Expression of hepatitis C virus envelope glycoproteins by herpes simplex virus type 1-based amplicon vectors.
Herpes simplex virus type 1 (HSV-1)-based amplicon vectors expressing hepatitis C virus (HCV) E1 and E2 glycoproteins were investigated. HSV-1 amplicon vectors carrying the E1E2p7- or E2p7-coding sequences of HCV type 1a under the control of the HSV-1 IE4 (alpha22/alpha47) promoter were constructed. Studies of infected HepG2, WRL 68 or Vero cells indicated that HSV-1-based amplicon vectors express high levels of HCV glycoproteins that are processed correctly. Immunofluorescence microscopy combined with immunoprecipitation and endoglycosidase treatment of cells infected with the HSV-1-based vectors expressing E1 and E2 showed that the two glycoproteins were retained in the endoplasmic reticulum and had the expected glycosylation patterns. Furthermore, although most of the E1 and E2 proteins formed disulfide-linked aggregates, significant amounts of monomeric forms of the two proteins were detected by SDS-PAGE under non-reducing conditions, suggesting the presence of non-covalently associated E1 and E2. Similar results were produced by a replication-competent recombinant HSV-1 vector expressing HCV E1 and E2. These results indicated that HSV-1-based amplicon vectors represent a useful expression system for the study of HCV glycoproteins.
Tsitoura
Eliza
E
Molecular Virology Laboratory, Hellenic Pasteur Institute, 127 Vas, Sofias Avenue, Athens, Greece1.
Lucas
Michaela
M
Molecular Virology Laboratory, Hellenic Pasteur Institute, 127 Vas, Sofias Avenue, Athens, Greece1.
Revol-Guyot
Valerie
V
Centre de Genetique Moleculaire et Cellulaire, UMR 5534 CNRS, Universite Claude Bernard Lyon I, 69622 Villeurbanne Cedex, France2.
Epstein
Alberto L
AL
Centre de Genetique Moleculaire et Cellulaire, UMR 5534 CNRS, Universite Claude Bernard Lyon I, 69622 Villeurbanne Cedex, France2.
Manservigi
Roberto
R
Section of Microbiology, University of Ferrara, Via Luigi Borsari 46, Ferrara 1-44100, Italy3.
Mavromara
Penelope
P
Molecular Virology Laboratory, Hellenic Pasteur Institute, 127 Vas, Sofias Avenue, Athens, Greece1.
D000818:Animals; D002460:Cell Line; D002522:Chlorocebus aethiops / Q000821:virology; D006224:Cricetinae; D004220:Disulfides / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism; D005784:Gene Amplification / Q000235:genetics*; D015870:Gene Expression; D005822:Genetic Vectors / Q000235:genetics*; D006023:Glycoproteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D006026:Glycoside Hydrolases / Q000378:metabolism; D006031:Glycosylation; D016174:Hepacivirus / Q000235:genetics*; D018259:Herpesvirus 1, Human / Q000235:genetics* / Q000502:physiology; D006801:Humans; D008856:Microscopy, Fluorescence; D011233:Precipitin Tests; D011485:Protein Binding; D014709:Vero Cells; D014759:Viral Envelope Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D014779:Virus Replication
D004220:Disulfides; C087035:E1 protein, Hepatitis C virus; D006023:Glycoproteins; D014759:Viral Envelope Proteins; C091015:glycoprotein E2, Hepatitis C virus; D006026:Glycoside Hydrolases