Global and local envelope protein dynamics of hepatitis C virus determine broad antibody sensitivity

Broad antibody sensitivity differences of hepatitis C virus (HCV) isolates and their ability to persist in the presence of neutralizing antibodies (NAbs) remain poorly understood. Here, we show that polymorphisms within glycoprotein E2, including hypervariable region 1 (HVR1) and antigenic site 412 (AS412), broadly affect NAb sensitivity by shifting global envelope protein conformation dynamics between theoretical “closed,” neutralization-resistant and “open,” neutralization-sensitive states. The conformational space of AS412 was skewed toward β-hairpin–like conformations in closed states, which also depended on HVR1, assigning function to these enigmatic E2 regions. Scavenger receptor class B, type I entry dependency of HCV was associated with NAb resistance and correlated perfectly with decreased virus propensity to interact with HCV co-receptor CD81, indicating that decreased NAb sensitivity resulted in a more complex entry pathway. This link between global E1/E2 states and functionally distinct AS412 conformations has important implications for targeting AS412 in rational HCV vaccine designs.

Download Full-text

Hepatitis C Virus Escape Studies of Human Antibody AR3A Reveal a High Barrier to Resistance and Novel Insights on Viral Antibody Evasion Mechanisms

Journal of Virology ◽

10.1128/jvi.01909-18 ◽

2018 ◽

Vol 93 (4) ◽

Cited By ~ 5

Author(s):

Rodrigo Velázquez-Moctezuma ◽

Andrea Galli ◽

Mansun Law ◽

Jens Bukh ◽

Jannick Prentoe

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Vaccine Development ◽

Scavenger Receptor ◽

Hypervariable Region ◽

Type I ◽

Low Level ◽

Class B ◽

High Level

ABSTRACTYearly, ∼2 million people become hepatitis C virus (HCV) infected, resulting in an elevated lifetime risk for severe liver-related chronic illnesses. Characterizing epitopes of broadly neutralizing antibodies (NAbs), such as AR3A, is critical to guide vaccine development. Previously identified alanine substitutions that can reduce AR3A binding to expressed H77 envelope were introduced into chimeric cell culture-infectious HCV recombinants (HCVcc) H77(core-NS2)/JFH1. Substitutions G523A, G530A, and D535A greatly reduced fitness, and S424A, P525A, and N540A, although viable, conferred only low-level AR3A resistance. Using highly NAb-sensitive hypervariable region 1 (HVR1)-deleted HCVcc, H77/JFH1ΔHVR1and J6(core-NS2)/JFH1ΔHVR1, we previously reported a low barrier to developing AR5A NAb resistance substitutions. Here, we cultured Huh7.5 cells infected with H77/JFH1, H77/JFH1ΔHVR1, or J6/JFH1ΔHVR1with AR3A. We identified the resistance envelope substitutions M345T in H77/JFH1, L438S and F442Y in H77/JFH1ΔHVR1, and D431G in J6/JFH1ΔHVR1. M345T increased infectivity and conferred low-level AR3A resistance to H77/JFH1 but not H77/JFH1ΔHVR1. L438S and F442Y conferred high-level AR3A resistance to H77/JFH1ΔHVR1but abrogated the infectivity of H77/JFH1. D431G conferred AR3A resistance to J6/JFH1ΔHVR1but not J6/JFH1. This was possibly because D431G conferred broadly increased neutralization sensitivity to J6/JFH1D431Gbut not J6/JFH1ΔHVR1/D431Gwhile decreasing scavenger receptor class B type I coreceptor dependency. Common substitutions at positions 431 and 442 did not confer high-level resistance in other genotype 2a recombinants [JFH1 or T9(core-NS2)/JFH1]. Although the data indicate that AR3A has a high barrier to resistance, our approach permitted identification of low-level resistance substitutions. Also, the HVR1-dependent effects on AR3A resistance substitutions suggest a complex role of HVR1 in virus escape and receptor usage, with important implications for HCV vaccine development.IMPORTANCEHepatitis C virus (HCV) is a leading cause of liver-related mortality, and limited treatment accessibility makes vaccine development a high priority. The vaccine-relevant cross-genotype-reactive antibody AR3A has shown high potency, but the ability of the virus to rapidly escape by mutating the AR3A epitope (barrier to resistance) remains unexplored. Here, we succeeded in inducing only low-level AR3A resistance, indicating a higher barrier to resistance than what we have previously reported for AR5A. Furthermore, we identify AR3A resistance substitutions that have hypervariable region 1 (HVR1)-dependent effects on HCV viability and on broad neutralization sensitivity. One of these substitutions increased envelope breathing and decreased scavenger receptor class B type I HCV coreceptor dependency, both in an HVR1-dependent fashion. Thus, we identify novel AR3A-specific resistance substitutions and the role of HVR1 in protecting HCV from AR3-targeting antibodies. These viral escape mechanisms should be taken into consideration in future HCV vaccine development.

