The duck hepatitis B virus polymerase is activated by its RNA packaging signal, epsilon.
The epsilon stem-loop at the 5' end of the pregenomic RNA of the hepatitis B viruses is both the primary element of the RNA packaging signal and the origin of reverse transcription. We have previously presented evidence for a third essential role for epsilon, that of an essential cofactor in the maturation of the viral polymerase (J. E. Tavis and D. Ganem, J. Virol. 70:5741-5750, 1996). In this case, binding of epsilon to the polymerase is proposed to induce a physical alteration to the polymerase that is needed for it to develop enzymatic activity. Three lines of evidence employing duck hepatitis B virus supporting this hypothesis are presented here. First, an unusual DNA polymerase activity employing exogenous RNAs (the trans reaction) that was originally discovered with recombinant duck hepatitis B virus polymerase expressed in Saccharomyces cerevisiae yeasts was shown to be an authentic property of the viral polymerase. The trans reaction was found to be template-dependent reverse transcription of the exogenous RNA. The trans reaction occurred independently of the hepadnavirus protein-priming mechanism, yet it was still strongly stimulated by epsilon. This directly demonstrates a role for epsilon in activation of the polymerase. Second, the reverse transcriptase domain of the polymerase was shown to be physically altered following binding to epsilon, as would be expected if the alteration was required for maturation of the polymerase to an enzymatically active form. Finally, analysis of 15 mutations throughout the duck hepatitis B virus polymerase demonstrated that the epsilon-dependent alteration to the polymerase was a prerequisite for DNA priming, reverse transcription, and the trans reaction.
Tavis
J E
JE
Department of Molecular Microbiology and Immunology, St. Louis University School of Medicine, St. Louis, Missouri 63104, USA. tavisje@wpogate.slu.edu
Massey
B
B
Gong
Y
Y
D000818:Animals; D002645:Chickens; D004789:Enzyme Activation; D015611:Hepatitis B Virus, Duck / Q000201:enzymology*; D012367:RNA, Viral / Q000502:physiology*; D012194:RNA-Directed DNA Polymerase / Q000378:metabolism*; D014407:Tumor Cells, Cultured; D019065:Virus Assembly
D012367:RNA, Viral; D012194:RNA-Directed DNA Polymerase