Stable expression of hepatitis delta virus antigen in a eukaryotic cell line.
The gene encoding the hepatitis delta virus structural antigen (HDAg) was linked to a neomycin resistance gene in a retrovirus expression vector, and human HepG2 cells were transfected with the recombinant plasmid. A stable cell line was cloned that expressed HDAg in the nuclei of 100% of cells, in a pattern indicating a close relationship with cell nucleoli. Analysis of partially purified recombinant HDAg by HPLC showed an Mr in the range of 7 x 10(5) to 2 x 10(6), which appeared to contain conformation-dependent epitopes, whereas the density of the antigen was 1.19 g/ml by equilibrium centrifugation in caesium chloride, and in rate zonal centrifugation it sedimented with a value of 50S, close to that of particulate hepatitis B virus surface antigen. Immunoblotting demonstrated a single polypeptide with an Mr of 24K which corresponded to the smaller of the two HDAg-specific polypeptides present in infected sera. The recombinant HDAg polypeptide was shown to be a RNA-binding protein with specificity for both genomic and antigenomic species of hepatitis delta virus RNA.
Macnaughton
T B
TB
Division of Medical Virology, Institute of Medical and Veterinary Science, Adelaide, South Australia.
Gowans
E J
EJ
Reinboth
B
B
Jilbert
A R
AR
Burrell
C J
CJ
D000956:Antigens, Viral / Q000032:analysis / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D002466:Cell Nucleolus / Q000276:immunology; D002467:Cell Nucleus / Q000276:immunology / Q000378:metabolism*; D002499:Centrifugation, Density Gradient; D002851:Chromatography, High Pressure Liquid; D003673:Defective Viruses / Q000235:genetics / Q000276:immunology; D005455:Fluorescent Antibody Technique; D005822:Genetic Vectors; D006367:HeLa Cells; D003698:Hepatitis Delta Virus / Q000235:genetics / Q000276:immunology*; D035923:Hepatitis delta Antigens; D006801:Humans; D015151:Immunoblotting; D012313:RNA / Q000378:metabolism; D011994:Recombinant Proteins / Q000032:analysis / Q000096:biosynthesis / Q000378:metabolism; D014162:Transfection
D000956:Antigens, Viral; D035923:Hepatitis delta Antigens; D011994:Recombinant Proteins; C442244:hepatitis delta virus large antigen; D012313:RNA