Download Full-text

Scavenger Receptor Class B Type I Mediates Cell Entry of Hepatitis C Virus

Journal of International Medical Research ◽

10.1177/147323000803600620 ◽

2008 ◽

Vol 36 (6) ◽

pp. 1319-1325 ◽

Cited By ~ 9

Author(s):

ZS Jia ◽

DW Du ◽

YF Lei ◽

X Wei ◽

W Yin ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Cho Cells ◽

Hepg2 Cells ◽

Scavenger Receptor ◽

Chinese Hamster ◽

Hcv Infection ◽

Type I ◽

Scavenger Receptor Class ◽

Class B

This study assessed the functional role of human scavenger receptor class B type I (SR-BI) as a putative hepatitis C virus (HCV) receptor using Chinese hamster ovary (CHO) cells transfected with human SR-BI (CHO–huSR-BI). The expression of SR-BI by primary Tupaia hepatocytes (PTHs), human hepatocarcinoma cell line (HepG2) cells, untransfected CHO cells and CHO–huSR-BI cells was analysed by Western blotting. Receptor competition assays showed that anti-SR-BI antibodies that block the binding of soluble envelope glycoprotein E2 could prevent HCV infection. Pre-incubation of CHO–huSR-BI and HepG2 cells with anti-SR-BI antibodies resulted in marked inhibition of E2 binding. After incubation with HCV RNA-positive serum from a patient with chronic HCV infection, however, HCV infection could not be detected in CHO–huSR-BI cells, but was detected in PTHs. These results demonstrate that, whilst SR-BI represents an important cell surface molecule for HCV infection, the presence of SR-BI alone is insufficient for HCV entry.

Download Full-text

56 Scavenger receptor class B type i mediates uptake and cross-presentation of hepatitis C virus by human dendritic cells

Journal of Hepatology ◽

10.1016/s0168-8278(06)80057-7 ◽

2006 ◽

Vol 44 ◽

pp. S26

Author(s):

H. Barth ◽

E.K. Schnober ◽

C. Neumann-Haefelin ◽

R. Thimme ◽

H.E. Blum ◽

...

Keyword(s):

Dendritic Cells ◽

Hepatitis C Virus ◽

Hepatitis C ◽

Scavenger Receptor ◽

Type I ◽

Cross Presentation ◽

Scavenger Receptor Class ◽

Class B ◽

Human Dendritic Cells

Download Full-text

The postbinding activity of scavenger receptor class B type I mediates initiation of hepatitis C virus infection and viral dissemination

Hepatology ◽

10.1002/hep.26097 ◽

2012 ◽

Vol 57 (2) ◽

pp. 492-504 ◽

Cited By ~ 58

Author(s):

Muhammad N. Zahid ◽

Marine Turek ◽

Fei Xiao ◽

Viet Loan Dao Thi ◽

Maryse Guérin ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Virus Infection ◽

Scavenger Receptor ◽

Type I ◽

Hepatitis C Virus Infection ◽

Viral Dissemination ◽

Scavenger Receptor Class ◽

Class B

Download Full-text

Role of Scavenger Receptor Class B Type I in Hepatitis C Virus Entry: Kinetics and Molecular Determinants

Journal of Virology ◽

10.1128/jvi.02199-08 ◽

2009 ◽

Vol 84 (1) ◽

pp. 34-43 ◽

Cited By ~ 105

Author(s):

Maria Teresa Catanese ◽

Helenia Ansuini ◽

Rita Graziani ◽

Thierry Huby ◽

Martine Moreau ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Virus Entry ◽

Scavenger Receptor ◽

Specific Protein ◽

Early Event ◽

Type I ◽

Direct Role ◽

Scavenger Receptor Class ◽

Class B

ABSTRACT Scavenger receptor class B type I (SR-BI) is an essential receptor for hepatitis C virus (HCV) and a cell surface high-density-lipoprotein (HDL) receptor. The mechanism of SR-BI-mediated HCV entry, however, is not clearly understood, and the specific protein determinants required for the recognition of the virus envelope are not known. HCV infection is strictly linked to lipoprotein metabolism, and HCV virions may initially interact with SR-BI through associated lipoproteins before subsequent direct interactions of the viral glycoproteins with SR-BI occur. The kinetics of inhibition of cell culture-derived HCV (HCVcc) infection with an anti-SR-BI monoclonal antibody imply that the recognition of SR-BI by HCV is an early event of the infection process. Swapping and single-substitution mutants between mouse and human SR-BI sequences showed reduced binding to the recombinant soluble E2 (sE2) envelope glycoprotein, thus suggesting that the SR-BI interaction with the HCV envelope is likely to involve species-specific protein elements. Most importantly, SR-BI mutants defective for sE2 binding, although retaining wild-type activity for receptor oligomerization and binding to the physiological ligand HDL, were impaired in their ability to fully restore HCVcc infectivity when transduced into an SR-BI-knocked-down Huh-7.5 cell line. These findings suggest a specific and direct role for the identified residues in binding HCV and mediating virus entry. Moreover, the observation that different regions of SR-BI are involved in HCV and HDL binding supports the hypothesis that new therapeutic strategies aimed at interfering with virus/SR-BI recognition are feasible.

