Cyclophilin B Is a Functional Regulator of Hepatitis C Virus RNA Polymerase

Watashi, Koichi; Ishii, Naoto; Hijikata, Makoto; Inoue, Daisuke; Murata, Takayuki; Miyanari, Yusuke; Shimotohno, Kunitada

doi:10.1016/j.molcel.2005.05.014

Cited by 410 publications

(443 citation statements)

References 46 publications

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“…Mutations in NS5B, the viral RNA-dependent RNA polymerase, fit with a model of CyPB as a functional regulator of the viral polymerase. 14 Our study is the first to demonstrate a role of NS5A in the CsA susceptibility of HCV. We show that NS5A has a larger effect than NS5B by separating the NS5A and NS5B mutations.…”

Section: H Epatitis C Virus (Hcv) Chronically Infects Ap-mentioning

confidence: 61%

“…One proposed mechanism for the antiviral effect of CsA is that it inhibits either a CyPB-NS5B interaction or alters the interaction between NS5B and RNA, which is regulated by CyPB. 14 We found 2 mutated NS5B residues in our CsA selection. One of these residues is a proline to threonine change at residue 538.…”

Section: Resultsmentioning

confidence: 85%

“…Cyclophilin B (CyPB) has been shown to bind the HCV polymerase and regulate the binding of RNA template to the polymerase. 14 Another model argues that replication of the HCV replicon depends upon CyPs A, B, and C. 6 Both models were based on experiments that used RNA silencing to target specific host CyPs that are known targets of CsA. We undertook a forward genetic approach in which no assumptions were made about the mechanism of CsA inhibition of HCV.…”

Section: H Epatitis C Virus (Hcv) Chronically Infects Ap-mentioning

confidence: 99%

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Sensitivity of hepatitis C virus to cyclosporine A depends on nonstructural proteins NS5A and NS5B

et al. 2007

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HCV reoccurs after liver transplantation and increases mortality. Cyclosporine, but not tacrolimus, has potent antiviral effects against HCV replication in cell culture. To determine the conditions, if any, under which HCV is susceptible to cyclosporine in vivo, we selected for cyclosporine-resistant mutant HCV in vitro. The resulting mutations were mapped to x-ray crystallographic structures and sequence databases. Mutations selected by cyclosporine were clustered in the nonstructural (NS) proteins NS5A and NS5B. Different sets of mutations in NS5A, paired with the same 2 NS5B mutations, conferred different levels of cyclosporine resistance when engineered back into the HCV replicon. Mutations in NS5B are structurally consistent with a proposed model of regulation of RNA binding by cyclophilin B (CyPB). These mutations also highlight a natural polymorphism between different HCV genotypes that correlates with the variation in response to cyclosporine A (CsA) noted in some clinical trials. Replicons engineered to have mutations in only NS5A (P < 0.0001) or only NS5B (P ‫؍‬ 0.002) suggest that while both NS5A or NS5B variants alter cyclosporine susceptibility, NS5A has the largest effect. Conclusion: Preexisting sequence variation could alter the effect of cyclosporine on HCV in vivo. (HEPATOLOGY

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Section: H Epatitis C Virus (Hcv) Chronically Infects Ap-mentioning

confidence: 61%

Section: Resultsmentioning

confidence: 85%

Section: H Epatitis C Virus (Hcv) Chronically Infects Ap-mentioning

confidence: 99%

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Sensitivity of hepatitis C virus to cyclosporine A depends on nonstructural proteins NS5A and NS5B

et al. 2007

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“…Host Cell Factors Involved in HCV RNA Replication-Apart from the viral proteins and CREs, several host cell factors play an important role for HCV RNA replication based on the criteria that RNA interference-mediated knockdown of each of these cellular genes and/or overexpression of dominant negative mutants decreases viral replication (55)(56)(57). The first one is the human vesicle-associated membrane protein-associated protein A (VAP-A) that was initially identified as an interaction partner of NS5A and NS5B (58).…”

