Extracellular maturation of secreted hepatitis C virus particles by incorporation of Apoliporotein E enhances infectivity

Aim of the study: Hepatitis C virus (HCV) efficiently evades humoral immune responses to establish a chronic infection. Virus particles circulate in complex with host-derived lipoproteins, thus facilitating escape from antibodies. The exchangeable apolipoprotein E (ApoE) is essential for intracellular HCV assembly as well as for HCV cell entry.

To gain insights into key principles of HCV assembly and immune evasion, we explored if ApoE released from non-infected human liver cells interacts with secreted HCV particles and modulates their properties.

Materials & methods: ApoE was expressed in human hepatoma cells and cell-free culture fluid enriched with ApoE was incubated with secreted HCV particles at ApoE doses comparable to typical levels in human sera. A direct interaction between cell culture derived and patient-derived HCV and exogenously added ApoE was quantified using an HA-tagged ApoE variant and immunoprecipitation. Infectivity of ApoE-conditioned HCV particles was determined using luciferase reporter virus and limiting dilution infection assays.

Results: Extracellular ApoE increased infectivity of secreted HCV particles from all genotypes as well as of HCV particles produced in primary human hepatocytes. Incubation of HCV particles with HA-tagged ApoE at physiological doses resulted in co-precitation of HCV core protein with HA-ApoE indicating that ApoE is loaded onto secreted HCV particles after release from infected cells. Infection of SR-BI knockout cells and heparinase treated cells as well as competition assays with SR-BI-targeting molecules and heparin indicated that ApoE enriched particles have increased entry efficiency due to improved virus attachment to cell surface proteoglycans and not -as shown for HDL-mediated enhancement- due to an interplay with SR-BI. Finally, extracellular loading of ApoE onto HCV particles decreased neutralization efficiency of E2-protein targeting monoclonal antibodies.

Conclusions: Our data provide evidence that secreted ApoE is incorporated into released HCV particles. Thus, secreted particles undergo maturation and remodelling to enhance infectivity and thereby facilitate evasion from antibodies.