Download Full-text

Hepatitis C virus entry: potential receptors and their biological functions

Journal of General Virology ◽

10.1099/vir.0.81646-0 ◽

2006 ◽

Vol 87 (5) ◽

pp. 1075-1084 ◽

Cited By ~ 124

Author(s):

Laurence Cocquerel ◽

Cécile Voisset ◽

Jean Dubuisson

Keyword(s):

Cell Culture ◽

Hepatitis C Virus ◽

Hepatitis C ◽

Low Density Lipoprotein ◽

Heparan Sulphate ◽

Scavenger Receptor ◽

Density Lipoprotein ◽

Type I ◽

Major Step ◽

Class B

Several cellular molecules have been identified as putative receptors for Hepatitis C virus (HCV): CD81 tetraspanin, scavenger receptor class B type I (SR-BI), mannose-binding lectins DC-SIGN and L-SIGN, low-density lipoprotein receptor, heparan sulphate proteoglycans and the asialoglycoprotein receptor. Due to difficulties in propagating HCV in cell culture, most of these molecules have been identified by analysing their interaction with a soluble, truncated form of HCV glycoprotein E2. A recent major step in investigating HCV entry was the development of pseudoparticles (HCVpp), consisting of unmodified HCV envelope glycoproteins assembled onto retroviral core particles. This system has allowed the investigation of the role of candidate receptors in the early steps of the HCV life cycle and the data obtained can now be confirmed with the help of a newly developed cell-culture system that allows efficient amplification of HCV (HCVcc). Interestingly, CD81 and SR-BI have been shown to play direct roles in HCVpp and/or HCVcc entry. However, co-expression of CD81 and SR-BI in non-hepatic cell lines does not lead to HCVpp entry, indicating that other molecule(s), expressed only in hepatic cells, are necessary for HCV entry. In this review, the molecules that have been proposed as potential HCV receptors are described and the experimental data indicating that CD81 and SR-BI are potentially involved in HCV entry are presented.

Download Full-text

An Interplay between Hypervariable Region 1 of the Hepatitis C Virus E2 Glycoprotein, the Scavenger Receptor BI, and High-Density Lipoprotein Promotes both Enhancement of Infection and Protection against Neutralizing Antibodies

Journal of Virology ◽

10.1128/jvi.79.13.8217-8229.2005 ◽

2005 ◽

Vol 79 (13) ◽

pp. 8217-8229 ◽

Cited By ~ 217

Author(s):

Birke Bartosch ◽

Géraldine Verney ◽

Marlène Dreux ◽

Peggy Donot ◽

Yoann Morice ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Neutralizing Antibodies ◽

Scavenger Receptor ◽

Hypervariable Region ◽

Density Lipoprotein ◽

Cell Entry ◽

Scavenger Receptor Bi ◽

E2 Glycoprotein ◽

Hypervariable Region 1

ABSTRACT Hepatitis C virus (HCV) circulates in the bloodstream in different forms, including complexes with immunoglobulins and/or lipoproteins. To address the significance of such associations, we produced or treated HCV pseudoparticles (HCVpp), a valid model of HCV cell entry and its inhibition, with naïve or patient-derived sera. We demonstrate that infection of hepatocarcinoma cells by HCVpp is increased more than 10-fold by human serum factors, of which high-density lipoprotein (HDL) is a major component. Infection enhancement requires scavenger receptor BI, a molecule known to mediate HDL uptake into cells as well as HCVpp entry, and involves conserved amino acid positions in hypervariable region 1 (HVR1) of the E2 glycoprotein. Additionally, we show that the interaction with human serum or HDL, but not with low-density lipoprotein, leads to the protection of HCVpp from neutralizing antibodies, including monoclonal antibodies and antibodies present in patient sera. Finally, the deletion or mutation of HVR1 in HCVpp abolishes infection enhancement and leads to increased sensitivity to neutralizing antibodies/sera compared to that of parental HCVpp. Altogether, these results assign to HVR1 new roles which are complementary in helping HCV to survive within its host. Besides immune escape by mutation, HRV1 can mediate the enhancement of cell entry and the protection of virions from neutralizing antibodies. By preserving a balance between these functions, HVR1 may be essential for the viral persistence of HCV.

Download Full-text