This is a competing renewal of a grant """"""""Identification and characterization of cellular factors involved in HCV entry"""""""" first funded in 2006. Our laboratory has been studying the mechanisms of hepatitis C virus (HCV) gene expression and replication for over twenty years, but our understanding of viral entry was limited by inadequate study systems. We and others overcame this hurdle by developing the first platforms for studying HCV entry in vitro - HCV pseudoparticles (HCVpp) and the fully-infectious HCV cell culture system (HCVcc). With the advent of these new tools, we proposed to identify and characterize factors required for HCV uptake. A long- term goal of these studies was to use this information to create much needed small animal models for HCV. Using lentivirus-based cyclic repackaging schemes, we succeeded in identifying two tight junction proteins, claudin-1 and occludin, as critical HCV entry factors. We built on this finding by elucidating the human factors required for viral entry into mouse cells, and have now gone on to create the first genetically humanized mouse model supporting the entire HCV lifecycle. This achievement opens unprecedented opportunities to study HCV entry in vivo and to dissect it using the power of mouse genetics. We have also shown proof-of-concept for this animal model in passive and active vaccine strategies. In addition to advancing in vivo models, we are pursuing more biologically relevant cell culture systems for the virus. These include primary adult and fetal hepatocyte cultures and three-dimensional liver organoids. In this competing renewal, we employ our novel in vitro and in vivo systems to further understand the virus-host interactions required for HCV entry. We propose genetic and biophysical approaches to uncover the mechanisms of uptake. Our studies will, for the first time, define the composition and structure of the infecting particle, probe the interplay between virus and host- associated lipid metabolism factors, and map the molecular interaction network active during viral internalization.

Public Health Relevance

Hepatitis C virus, a causative agent of cirrhosis, fibrosis and hepatocellular carcinoma, infects an estimated 2% of the global population. We are studying entry of the virus into cells in the hopes of identifying new targets for inhibitors.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI072613-08
Application #
8658799
Study Section
Virology - B Study Section (VIRB)
Program Officer
Koshy, Rajen
Project Start
2006-12-15
Project End
2017-04-30
Budget Start
2014-05-01
Budget End
2015-04-30
Support Year
8
Fiscal Year
2014
Total Cost
Indirect Cost
Name
Rockefeller University
Department
Microbiology/Immun/Virology
Type
Graduate Schools
DUNS #
City
New York
State
NY
Country
United States
Zip Code
10065
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Swanson, Michael D; Boudreaux, Daniel M; Salmon, Loïc et al. (2015) Engineering a therapeutic lectin by uncoupling mitogenicity from antiviral activity. Cell 163:746-58
Scheel, Troels K H; Kapoor, Amit; Nishiuchi, Eiko et al. (2015) Characterization of nonprimate hepacivirus and construction of a functional molecular clone. Proc Natl Acad Sci U S A 112:2192-7
Scull, Margaret A; Shi, Chao; de Jong, Ype P et al. (2015) Hepatitis C virus infects rhesus macaque hepatocytes and simianized mice. Hepatology 62:57-67
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Saeed, Mohsan; Andreo, Ursula; Chung, Hyo-Young et al. (2015) SEC14L2 enables pan-genotype HCV replication in cell culture. Nature 524:471-5
Ramanan, Vyas; Scull, Margaret A; Sheahan, Timothy P et al. (2014) New Methods in Tissue Engineering: Improved Models for Viral Infection. Annu Rev Virol 1:475-499
Vercauteren, Koen; Van Den Eede, Naomi; Mesalam, Ahmed Atef et al. (2014) Successful anti-scavenger receptor class B type I (SR-BI) monoclonal antibody therapy in humanized mice after challenge with HCV variants with in vitro resistance to SR-BI-targeting agents. Hepatology 60:1508-18
Sheldon, Julie; Beach, Nathan M; Moreno, Elena et al. (2014) Increased replicative fitness can lead to decreased drug sensitivity of hepatitis C virus. J Virol 88:12098-111
Anggakusuma; Colpitts, Che C; Schang, Luis M et al. (2014) Turmeric curcumin inhibits entry of all hepatitis C virus genotypes into human liver cells. Gut 63:1137-49

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