Hepatitis C Virus Hypervariable Region 1 Modulates Receptor Interactions, Conceals the CD81 Binding Site, and Protects Conserved Neutralizing Epitopes

Abstract

The variability of the hepatitis C virus (HCV), which likely contributes to immune escape, is most pronounced in hypervariable region 1 (HVR1) of viral envelope protein 2. This domain is the target for neutralizing antibodies, and its deletion attenuates replication in vivo. Here we characterized the relevance of HVR1 for virus replication in vitro using cell culture-derived HCV. We show that HVR1 is dispensable for RNA replication. However, viruses lacking HVR1 (⌬HVR1) are less infectious, and separation by density gradients revealed that the population of ⌬HVR1 virions comprises fewer particles with low density. Strikingly, ⌬HVR1 particles with intermediate density (1.12 g/ml) are as infectious as wild-type virions, while those with low density (1.02 to 1.08 g/ml) are poorly infectious, despite quantities of RNA and core similar to those in wild-type particles. Moreover, ⌬HVR1 particles exhibited impaired fusion, a defect that was partially restored by an E1 mutation (I347L), which also rescues infectivity and which was selected during long-term culture. Finally, ⌬HVR1 particles were no longer neutralized by SR-B1-specific immunoglobulins but were more prone to neutralization and precipitation by soluble CD81, E2-specific monoclonal antibodies, and patient sera. These results suggest that HVR1 influences the biophysical properties of released viruses and that this domain is particularly important for infectivity of low-density particles. Moreover, they indicate that HVR1 obstructs the viral CD81 binding site and conserved neutralizing epitopes. These functions likely optimize virus replication, facilitate immune escape, and thus foster establishment and maintenance of a chronic infection.Hepatitis C virus (HCV) is a single-stranded positive-sense RNA virus of the family Flaviviridae that has infected an estimated 130 million people worldwide (1). Acute HCV infection is mostly asymptomatic; however, virus persistence can lead to severe liver disease, and within 20 years ca. 20% of chronically infected adults develop cirrhosis (46). In fact, morbidity associated with chronic HCV infection is the most common indication for orthotopic liver transplantation (7). The mechanisms that permit the virus to establish chronic infection in ca. 55 to 85% of cases (24) despite vigorous immune responses are incompletely understood.A number of studies have highlighted the pivotal role of strong, multispecific, and sustained T-cell responses for control of HCV infection (summarized in reference 53). Although resolution of acute HCV infection can occur in the absence of antibodies (47), mounting evidence indicates that neutralizing antibodies also contribute to protective immunity (summarized in reference 62). Nevertheless, HCV often successfully evades cellular and humoral immune pressure likely at least in part via the constant generation of variants created by an error-prone RNA replication machinery. In line with this notion, a high degree of HCV sequence evolution is associated with chronic disease, while a comparatively st...

Characterization of Hepatitis C Virus Particle Subpopulations Reveals Multiple Usage of the Scavenger Receptor BI for Entry Steps

“…In agreement with recent reports (9,14,15,19), HCVcc particles had more than 65% of their infectivity in fractions of densities of 1.08 -1.13 g/ml, i.e. slightly shifted toward low densities as compared with the peak of HCV RNA level (1.10 -1.16 g/ml) (Fig.…”

Characterization of the inhibition of hepatitis C virus entry byIn vitro-generated and patient-derived oxidized low-density lipoprotein

“…The IC 50 value of oxLDL against two HCV mutants with reduced-but not absent-SR-BI dependency, DHVR1 35 and G451R, 36 did not differ significantly from WT. However, in the case of DHVR1, numerically, the IC 50 value was markedly (18-fold) higher, compared to WT.…”

“…The IC 50 value for oxLDL inhibition of the DHVR1 variant, where the entire hypervariable region 1 (HVR1) has been deleted from the E2 glycoprotein, 35 was approximately 18-fold higher, compared to unmodified HCV (Fig. 6A), but this observation did not reach statistical significance.…”

“…35 Interestingly, DHVR1 and G451R share a decreased sensitivity to anti-SR-BI sera and increased sensitivity to soluble CD81. Of note, their major difference-G451R, but not DHVR1, directly bind SR-BI-has been shown only with sE2, but not complete virions.…”

“…Of note, their major difference-G451R, but not DHVR1, directly bind SR-BI-has been shown only with sE2, but not complete virions. 35,36 Given that SR-BI is thought to act at more than one step of the HCV-entry process, 21,22 further work will clearly be needed to define the exact effects of the different viral variants and inhibitors of the virus-SR-BI interactions. In addition, it is interesting to speculate how oxidative modification of apolipoproteins, such as ApoE and ApoC1, currently known to be associated with HCV, would affect HCV infectivity.…”