Initiation of replication of the human hepatitis delta virus genome from cloned DNA: role of delta antigen.
Beginning with three partial cDNA clones of the RNA genome of human hepatitis delta virus (HDV), we assembled the complete 1,679-base sequence on a single molecule and then inserted a trimer of this into plasmid pSLV, a simian virus 40-based eucaryotic expression vector. This construct was used to transfect both monkey kidney (COS7) and human hepatocellular carcinoma (HuH7) cell lines. In this way we obtained replication of the HDV RNA genome and the appearance, in the nucleoli, of the delta antigen, the only known virus-coded protein. This proved both that the HDV genome could replicate in nonliver as well as liver cells and that there was no requirement for the presence of hepatitis B virus sequences or proteins. When the pSVL construct was made with a dimer of an HDV sequence with a 2-base-pair deletion in the open reading frame, genome replication was reduced at least 40-fold. However, when we cotransfected with a plasmid that expressed the correct delta antigen, the mutated dimer achieved a level of genome replication comparable to that of the nonmutated sequence. We thus conclude that the delta antigen can act in trans and is essential for replication of the HDV genome.
Kuo
M Y
MY
Fox Chase Cancer Center, Philadelphia, Pennsylvania 19111.
Chao
M
M
Taylor
J
J
D000956:Antigens, Viral / Q000502:physiology*; D015153:Blotting, Western; D002460:Cell Line; D003001:Cloning, Molecular; D005455:Fluorescent Antibody Technique; D005814:Genes, Viral; D003698:Hepatitis Delta Virus / Q000235:genetics / Q000254:growth & development*; D035923:Hepatitis delta Antigens; D006801:Humans; D066298:In Vitro Techniques; D012367:RNA, Viral / Q000096:biosynthesis* / Q000235:genetics; D014162:Transfection; D014779:Virus Replication
D000956:Antigens, Viral; D035923:Hepatitis delta Antigens; D012367:RNA, Viral; C442244:hepatitis delta virus large antigen