Regulatory Role of Phospholipids in Hepatitis C Virus Replication and Protein Function

Abstract

Positive-strand RNA viruses such as hepatitis C virus (HCV) hijack key factors of lipid metabolism of infected cells and extensively modify intracellular membranes to support the viral lifecycle. While lipid metabolism plays key roles in viral particle assembly and maturation, viral RNA synthesis is closely linked to the remodeling of intracellular membranes. The formation of viral replication factories requires a number of interactions between virus proteins and host factors including lipids. The structure-function relationship of those proteins is influenced by their lipid environments and lipids that selectively modulate protein function. Here, we review our current understanding on the roles of phospholipids in HCV replication and of lipid-protein interactions in the structure-function relationship of the NS5A protein. NS5A is a key factor in membrane remodeling in HCV-infected cells and is known to recruit phosphatidylinositol 4-kinase III alpha to generate phosphatidylinositol 4-phosphate at the sites of replication. The dynamic interplay between lipids and viral proteins within intracellular membranes is likely key towards understanding basic mechanisms in the pathobiology of virus diseases, the mode of action of specific antiviral agents and related drug resistance mechanisms.

Keywords: NS5A; RNA virus; hepatitis C virus; interactions; membrane remodeling; phospholipid; viral replication.

Figure 1

Schematic representation of the HCV life cycle. Steps of the HCV genome replication cycle involving interactions between nonstructural viral proteins and host phospholipids are shown in red. Interaction of HCV lipo-viro-particles (LVPs) with hepatocytes is mediated by attachment receptors, followed by tight binding to internalization receptors, which are required for HCV endocytosis. After fusion of the enveloping HCV-LVP membrane with the endosome, the HCV +RNA genome is released into the cytoplasm and translated at the ER. HCV proteins interact with the membrane of the ER and induce membrane remodeling and formation of replication organelles (comprising double-membrane vesicles, DMVs). Viral assembly occurs at detergent-resistant membranes (DRMs) of the ER in close proximity to DMVs and cytoplasmic lipid droplets (LDs). The maturing LVPs are finally released via the secretory pathway.

Figure 2

Hypothetical models of the NS5A interaction with membranes. (A) NS5A model of a full-length dimer comprising N-terminal membrane attachment portions—amphipathic α-helices (AHs) based on Protein Data Bank (PDB) entries 1ZH1 for NS5A D1 dimer and 1R7G for AH. Domains (D) 2 and 3 of NS5A are intrinsically unfolded and likely interact with multiple host factors. The N-terminal AH specifically interacts with phosphatidylinositol 4,5-bisphosphate (PIP2) via a pincer motif (residues as stick models). Domain 1 of NS5A interacts with the lipid kinase phosphatidylinositol 4-kinase III alpha (PI4KIIIα) via PFIS motif; (B) Binding of NS5A at the replication complex (negative membrane curvature) or lipid droplets (LD) (positive membrane curvature). NS5A-mediated subversion of lipid metabolism at the replication complex by sequential recruitment and activation of PI4KIIIα and oxysterol binding protein (OSBP) with accumulation of PI4P and subsequent exchange for cholesterol by OSBP or for ceramide by ceramide transfer protein (CERT).