The roles of CD81 and glycosaminoglycans in the adsorption and uptake of infectious HCV particles

Abstract

Because appropriate cell-culture systems or small-animal models have been lacking, the early steps in the HCV life cycle have been difficult to study. A cell culture system was developed recently that allows production of infectious HCV. In this study, infectious HCV particles produced in cultured cells were used. To clarify the role of CD81 in HCV attachment and entry, the effect of anti-CD81 antibody was examined. The antibody blocked HCV virion entry but not particle attachment. Only the fraction bound to a heparin affinity column and eluted with 0.3 M NaCl productively infected Huh7 cells, indicating that infectious HCV particles bind to heparin. Both heparin treatment of the virus particles and heparinase treatment of the Huh7 cells reduced virus-cell binding without substantially inhibiting HCV infectivity. Finally, to confirm the role of both heparin sulfate proteoglycan (HSPG) and CD81 in HCV entry, the effects of heparinase I and anti-CD81 antibody were analyzed. No productive RNA replication was detected in the Huh7 cells in the presence of both heparinase I and anti-CD81 antibody. In conclusion, these data suggested that both HSPG and CD81 are important for HCV entry. HSPG may play a role in the initial cell surface binding of infectious HCV particles and CD81 is conceivably correlated with HCV entry after viral attachment.

Cellular models for the screening and development of anti-hepatitis C virus agents

“…It is likely that each step is necessary for the next one and for full coordination of the entry. The participation of glycosaminoglycans (Barth et al, 2006;Morikawa et al, 2007) and LDL receptor (LDLR) (Agnello et al, 1999;Molina et al, 2007) as attachment factors to concentrate HCV at the host cell membrane has been documented and is consistent with the fact that both E1 and E2 are highly glycosylated and with the nature of the HCV LVPs (Andre et al, 2002) (Fig 1, step 1). After initial recruitment at the membrane, HCV particles appear to bind to high affinity entry factors in a sequential manner.…”

Critical Role of Virion-Associated Cholesterol and Sphingolipid in Hepatitis C Virus Infection

“…Plasmid pJFH1, containing full-length cDNA of the JFH-1 isolate, was used to generate HCVcc as described elsewhere (23,33,34,54). pJ6/JFH was obtained from JFH1 by replacement of the 5Ј untranslated region to the p7 region (EcoRI-BclI) of J6.…”

“…It has been shown that CD81 plays an important role in HCV internalization but is not correlated with viral attachment (7,33). An anti-CD81 antibody was used as a negative control for reduced viral attachment.…”

“…Cell-associated virus was quantified by measuring the amount of HCV RNA in the cell lysate by the real-time reverse transcription-PCR method (2,34). Cells were treated with heparinase as previously described (33).…”

Tetraspanin-3 is an organizer of the multi-subunit Nogo-A signaling complex

“…during viral infection of cells -where distinct tetraspanin proteins can have crucial roles in the specific post-attachment binding phase of the virus to the cell surface, after initial viral attachment to heparin sulfate proteoglycans and/or low-density lipoprotein receptorrelated proteins (LRPs) during viral infection of cells (Dahmane et al, 2014;Lindenbach and Rice, 2013;Monk and Partridge, 2012;Morikawa et al, 2007;Nydegger et al, 2006;Scheffer et al, 2013). To gain more insights into the functions of TSPAN3 in Nogo-A-Δ20 binding, we followed TSPAN3 lateral dynamics in the cell membrane at a nanoscopic level with single-particle-tracking.…”