FORMATION AND INTRACELLULAR-LOCALIZATION OF HEPATITIS-C VIRUS ENVELOPE GLYCOPROTEIN COMPLEXES EXPRESSED BY RECOMBINANT VACCINIA AND SINDBISVIRUSES

Hepatitis C virus (HCV) encodes two putative virion glycoproteins (E1 and E2) which are released from the polyprotein by signal peptidase cl eavage. In this report, we have characterized the complexes formed bet ween E1 and E2 (called E1E2) for two different HCV strains (H and BK) and studied their intracellular localization. Vaccinia virus and Sindb is virus vectors were used to express the HCV structural proteins in t hree different cell lines (HepG2, BHK-21, and PK-15). The kinetics of association between E1 and E2, as studied by pulse-chase analysis and coprecipitation of E2 with an anti-E1 monoclonal antibody, indicated t hat formation of stable E1E2 complexes is slow. The times required for half-maximal association between E1 and E2 were 60 to 85 min for the H strain and more than 165 min for the BK strain. In the presence of n onionic detergents, two forms of E1E2 complexes were detected. The pre dominant form was a heterodimer of E1 and E2 stabilized by noncovalent interactions. A minor fraction consisted of heterogeneous disulfide-l inked aggregates, which most likely represent misfolded complexes. Pos ttranslational processing and localization of the HCV glycoproteins we re examined by acquisition of endoglycosidase H resistance, subcellula r fractionation, immunofluorescence, cell surface immunostaining, and immunoelectron microscopy. HCV glycoproteins containing complex N-link ed glycans were not observed, and the proteins were not detected at th e cell surface. Rather, the proteins localized predominantly to the en doplasmic reticular network, suggesting that some mechanism exists for their retention in this compartment.