Section: Components Of the Hcv Replication Complexmentioning

confidence: 99%

From Structure to Function: New Insights into Hepatitis C Virus RNA Replication

Appel

Schaller

Pénin

et al. 2006

Journal of Biological Chemistry

177

120

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With an estimated number of about 180 million affected people worldwide and a limited therapy option, infection with the hepatitis C virus (HCV) 2 is an important medical problem. Initial limitations in propagating HCV in cell culture have been overcome step-by-step in the past few years and culminated in the recent development of a system allowing efficient propagation of infectious HCV in tissue culture. Nevertheless, structural and biochemical studies of viral proteins as well as molecular analysis of viral RNA replication are still major challenges. The recent resolution of the three-dimensional structures of some HCV proteins or domains along with elegant reverse genetic and cell biological studies provided first insights into how HCV amplifies its RNA, exploits the cellular machinery, and eventually overcomes innate immune responses. Organization of the Viral GenomeHCV is a positive strand RNA virus that has been classified as the genus Hepacivirus in the Flaviviridae family. The HCV genome is an uncapped, linear molecule with a length of ϳ9600 nucleotides (nt; Fig. 1A). It carries a long open reading frame that is flanked at the 5Ј-and 3Ј-ends by short highly structured non-translated regions (NTRs). The 5Ј-NTR has a length of about 340 nt and contains an internal ribosome entry site (IRES) required for translation of the HCV genome (1). This RNA element binds the 40 S ribosomal subunit in the absence of other translation initiation factors in a way that the initiation codon is placed in the immediate vicinity of the P site (2). Part of the IRES (domain II) overlaps with RNA signals essential for viral replication (Fig. 1A) arguing for a possible role of domain II in regulating a translation-RNA replication switch. The 3Ј-NTR has a tripartite structure composed of an ϳ40-nt-long variable region downstream of the HCV coding sequence, a poly(U/UC) tract of heterogeneous length, and a highly conserved 98-nt-long sequence designated X-tail (Fig. 1A). Genetic studies have shown that a poly(U/UC) tract of at least ϳ25 nt as well as the complete X-tail are required for RNA replication in cell culture and for infectivity of the viral genome in vivo. An additional cis-acting RNA element (CRE) has been identified in the 3Ј-terminal coding region of NS5B (Fig. 1A). This CRE (designated 5BSL3.2) (3) forms a long distance RNA-RNA interaction with SL2 in the X-tail (4), which is indispensable for RNA replication.Expression of the viral proteins from the monocistronic genome is primarily achieved by production of a polyprotein that is proteolytically cleaved into the structural proteins (core, envelope proteins E1 and E2), the hydrophobic peptide p7, and the non-structural (NS) proteins NS2, NS3, NS4A, NS4B, NS5A, and NS5B (Fig. 1A) (1, 5). Processing of the core to p7 region is mediated by host cell signalases, and in the case of the core protein, in addition by signal peptide peptidase. All remaining cleavages are carried out by two viral proteases: the NS2/3 protease mediating cleavage between NS2 and NS3 and the NS3...

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“…Moreover, the replicon system has been exploited for analyses of the effect of cytokines on HCV RNA replication 46,47 and was instrumental in a recent series of elegant studies looking at host proteins involved in HCV RNA replication as well as mechanisms of evasion from innate immune responses. [48][49][50][51][52][53] Since the original reports of functional genotype 1b replicons, replicons for genotype 1a 54 and 2a 55 as well as derivatives expressing easily quantifiable marker enzymes in a separate cistron have been made to facilitate genetic studies as well as drug screening and evaluation. [56][57][58] In addition, full-length replicons and HCV genomes efficiently replicating in tissue culture have been developed, 44,59,60 and the spectrum of permissive host cells has been expanded.…”

Section: The Replicon Systemmentioning

confidence: 99%