Human RPB5, a subunit shared by eukaryotic nuclear RNA polymerases, binds human hepatitis B virus X protein and may play a role in X transactivation.
The X gene of human hepatitis B virus encodes the polypeptide HBx which transactivates viral and host genes through a variety of cis-acting enhancer elements present in RNA polymerases I, II and III promoters. To better understand the mechanism of X transactivation, we cloned cDNAs of proteins that bind HBx. Here we demonstrate that one of these cDNAs is a full-length cDNA of human RPB5, a subunit shared by RNA polymerases. The HBx transactivation domain and the central region of human RPB5 were necessary for the specific binding of the two proteins as shown by: (i) in vitro assays using deletion mutants of fusion proteins; (ii) in vivo assays which detect associated proteins by co-immunoprecipitation of the non-fused proteins from transfected HepG2 cells. Over-expressed HBx seemed to associate with assembled forms of endogenous human RPB5 in HBx-transfected cells, since the endogenous RPB5 co-immunoprecipitated with HBx. The HBx binding region of human RPB5 by itself stimulated chloramphenicol acetyltransferase activities from several different reporters having X-responsive element(s). Our results support the idea that the interaction of HBx and human RPB5 can facilitate HBx transactivation and that human RPB5 has a domain which can communicate with transcriptional regulators.
Cheong
J H
JH
Department of Molecular Biology, Kanazawa University, Japan.
Yi
M
M
Lin
Y
Y
Murakami
S
S
D000595:Amino Acid Sequence; D015153:Blotting, Western; D002478:Cells, Cultured; D003001:Cloning, Molecular; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D005057:Eukaryotic Cells; D017930:Genes, Reporter; D006801:Humans; D008099:Liver / Q000166:cytology; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism*; D011233:Precipitin Tests; D011485:Protein Binding; D011487:Protein Conformation; D015534:Trans-Activators / Q000378:metabolism*; D015533:Transcriptional Activation; D014162:Transfection; D054334:Viral Regulatory and Accessory Proteins
D009687:Nuclear Proteins; D015534:Trans-Activators; D054334:Viral Regulatory and Accessory Proteins; C057628:hepatitis B virus X protein; D012321:DNA-Directed RNA Polymerases; C517511:POLR2E protein, human