Identification of a Residue in Hepatitis C Virus E2 Glycoprotein That Determines Scavenger Receptor BI and CD81 Receptor Dependency and Sensitivity to Neutralizing Antibodies

Abstract

Hepatitis C virus (HCV) infection is dependent on at least three coreceptors: CD81, scavenger receptor BI (SR-BI), and claudin-1. The mechanism of how these molecules coordinate HCV entry is unknown. In this study we demonstrate that a cell culture-adapted JFH-1 mutant, with an amino acid change in E2 at position 451 (G451R), has a reduced dependency on SR-BI. This altered receptor dependency is accompanied by an increased sensitivity to neutralization by soluble CD81 and enhanced binding of recombinant E2 to cell surface-expressed and soluble CD81. Fractionation of HCV by density gradient centrifugation allows the analysis of particle-lipoprotein associations. The cell culture-adapted mutation alters the relationship between particle density and infectivity, with the peak infectivity occurring at higher density than the parental virus. No association was observed between particle density and SR-BI or CD81 coreceptor dependence. JFH-1 G451R is highly sensitive to neutralization by gp-specific antibodies, suggesting increased epitope exposure at the virion surface. Finally, an association was observed between JFH-1 particle density and sensitivity to neutralizing antibodies (NAbs), suggesting that lipoprotein association reduces the sensitivity of particles to NAbs. In summary, mutation of E2 at position 451 alters the relationship between particle density and infectivity, disrupts coreceptor dependence, and increases virion sensitivity to receptor mimics and NAbs. Our data suggest that a balanced interplay between HCV particles, lipoprotein components, and viral receptors allows the evasion of host immune responses.Hepatitis C virus (HCV), the sole member of the Hepacivirus genus within the Flaviviridae, poses a global health burden, with an estimated 170 million infected individuals (according to the WHO). The majority of patients suffer a chronic infection that is associated with a progressive liver disease (1). HCV has a short positive-sense RNA genome encoding three structural (core protein, E1, and E2) glycoproteins (gps) and seven nonstructural proteins (p7 and NS2 to NS5) (40). The E1 and E2 gps interact with cell surface receptors to facilitate particle entry via low-pH and clathrin-dependent endocytosis (9,15,29,47,71). The recent discovery that the JFH-1 strain of HCV can replicate and assemble infectious particles in cultured cells (HCVcc) has allowed investigation into the viral life cycle for the first time since its identification almost 20 years ago (41,75,79).Early studies with truncated soluble HCV E2 (sE2) identified interactions with the tetraspanin CD81 and scavenger receptor class B type I (SR-BI) (56, 62). The recent availability of HCVcc and HCV pseudoparticles (HCVpp) provided the tools to validate receptor candidates. HCV entry is thought to require at least three cellular receptors: CD81, SR-BI, and the tight junction protein claudin-1 (reviewed in references 21 and 74). Other candidate components include glycosaminoglycans (5, 6, 51), low-density lipoprotein receptor (49,77), and the C...

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

“…6A), but this observation did not reach statistical significance. The IC 50 value for oxLDL against the G451R point mutant 36 was similar to WT (Fig. 6B).…”

“…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.…”

“…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.…”

Apolipoprotein E Mediates Evasion From Hepatitis C Virus Neutralizing Antibodies

“…TRL association, measured by buoyant density distribution, could shield HCV glycoproteins from nAbs. 12 Two distinct models have been proposed regarding HCVÀapoE interaction and lipoviral particles formation: a 1-particle model presenting the virion inextricably fused to the lipoprotein with apoE on its surface, and a 2-particle model where HCV glycoproteins interact with apoE directly, which may mediate lipoprotein attachment. 30 In addition, the envelope glycoprotein, and the transmembrane domain of E2 specifically is critical for = -specific antibody and captured complexes were analyzed by immunoblotting using anti-E2 antibodies.…”

“…ApoCI may be involved in viral fusion, 11 and apoAI can affect HCV replication and production. Although association with TRL and LDL has been hypothesized to contribute to viral evasion, 7,12 the role of specific apolipoproteins in HCV persistence is unknown. Given that apoE is a host protein incorporated into the HCV particle and is required for virion production, we sought to determine its functional role in viral evasion from host nAbs.…”