Hepatitis B viruses (HBV) are a family of DNA viruses that can persistently infect the liver of a variety of animal hosts including humans. There is a close association between chronic HBV infection and hepatocellular carcinoma, though the mechanism of oncogenesis is not understood. Although they have a DNA genome, hepadnaviruses replicate via reverse transcription of an RNA intermediate (RNA pregenome) resulting in a relaxed circular DNA genome. The major goal our laboratory is understanding HBV replication. The HBV capsid is comprised of 120 dimers of the core protein (Cp), held together as a lattice of dimer-dimer associations. We have found that perturbation of the dimer-dimer interface, either chemically or genetically, has profound effects on different processes in HBV replication, such as capsid assembly, DNA synthesis, and virion morphogenesis. The capsid is best known for its structural role as the “vessel” for the HBV genome. In this capacity, its static features are well characterized. It is also clear that the capsid is a dynamic entity that makes multiple contributions to the life cycle of HBV. Reverse transcription/DNA genome synthesis occurs in the interior of the capsid. Capsids containing mature DNA genomes are preferentially secreted as virions over capsids containing incompletely synthesized, immature genomes. As such, the capsid is a dynamic interface between the status of the viral genome and the virus envelope proteins. We hypothesize that: 1) unregulated assembly of capsids leads to the production of empty capsids; 2) productive reverse transcription is dependent on specific conformational changes within the dimer-dimer interface of the capsid; and 3) productive intracellular localization of the capsid is dependent on specific conformations of the capsid. To address these hypotheses, we propose to understand how the dimer-dimer interface plays multiple roles in the HBV life cycle. Little is known about cellular localization and trafficking of capsids during their assembly and genome replication. There is a significant gap in our knowledge of the subcellular locations the capsid uses during the assembly of nascent nucleocapsids, the process of reverse transcription, and morphogenesis of virions. Using live cell imaging we propose to learn about subcellular localization and trafficking of capsids during HBV replication and identify important host processes. In addition, we propose to understand how snow goose hepatitis B virus (SGHBV) supports the secretion of virions containing immature genomes to better understand the mechanistic link between genome maturation and virion envelopment.
Daniel D. Loeb, PhD
6453 Wi Institute Medical Research
1111 Highland Ave
Madison, WI 53705
PhD, Microbiology and Immunology, University of North Carolina at Chapel Hill
molecular biology of the hepatitis B viruses
Tan, Z., Maguire, M. L., Loeb, D. D., and Zlotnick, A. (2013). Genetically Altering the Thermodynamics and Kinetics of Hepatitis B Virus Capsid Assembly Has Profound Effects on Virus Replication in Cell Culture. J. Virol., 87: 3208-3216.
Lentz, T. B., and Loeb, D. D. (2011). Roles of the Envelope Proteins in the Amplification of Covalently Closed Circular DNA and Completion of Synthesis of the Plus-Strand DNA in Hepatitis B Virus. J. Virol., 85: 11916-11927.
Lewellyn, E. B., and Loeb, D. D. (2011). Serine phosphoacceptor sites within the core protein of hepatitis B virus contribute to genome replication pleiotropically. PLoS One, 6(2):e17202.
Lewellyn, E. B., and Loeb, D. D. (2011). The arginine clusters of the carboxy-terminal domain of the core protein of hepatitis B virus make pleiotropic contributions to genome replication. J. Virol., 85: 1298-1309.
Maguire, M. L., and Loeb, D. D. (2010). cis-acting sequences that contribute to synthesis of minus-strand DNA are not conserved between hepadnaviruses. J. Virol., 84: 12824-12831.
Lentz, T. B., and Loeb, D. D. (2010). Development of cell cultures that express hepatitis b virus to high levels and accumulate cccDNA. J. Virol. Methods, 169: 52-60.
Porterfield, J. Z., Dhason, M. S., Loeb, D. D., Nassal, M., Stray, S. J., and Zlotnick, A. (2010). Full-Length hepatitis B virus core protein packages viral and heterologous RNA with similarly high levels of cooperativity. J. Virol., 84: 7174-7184.