title: Selection of a hepatitis C virus with altered entry factor requirements reveals a genetic interaction between the E1 glycoprotein and claudins.
abstract: Hepatitis C virus (HCV) cell entry is a complex, multistep process requiring numerous host cell factors, including the tight junction protein claudin-1 (CLDN1). It is not known whether CLDN1 and the HCV glycoproteins physically interact. Therefore, the focus of this work was to study genetic interactions between CLDN1 and HCV. We used CRISPR technology to generate CLDN1 knockout (KO) Huh-7.5 cells, which could not be infected by genotype 2a Jc1 HCV unless CLDN1 expression was restored. Passage of Jc1-transfected CLDN1 KO cells resulted in the selection of a virus that could infect these cells. This virus encoded a single mutation, H316N (numbered relative to the HCV polyprotein), in the E1 glycoprotein. Whereas Jc1 H316N efficiently infected cells lacking CLDN1, such infection was blocked by an antibody targeting CLDN6, another member of the claudin family that is expressed in these cells. Furthermore, HuH6 cells, which express CLDN6, but not CLDN1, were infectable only with the mutant virus. Thus, this mutant virus adapted to the loss of CLDN1 by developing the capacity to utilize other CLDNs. Indeed, CLDN1/CLDN6 double-KO Huh-7.5 cells supported infection by the mutant virus only when CLDN1, CLDN6, or CLDN9 was expressed. Finally, this phenotype was not genotype dependent, given that the H316N mutation rendered a Japanese fulminant hepatitis 1 chimeric HCV genome encoding the genotype 5a glycoproteins able to utilize CLDN6 for host cell entry.
  
  
  CONCLUSIONS
  These data demonstrate plasticity of HCV virus-host interactions, where a previously CLDN1-dependent virus was capable of evolving to use CLDN6. They also reveal a role for E1 in determining entry factor usage and imply a direct, physical interaction between E1 and CLDNs.
authors.affiliation: Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York, NY.
authors.forename: Sharon E
authors.initials: SE
authors.lastname: Hopcraft
authors.affiliation: Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York, NY.
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Evans
mesh_terms: D002478:Cells, Cultured; D062445:Claudin-1 / Q000235:genetics*; D016174:Hepacivirus / Q000235:genetics*; D006801:Humans; D014759:Viral Envelope Proteins / Q000235:genetics*; D053586:Virus Internalization
chemical_list: D062445:Claudin-1; C087035:E1 protein, Hepatitis C virus; D014759:Viral Envelope Proteins
references.citation: Hepatology. 2014 Mar;59(3):776-88
references.pmid: 24038151
references.citation: J Virol. 2008 Apr;82(7):3555-60
references.pmid: 18234789
references.citation: J Virol. 2013 Sep;87(18):10405-10
references.pmid: 23864633
references.citation: PLoS Pathog. 2013 Mar;9(3):e1003244
references.pmid: 23555257
references.citation: Methods. 2013 Feb;59(2):233-48
references.pmid: 23009812
references.citation: Science. 2013 Feb 15;339(6121):823-6
references.pmid: 23287722
references.citation: Science. 2013 Feb 15;339(6121):819-23
references.pmid: 23287718
references.citation: Science. 2012 Aug 17;337(6096):816-21
references.pmid: 22745249
references.citation: PLoS One. 2012;7(4):e36029
references.pmid: 22558311
references.citation: Curr Opin Virol. 2012 Feb;2(1):14-9
references.pmid: 22440961
references.citation: Cell. 2000 Apr 14;101(2):173-85
references.pmid: 10786833
references.citation: Mol Ther. 2001 Apr;3(4):438-48
references.pmid: 11319904
references.citation: J Virol. 2002 Dec;76(24):13001-14
references.pmid: 12438626
references.citation: J Biol Chem. 2003 Apr 4;278(14):11979-84
references.pmid: 12554735
references.citation: Biochimie. 2003 Mar-Apr;85(3-4):295-301
references.pmid: 12770768
references.citation: Semin Liver Dis. 1995 Feb;15(1):41-63
references.pmid: 7597443
references.citation: Science. 2005 Jul 22;309(5734):623-6
references.pmid: 15947137
references.citation: Nature. 2005 Aug 18;436(7053):973-8
references.pmid: 16107838
references.citation: Proc Natl Acad Sci U S A. 2006 May 9;103(19):7408-13
references.pmid: 16651538
references.citation: J Biol Chem. 2008 Mar 28;283(13):8643-53
references.pmid: 18211898
references.citation: Hepatology. 2009 Feb;49(2):364-77
references.pmid: 19148942
references.citation: Nature. 2009 Feb 12;457(7231):882-6
references.pmid: 19182773
references.citation: J Virol. 2010 Jan;84(2):964-75
references.pmid: 19889758
references.citation: Hepatology. 2010 Apr;51(4):1144-57
references.pmid: 20069648
references.citation: PLoS Pathog. 2010;6:e1000978
references.pmid: 20617177
references.citation: Gastroenterology. 2010 Sep;139(3):953-64, 964.e1-4
references.pmid: 20685314
references.citation: J Virol. 2010 Nov;84(22):11696-708
references.pmid: 20844048
references.citation: J Virol. 2011 Jul;85(14):7005-19
references.pmid: 21543495
references.citation: Protein Eng Des Sel. 2011 Sep;24(9):679-89
references.pmid: 21810920
references.citation: Hepatology. 2014 Jan;59(1):24-34
references.pmid: 23775920
references.citation: J Virol. 2007 Nov;81(22):12465-71
references.pmid: 17804490
references.citation: Nature. 2007 Apr 12;446(7137):801-5
references.pmid: 17325668
references.citation: Hepatology. 2015 Jan;61(1):77-87
references.pmid: 25069599
references.citation: Nat Biotechnol. 2015 May;33(5):549-54
references.pmid: 25798937