The antiviral protein viperin inhibits hepatitis C virus replication via interaction with nonstructural protein 5A

Abstract

The interferon-stimulated gene viperin has been shown to have antiviral activity against hepatitis C virus (HCV) in the context of the HCV replicon, although the molecular mechanisms responsible are not well understood. Here we demonstrate that viperin plays an integral part in the ability of interferon to limit replication of cell culture derived HCV (JFH-1) that accurately reflects the complete viral life cycle. Using confocal microscopy and Fluorescence Resonance Energy Transfer (FRET) analysis we demonstrate that viperin localizes and interacts with HCV NS5A at the lipid droplet interface. In addition viperin also associates with NS5A and the pro-viral cellular factor, VAP-A at the HCV replication complex. The ability of viperin to limit HCV replication was dependent on residues within the C-terminus as well as an N-terminal amphipathic helix. Removal of the amphipathic helix redirected viperin from the cytosolic face of the ER and the lipid droplet to a homogenous cytoplasmic distribution, coinciding with a loss of antiviral effect. C-terminal viperin mutants still localized to the lipid droplet interface and replication complexes but did not interact with NS5A proteins as determined by FRET analysis. In conclusion we propose that viperin interacts with NS5A and the host factor VAP-A to limit HCV replication at the replication complex. This highlights the complexity of host control of viral replication by interferon stimulated gene expression.

Sequence analysis and subcellular localization of crucian carp Carassius auratus viperin

“…to prevent viral assembly and/or budding [15,17]. Consistently, removal of N-terminal amphipathic helix domain of human viperin results in a homogenous cytoplasmic distribution when transfection of Huh-7 cells, which coincides with a loss of its ability to inhibit HCV replication [17].…”

“…Consistently, removal of N-terminal amphipathic helix domain of human viperin results in a homogenous cytoplasmic distribution when transfection of Huh-7 cells, which coincides with a loss of its ability to inhibit HCV replication [17]. In another paper, sequential deletion of the N-terminal region disrupts ER-association of human viperin in the transfected HEK-293T cells, resulting in its progressive accumulation in cytosol [16].…”

Does Viperin Function as a Radical S-Adenosyl-l-methionine-dependent Enzyme in Regulating Farnesylpyrophosphate Synthase Expression and Activity?

“…The consensus from these studies is that viperin likely exerts its activity through different pathways that depend on the particular virus and that it interacts with multiple cellular and viral proteins in the process (19,27,31,33,34). Previous studies, using a truncated version of viperin, recombinantly produced and purified from E. coli, have demonstrated that the enzyme cleaves SAM to form 5Ј-dA (22), which is the hallmark reaction of radical SAM enzymes.…”

Fish viperin exerts a conserved antiviral function through RLR-triggered IFN signaling pathway

“…In mammals, viperin was characterized as one of IFN-inducible antiviral effectors (Carlton-Smith and Elliott, 2012;Fitzgerald, 2011;Helbig et al, 2011;Jiang et al, 2008;Mattijssen and Pruijn, 2012;Overby et al, 2009;Quraishi et al, 2010), and its expression regulation has been investigated in mammals (Boudinot et al, 2000;Chan et al, 2008;Chin and Cresswell, 2001;Olofsson et al, 2005;Saitoh et al, 2011;Severa et al, 2006;Stirnweiss et al, 2010). However, this information in fish is still unclear.…”

“…Viperin is capable to inhibit the release of influenza virus by perturbing lipid rafts (Wang et al, 2007) or to limit the replication of hepatitis C virus by interacting with nonstructural protein 5A (Helbig et al, 2011). Overexpression of viperin promotes TLR7-and TLR9-mediated production of type I IFN in mouse plasmacytoid dendritic cells (Saitoh et al, 2011).…”