A major determinant of cyclophilin dependence and cyclosporine susceptibility of Hepatitis C virus identified by a genetic approach

• Verfasst von: Yang, Feng [VerfasserIn]
• Verfasst von: Robotham, Jason M. [VerfasserIn]
• Verfasst von: Grise, Henry [VerfasserIn]
• Verfasst von: Frausto, Stephen [VerfasserIn]
• Verfasst von: Madan Renes, Vanesa [VerfasserIn]
• Verfasst von: Zayas López, Margarita Laura [VerfasserIn]
• Verfasst von: Bartenschlager, Ralf [VerfasserIn]
• Verfasst von: Robinson, Margaret [VerfasserIn]
• Verfasst von: Greenstein, Andrew E. [VerfasserIn]
• Verfasst von: Nag, Anita [VerfasserIn]
• Verfasst von: Logan, Timothy M. [VerfasserIn]
• Verfasst von: Bienkiewicz, Ewa [VerfasserIn]
• Verfasst von: Tang, Hengli [VerfasserIn]
• Titel: A major determinant of cyclophilin dependence and cyclosporine susceptibility of Hepatitis C virus identified by a genetic approach
• Verf.angabe: Feng Yang, Jason M. Robotham, Henry Grise, Stephen Frausto, Vanesa Madan, Margarita Zayas, Ralf Bartenschlager, Margaret Robinson, Andrew E. Greenstein, Anita Nag, Timothy M. Logan, Ewa Bienkiewicz, Hengli Tang
• E-Jahr: 2010
• Jahr: September 2010
• Umfang: 16 S.
• Illustrationen: Illustrationen
• Fussnoten: Gesehen am 22.05.2024
• Titel Quelle: Enthalten in: Public Library of SciencePLoS pathogens
• Ort Quelle: Lawrence, Kan. : PLoS, 2005
• Jahr Quelle: 2010
• Band/Heft Quelle: 6(2010), 9 vom: Sept., Artikel-ID e1001118, Seite 1-16
• ISSN Quelle: 1553-7374
• DOI: doi:10.1371/journal.ppat.1001118
• Datenträger: Online-Ressource
• Sprache: eng
• Sach-SW: Circular dichroism spectroscopy
• Sach-SW: Genomics
• Sach-SW: Hepatitis C virus
• Sach-SW: Luciferase assay
• Sach-SW: Microbial mutation
• Sach-SW: Point mutation
• Sach-SW: Proline
• Sach-SW: Viral replication
• K10plus-PPN: 1889577022

Abstract

Since the advent of genome-wide small interfering RNA screening, large numbers of cellular cofactors important for viral infection have been discovered at a rapid pace, but the viral targets and the mechanism of action for many of these cofactors remain undefined. One such cofactor is cyclophilin A (CyPA), upon which hepatitis C virus (HCV) replication critically depends. Here we report a new genetic selection scheme that identified a major viral determinant of HCV's dependence on CyPA and susceptibility to cyclosporine A. We selected mutant viruses that were able to infect CyPA-knockdown cells which were refractory to infection by wild-type HCV produced in cell culture. Five independent selections revealed related mutations in a single dipeptide motif (D316 and Y317) located in a proline-rich region of NS5A domain II, which has been implicated in CyPA binding. Engineering the mutations into wild-type HCV fully recapitulated the CyPA-independent and CsA-resistant phenotype and four putative proline substrates of CyPA were mapped to the vicinity of the DY motif. Circular dichroism analysis of wild-type and mutant NS5A peptides indicated that the D316E/Y317N mutations (DEYN) induced a conformational change at a major CyPA-binding site. Furthermore, nuclear magnetic resonance experiments suggested that NS5A with DEYN mutations adopts a more extended, functional conformation in the putative CyPA substrate site in domain II. Finally, the importance of this major CsA-sensitivity determinant was confirmed in additional genotypes (GT) other than GT 2a. This study describes a new genetic approach to identifying viral targets of cellular cofactors and identifies a major regulator of HCV's susceptibility to CsA and its derivatives that are currently in clinical trials.