Abstract
Amphipathic α-helices of exchangeable apolipoproteins have shown to play crucial roles in the formation of infectious hepatitis C virus (HCV) particles through the interaction with viral particles. Among the Flaviviridae members, pestivirus and flavivirus possess a viral structural protein Erns or a non-structural protein 1 (NS1) as secretory glycoproteins, respectively, while Hepacivirus including HCV has no secretory glycoprotein. In case of pestivirus replication, the C-terminal long amphipathic α-helices of Erns are important for anchoring to viral membrane. Here we show that host-derived apolipoproteins play functional roles similar to those of virally encoded Erns and NS1 in the formation of infectious particles. We examined whether Erns and NS1 could compensate for the role of apolipoproteins in particle formation of HCV in apolipoprotein B (ApoB) and ApoE double-knockout Huh7 (BE-KO), and non-hepatic 293T cells. We found that exogenous expression of either Erns or NS1 rescued infectious particle formation of HCV in the BE-KO and 293T cells. In addition, expression of apolipoproteins or NS1 partially rescued the production of infectious pestivirus particles in cells upon electroporation with an Erns-deleted non-infectious RNA. As with exchangeable apolipoproteins, the C-terminal amphipathic α-helices of Erns play the functional roles in the formation of infectious HCV or pestivirus particles. These results strongly suggest that the host- and virus-derived secretory glycoproteins have overlapping roles in the viral life cycle of Flaviviridae, especially in the maturation of infectious particles, while Erns and NS1 also participate in replication complex formation and viral entry, respectively. Considering the abundant hepatic expression and liver-specific propagation of these apolipoproteins, HCV might have evolved to utilize them in the formation of infectious particles through deletion of a secretory viral glycoprotein gene.
Highlights
The family Flaviviridae consists of 4 genera, namely Flavivirus, Pestivirus, Pegivirus, and Hepacivirus
To examine whether Erns of pestivirus can compensate for the role of apolipoproteins in the formation of infectious hepatitis C virus (HCV) particles, apolipoprotein E (ApoE) or HA-tagged Erns (HA-Erns) was lentivirally expressed in BE-KO cells and intracellular HCV RNA and infectious viral titers in the supernatants were determined at 72-h postinfection with HCV (Fig 1A–1D)
Expression of HA-Erns derived from various pestiviruses, including bovine viral diarrhea virus (BVDV), classical swine fever virus (CSFV), and border disease virus (BDV), in BE-KO cells led to a significant recovery of the formation of HCV infectious particles to nearly the level in the control cells, while HCV RNA replication was not affected by the expression of these proteins
Summary
The family Flaviviridae consists of 4 genera, Flavivirus, Pestivirus, Pegivirus, and Hepacivirus. Flaviviridae viruses commonly possess a single-stranded positive-sense RNA encoding 2–4 structural proteins and 7–8 non-structural proteins [1]. The host range and tissue tropism of these proteins are strikingly different. The genus Flavivirus is composed of more than 50 species with a wide host range from reptiles to mammals; these mosquito- or tick-borne viruses include dengue virus (DENV), Japanese encephalitis virus (JEV), and tick-borne encephalitis virus (TBEV) [2]. The genus Pestivirus comprises economically important pathogens of eventoed ungulate animals, including classical swine fever virus (CSFV), bovine viral diarrhea virus (BVDV), and border disease virus (BDV), which cause major losses in livestock farming [3]. In the case of hepatitis C virus (HCV) from the genus Hepacivirus, the species-specificity is restricted to humans and chimpanzees [4]
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