal: Sci Transl Med Date: 2018-06-27 Impact factor: 17.956